Researchers at The University of Nottingham have developed a unique technology that will allow scientists to look at microscopic activity within the body's chemical messenger system for the very first time, live as it happens.
The cutting edge laser technology has helped to attract .3 million pounds from the MRC (Medical Research Council) for a five-year project that will offer a new insight into the tiny world of activity taking place within single cells and could contribute to the design of new drugs to treat human diseases such as asthma and arthritis with fewer side effects.
The team, involving scientists from the University's Schools of Biomedical Science (Professor Steve Hill and Dr Steve Briddon) and Pharmacy (Dr Barrie Kellam), is concentrating on a type of specialised docking site (receptor) on the surface of a cell that recognises and responds to a natural chemical within the body called adenosine.
These A3-adenosine receptors work within the body by binding with proteins to cause a response within cells and are found in very tiny and highly specialised area of a cell membrane called microdomains. Microdomains contain a collection of different molecules that are involved in telling the cell how to respond to drugs or hormones.
It is believed that these receptors play an important role in inflammation within the body and knowing more about how they operate could inform the future development of anti-inflammatory drugs that target just those receptors in the relevant microdomain of the cell, without influencing the same receptors in other areas of the cell. However, scientists have never before been able to look in detail at their activity within these tiny microscopic regions of a living cell.
The Nottingham researchers have solved this problem by creating novel drug molecules which have fluorescent labels attached. Using a cutting edge laser technology called fluorescence correlation spectroscopy, the fluorescent drug molecules can be detected as they glow under the laser beam of a highly sensitive microscope. This allows their binding to the receptor to be followed for the first time in real time at the single molecule level.
Leading the project, Professor Steve Hill in the School of Biomedical Sciences said: "These microdomains are so tiny you could fit five million on them on a full stop. There are 10,000 receptors on each cell, and we are able to follow how single drug molecules bind to individual receptors in these specialised microdomains.
"What makes this single molecule laser technique unique is that we are looking at them in real time on a living cell. Other techniques that investigate how drugs bind to their receptors require many millions of cells to get a big enough signal and this normally involves destroying the cells in the process"
The researchers will be using donated blood as a source of A3-receptors in specialised human blood cells (neutrophils) that have important roles during inflammation.
Different types of adenosine receptors are found all over the body and can exist in different areas of the cell membrane and have different properties. Scientists hope that eventually the new technology could also be used to unlock the secrets of the role they play in a whole host of human diseases.
The fluorescent molecules developed as part of the research project will also be useful in drug screening programmes and The University of Nottingham will be making these fluorescent drugs available to the wider scientific community through its links with its spin-out company CellAura Technologies Ltd.
The University of Nottingham is ranked in the UK's Top 10 and the World's Top 70 universities by the Shanghai Jiao Tong (SJTU) and Times Higher (THES) World University Rankings.
It provides innovative and top quality teaching, undertakes world-changing research, and attracts talented staff and students from 150 nations. Described by The Times as Britain's "only truly global university", it has invested continuously in award-winning campuses in the United Kingdom, China and Malaysia. Twice since 2003 its research and teaching academics have won Nobel Prizes. The University has won the Queen's Award for Enterprise in both 2006 (International Trade) and 2007 (Innovation - School of Pharmacy).
Its students are much in demand from 'blue-chip' employers. Winners of Students in Free Enterprise for four years in succession, and current holder of UK Graduate of the Year, they are accomplished artists, scientists, engineers, entrepreneurs, innovators and fundraisers. Nottingham graduates consistently excel in business, the media, the arts and sport. Undergraduate and postgraduate degree completion rates are amongst the highest in the United Kingdom.
Video footage can be found at wirksworthii.nottingham.ac.uk/Podcasts/files/rmg/public/science/laser.mp4
Source: Emma Thorne
University of Nottingham
понедельник, 30 мая 2011 г.
Vaporized Viral Vector Shows Promise In Anti-Cancer Gene Therapy
A new lung cancer therapy employing a vaporized viral vector to deliver a cancer-inhibiting molecule directly to lung tissue shows early promise in mouse trials, according to researchers at the Ministry of Education, Science and Technology in Korea.
Gene therapy is an area of great promise, but delivery mechanisms, which have included intravenous injection and intratracheal instillation, have proven problematic for effective delivery of genetic therapy to lung tissues.
"Aerosol delivery targets the lungs specifically and represents a noninvasive alternative for targeting genes to the lung," wrote Myung-Haing Cho, D.V.M., Ph.D., professor at Seoul National University and principal investigator of the study. "The delivery of genes via aerosol holds promise for the treatment of a broad spectrum of pulmonary disorders and offers numerous advantages over more invasive modes of delivery."
The results of Dr. Cho's promising research will be published in the June 15 issue of the American Journal of Respiratory and Critical Care Medicine.
Lung cancer is the most common cause of cancer deaths worldwide, killing more people each year than breast, prostate and colon cancers combined. It costs the U.S. alone more than $9 billion a year, according to the Centers for Disease Control and Prevention. Most available therapies - surgery, radiation and chemotherapy - offer transient relief at best and are typically ineffective in advanced stages of the disease. For this reason, novel therapies for lung cancer are of great interest.
Dr. Cho and colleagues targeted the Akt signaling pathway, which has been shown to be an important regulator of cell proliferation and cancer progression. A recent report found that 90 percent of non-small cell lung carcinomas were associated with the activation of the Akt signaling pathway. They chose a lentiviral vector, derived from a retrovirus and known for its ability to infect nondividing cells and effect persistent genetic changes. They transfected the lentiviral vector with a negative regulator of Akt signaling, carboxyl-terminal modulator protein (CTMP), which would theoretically inhibit Akt signaling, thus suppressing cancer cell proliferation and tumor growth.
Using a mouse model of lung cancer, the researchers designed a double-control study, exposing one-third of the mice to the aerosolized CTMP vector, one-third to the vector alone and one third were untreated.
"In this study, our main purpose was to determine if viral delivery of CTMP can provide useful tool for designing lung tumor treatment," said Dr. Cho. "We would like to demonstrate that CTMP can suppress lung tumor mass in the lungs and lentivirus may act as an effective carrier of CTMP."
After four weeks of twice-weekly treatments, the researchers found exactly that: both pathological and histological examination of the mice revealed that CTMP delivery suppressed lung tumor mass in the lungs of the mice. Furthermore, the number and volume of tumors were significantly decreased in CTMP-treated mice.
The researchers also found that CTMP increased apoptosis, inhibited angiogenesis and suppressed production of several proteins, such as cyclin D1, CDK4 and CDK2, which are important in cancer cell growth.
"Our results demonstrated that lentivirus-mediated CTMP overexpression suppressed Akt activity and inhibited tumor progression," wrote Dr. Cho. "Repeated aerosol gene delivery may provide an effective noninvasive model of gene delivery and understanding the role of CTMP in the multistage lung tumorigenesis may be essential in developing effective therapeutics for lung cancer."
Source:
Keely Savoie
American Thoracic Society
Gene therapy is an area of great promise, but delivery mechanisms, which have included intravenous injection and intratracheal instillation, have proven problematic for effective delivery of genetic therapy to lung tissues.
"Aerosol delivery targets the lungs specifically and represents a noninvasive alternative for targeting genes to the lung," wrote Myung-Haing Cho, D.V.M., Ph.D., professor at Seoul National University and principal investigator of the study. "The delivery of genes via aerosol holds promise for the treatment of a broad spectrum of pulmonary disorders and offers numerous advantages over more invasive modes of delivery."
The results of Dr. Cho's promising research will be published in the June 15 issue of the American Journal of Respiratory and Critical Care Medicine.
Lung cancer is the most common cause of cancer deaths worldwide, killing more people each year than breast, prostate and colon cancers combined. It costs the U.S. alone more than $9 billion a year, according to the Centers for Disease Control and Prevention. Most available therapies - surgery, radiation and chemotherapy - offer transient relief at best and are typically ineffective in advanced stages of the disease. For this reason, novel therapies for lung cancer are of great interest.
Dr. Cho and colleagues targeted the Akt signaling pathway, which has been shown to be an important regulator of cell proliferation and cancer progression. A recent report found that 90 percent of non-small cell lung carcinomas were associated with the activation of the Akt signaling pathway. They chose a lentiviral vector, derived from a retrovirus and known for its ability to infect nondividing cells and effect persistent genetic changes. They transfected the lentiviral vector with a negative regulator of Akt signaling, carboxyl-terminal modulator protein (CTMP), which would theoretically inhibit Akt signaling, thus suppressing cancer cell proliferation and tumor growth.
Using a mouse model of lung cancer, the researchers designed a double-control study, exposing one-third of the mice to the aerosolized CTMP vector, one-third to the vector alone and one third were untreated.
"In this study, our main purpose was to determine if viral delivery of CTMP can provide useful tool for designing lung tumor treatment," said Dr. Cho. "We would like to demonstrate that CTMP can suppress lung tumor mass in the lungs and lentivirus may act as an effective carrier of CTMP."
After four weeks of twice-weekly treatments, the researchers found exactly that: both pathological and histological examination of the mice revealed that CTMP delivery suppressed lung tumor mass in the lungs of the mice. Furthermore, the number and volume of tumors were significantly decreased in CTMP-treated mice.
The researchers also found that CTMP increased apoptosis, inhibited angiogenesis and suppressed production of several proteins, such as cyclin D1, CDK4 and CDK2, which are important in cancer cell growth.
"Our results demonstrated that lentivirus-mediated CTMP overexpression suppressed Akt activity and inhibited tumor progression," wrote Dr. Cho. "Repeated aerosol gene delivery may provide an effective noninvasive model of gene delivery and understanding the role of CTMP in the multistage lung tumorigenesis may be essential in developing effective therapeutics for lung cancer."
Source:
Keely Savoie
American Thoracic Society
DNA Repair Enzyme Probed By Researchers
Researchers have taken the first steps toward understanding how an enzyme repairs DNA.
Enzymes called helicases play a key role in human health, according to Maria Spies, a University of Illinois biochemistry professor.
"DNA helicases act as critical components in many molecular machineries orchestrating DNA repair in the cell." she said. "Multiple diseases including cancer and aging are associated with malfunctions in these enzymes."
Spies' laboratory undertook a recent study of an enzyme, called Rad3, which defines a group of DNA helicases characterized by a unique structural domain containing iron. The findings appear in the Journal of Biological Chemistry.
Helicases are a special category of molecular motors that modify DNA (deoxyribonucleic acid, the fundamental building block of genes and chromosomes). They do so by moving along strands of DNA, much the same way cars move on roads, using an energy-packed molecule, adenosine triphosphate (ATP) as a fuel source.
Their primary function is to unzip double-stranded DNA, allowing replication and repair of the strands.
DNA is a fragile molecule that undergoes dramatic changes when exposed to radiation, ultraviolet light, toxic chemicals or byproducts of normal cellular processes. DNA damage, if not repaired in time, may lead to mutations, cancer or cell death. Many helicases in the Rad3 family are key players in the cell's elaborate machinery to prevent and repair such damage. Mutations in the human members of this helicase family impede DNA repair and may contribute to breast cancer, Fanconi Anemia and Xeroderma pigmentosum.
The researchers studied the archaeal version of Rad3. Archaea are microbes whose DNA repair systems are closely related to those of human cells.
"(The archaeal Rad3) is a very good representative of a unique family of structurally related DNA repair helicases, all of which have the same motor core and share an unprecedented (for helicases) structural feature - an accessory domain stabilized by an iron-sulfur cluster," Spies said.
Working with archaea has the advantage of allowing the researchers to increase the amount available protein and also permits easy genetic manipulation.
Like other helicases, Rad3 is composed of a chain of amino acids. It also contains an ancient prosthetic group called an iron-sulfur cluster, an assembly of four iron and four sulfur atoms incorporated into the protein structure through interaction with four cysteine residues of the amino acid chain.
"DNA helicases, which belong to the Rad3 family, have an auxiliary domain inserted within a conserved motor core. The structure of this domain is stabilized by an iron-sulfur cluster, whose integrity seems to be essential for proper function of these enzymes in DNA repair," Spies said. By mutating the cysteine ligands to the cluster, the researchers probed its role in the molecular mechanism of Rad3 enzymes. Some of these mutations uncoupled DNA translocation and ATP hydrolysis, meaning that the engine of the protein could still use the ATP fuel but was no longer capable of moving along the DNA.
This analysis also revealed that the integrity of the cluster and the iron-containing domain is crucial for recognition of specific DNA structures believed to be physiological targets for this helicase. "On making these mutations, the helicase no longer behaves like it's supposed to," said graduate student Robert Pugh, lead author on the study. "The cluster is still there but the environment around it is somehow changing."
This research was performed in collaboration with Isaac Caan's group from Animal Sciences whose lab is engaged in the study of nucleotide metabolism in archaea.
Source: Diana Yates
University of Illinois at Urbana-Champaign
Enzymes called helicases play a key role in human health, according to Maria Spies, a University of Illinois biochemistry professor.
"DNA helicases act as critical components in many molecular machineries orchestrating DNA repair in the cell." she said. "Multiple diseases including cancer and aging are associated with malfunctions in these enzymes."
Spies' laboratory undertook a recent study of an enzyme, called Rad3, which defines a group of DNA helicases characterized by a unique structural domain containing iron. The findings appear in the Journal of Biological Chemistry.
Helicases are a special category of molecular motors that modify DNA (deoxyribonucleic acid, the fundamental building block of genes and chromosomes). They do so by moving along strands of DNA, much the same way cars move on roads, using an energy-packed molecule, adenosine triphosphate (ATP) as a fuel source.
Their primary function is to unzip double-stranded DNA, allowing replication and repair of the strands.
DNA is a fragile molecule that undergoes dramatic changes when exposed to radiation, ultraviolet light, toxic chemicals or byproducts of normal cellular processes. DNA damage, if not repaired in time, may lead to mutations, cancer or cell death. Many helicases in the Rad3 family are key players in the cell's elaborate machinery to prevent and repair such damage. Mutations in the human members of this helicase family impede DNA repair and may contribute to breast cancer, Fanconi Anemia and Xeroderma pigmentosum.
The researchers studied the archaeal version of Rad3. Archaea are microbes whose DNA repair systems are closely related to those of human cells.
"(The archaeal Rad3) is a very good representative of a unique family of structurally related DNA repair helicases, all of which have the same motor core and share an unprecedented (for helicases) structural feature - an accessory domain stabilized by an iron-sulfur cluster," Spies said.
Working with archaea has the advantage of allowing the researchers to increase the amount available protein and also permits easy genetic manipulation.
Like other helicases, Rad3 is composed of a chain of amino acids. It also contains an ancient prosthetic group called an iron-sulfur cluster, an assembly of four iron and four sulfur atoms incorporated into the protein structure through interaction with four cysteine residues of the amino acid chain.
"DNA helicases, which belong to the Rad3 family, have an auxiliary domain inserted within a conserved motor core. The structure of this domain is stabilized by an iron-sulfur cluster, whose integrity seems to be essential for proper function of these enzymes in DNA repair," Spies said. By mutating the cysteine ligands to the cluster, the researchers probed its role in the molecular mechanism of Rad3 enzymes. Some of these mutations uncoupled DNA translocation and ATP hydrolysis, meaning that the engine of the protein could still use the ATP fuel but was no longer capable of moving along the DNA.
This analysis also revealed that the integrity of the cluster and the iron-containing domain is crucial for recognition of specific DNA structures believed to be physiological targets for this helicase. "On making these mutations, the helicase no longer behaves like it's supposed to," said graduate student Robert Pugh, lead author on the study. "The cluster is still there but the environment around it is somehow changing."
This research was performed in collaboration with Isaac Caan's group from Animal Sciences whose lab is engaged in the study of nucleotide metabolism in archaea.
Source: Diana Yates
University of Illinois at Urbana-Champaign
Super Sticky Barnacle Glue Cures Like Blood Clots
Barnacles are a big problem for boats. Adhering to the undersides of vessels, carpets of the crustaceans can increase fuel consumption by as much as 25%. Ship owners would love to know how to stop these hitchhikers gluing on, but before you can learn how to disrupt an adhesive, you have to understand the curing process. Curious about many aspects of the crustacean's lifestyle, Dan Rittschof from Duke University decided to find out how barnacle adhesive polymerizes. 'The process must be related to something because glue isn't de novo,' says Rittschof, so he wondered what else coagulates under water and came up with two answers: blood and semen. With a colossal body of blood clotting literature to draw on, Rittschof decided to follow his evolutionarily inspired theory to see whether barnacle glue polymerization is really an extreme example of scab formation and publishes his results on 16 October 2009 in the Journal of Experimental Biology at jeb.biologists.
Rittschof teamed up with Gary Dickinson and the first thing that Dickinson had to do was work out how to collect the unpolymerised glue and keep it fluid. Building on 30 years of Rittschof's experience and Beatriz Orihuela's expertise at growing and reattaching barnacles, Dickinson learned to gently lift polymerised glue away from the pores that secrete the adhesive and quickly collect the minute drops as they oozed from the shell. Working in the cold room to slow the polymerization process, Dickinson had only 5 minutes before each sample polymerized and the glue set solid.
Next the team had to convince themselves that the viscous secretion was glue and not some other body fluid. Dickinson found that the fluid polymerised rapidly and was packed full of protein, just like barnacle glue. Next Dickinson teamed up with Kathy Wahl to use atomic force microscopy to compare the molecular structures of naturally cured glue (from stuck-down barnacles) and his polymerized samples. The two samples were virtually indistinguishable and Dickinson could clearly see tangled webs of fibres in his glue drops, similar to the tangled fibres in blood clots.
But this evidence was still far from proving that barnacle glue cures by the same process as blood clots. Dickinson and Rittschof needed to identify the key proteins that polymerize the cement. Knowing that blood clots are formed when enzymes, known as trypsin-like serine proteases, trigger a cascade of events that culminates in the formation of the long fibres found in blood clots, Dickinson and Rittschoff began searching for the protease in the unpolymerised glue. Separating the glue's components on a gel, Dickinson could see the tell-tale pattern of bands that suggested that a trypsin-like serine protease was present. And when Dickinson added an inhibitor, to inactivate the protease, to a fresh sample of glue, the sample didn't set.
Having convinced themselves that the glue contained a trypsin-like serine protease, the team began to search for other blood-clot-like proteins in the barnacle's secretions. Teaming up with Joseph Bonaventura and Irving Vega, Dickinson chopped each glue component into minute fragments, measured their sizes with mass spectrometry and matched the fragment pattern to known protein sequences. Amazingly, one of the glue proteins was remarkably similar to human factor XIII: a human blood clotting factor that cross-links clot fibres to form a scab. In fact, some regions of the human and barnacle proteins were completely identical. Dickinson and Rittschof had stumbled across the crucial protein that cross-links the glue fibres to cure barnacle cement and it was very similar to factor XIII, an essential human blood-clotting factor.
Rittschof admits that he is shocked that his hypothesis stands up to the tests. 'It seems likely that barnacle glue polymerization is a specialized form of wound healing,' he says and suspects that many other marine animals that rely on glue to get a grip may use the same polymerization mechanism.
REFERENCE: Dickinson, G. H., Vega, I. E., Wahl, K. J., Orihuela, B., Beyley, V., Rodriguez, E. N., Everett, R. K., Bonaventura, J. and Rittschof, D. (2009). Barnacle cement: a polymerization model based on evolutionary concepts. J. Exp. Biol. 212, 3499-3510
Source:
Kathryn Knight
The Company of Biologists
Rittschof teamed up with Gary Dickinson and the first thing that Dickinson had to do was work out how to collect the unpolymerised glue and keep it fluid. Building on 30 years of Rittschof's experience and Beatriz Orihuela's expertise at growing and reattaching barnacles, Dickinson learned to gently lift polymerised glue away from the pores that secrete the adhesive and quickly collect the minute drops as they oozed from the shell. Working in the cold room to slow the polymerization process, Dickinson had only 5 minutes before each sample polymerized and the glue set solid.
Next the team had to convince themselves that the viscous secretion was glue and not some other body fluid. Dickinson found that the fluid polymerised rapidly and was packed full of protein, just like barnacle glue. Next Dickinson teamed up with Kathy Wahl to use atomic force microscopy to compare the molecular structures of naturally cured glue (from stuck-down barnacles) and his polymerized samples. The two samples were virtually indistinguishable and Dickinson could clearly see tangled webs of fibres in his glue drops, similar to the tangled fibres in blood clots.
But this evidence was still far from proving that barnacle glue cures by the same process as blood clots. Dickinson and Rittschof needed to identify the key proteins that polymerize the cement. Knowing that blood clots are formed when enzymes, known as trypsin-like serine proteases, trigger a cascade of events that culminates in the formation of the long fibres found in blood clots, Dickinson and Rittschoff began searching for the protease in the unpolymerised glue. Separating the glue's components on a gel, Dickinson could see the tell-tale pattern of bands that suggested that a trypsin-like serine protease was present. And when Dickinson added an inhibitor, to inactivate the protease, to a fresh sample of glue, the sample didn't set.
Having convinced themselves that the glue contained a trypsin-like serine protease, the team began to search for other blood-clot-like proteins in the barnacle's secretions. Teaming up with Joseph Bonaventura and Irving Vega, Dickinson chopped each glue component into minute fragments, measured their sizes with mass spectrometry and matched the fragment pattern to known protein sequences. Amazingly, one of the glue proteins was remarkably similar to human factor XIII: a human blood clotting factor that cross-links clot fibres to form a scab. In fact, some regions of the human and barnacle proteins were completely identical. Dickinson and Rittschof had stumbled across the crucial protein that cross-links the glue fibres to cure barnacle cement and it was very similar to factor XIII, an essential human blood-clotting factor.
Rittschof admits that he is shocked that his hypothesis stands up to the tests. 'It seems likely that barnacle glue polymerization is a specialized form of wound healing,' he says and suspects that many other marine animals that rely on glue to get a grip may use the same polymerization mechanism.
REFERENCE: Dickinson, G. H., Vega, I. E., Wahl, K. J., Orihuela, B., Beyley, V., Rodriguez, E. N., Everett, R. K., Bonaventura, J. and Rittschof, D. (2009). Barnacle cement: a polymerization model based on evolutionary concepts. J. Exp. Biol. 212, 3499-3510
Source:
Kathryn Knight
The Company of Biologists
For Liver Cancer Identification Proteomic Profiling Shown More Accurate Than Traditional Biomarkers
As the incidence of liver cancer continues to grow - fueled in large part, by rising rates of hepatitis C infections - so too does the need for tests to help diagnose the disease at an earlier stage. A study appearing in the January 15 issue of Clinical Cancer Research demonstrates that a novel mass-spectrometry based form of proteomic profiling is more accurate than traditional biomarkers in distinguishing liver cancer patients from patients with hepatitis C liver cirrhosis, particularly with regard to identifying patients with small, curable tumors. Led by researchers at Beth Israel Deaconess Medical Center (BIDMC), the study could help lead to earlier diagnostic methods - and subsequent treatments - for liver cancer.
"Proteomics represents a potentially powerful tool for the serologic recognition of protein profiles associated with cancer," explains co-senior author Towia Libermann, PhD, Director of the Genomics Center at BIDMC and Associate Professor of Medicine at Harvard Medical School.
"Although this particular proteomics technology, SELDI-TOF MS [surface enhanced laser desorption/ionization time of flight mass spectrometry] had already proven capable of identifying liver cancer in some limited studies, this was the first time that the technology was compared side-by-side with the clinical standard biomarker in a cohort of patients at risk for developing the disease," adds Liebermann, who is also Director of the Dana-Farber/Harvard Cancer Center Proteomics Core in the Division of Interdisciplinary Medicine and Biotechnology at BIDMC.
Over a single decade, the incidence of liver cancer (hepatocellular carcinoma) increased from 1.8 to 2.5 per 100,000 patients, in large part due to a rise in the spread of hepatitis C virus.
"Hepatitis C has become a tremendous public health problem," explains co-senior author Nezam Afdhal, MD, Director of the Liver Center at BIDMC and Associate Professor of Medicine at Harvard Medical School. "And a significant number of hepatitis C-infected patients will go on to develop liver cirrhosis." Cirrhosis results when healthy tissue is replaced by scar tissue, preventing the liver from properly functioning. Cirrhosis itself is responsible for more than 25,000 deaths each year. But, adds Afdhal, secondarily, cirrhosis greatly increases a person's chances of developing liver cancer.
"Each year, cirrhosis patients have a two to five percent chance that their condition will escalate to cancer," he explains. "And the problem is that, right now, there is no reliable means of detecting liver cancer at an early stage, when surgical treatment is an option. Typically by the time the disease is discovered, the cancer has advanced and treatment options become much more limited."
The best hope for early detection is cancer biomarkers, serum proteins found in altered amounts in blood or other body fluids. The current biomarker for liver cancer in clinical use is alpha fetoprotein (AFP). In many cases, patients with hepatitis C undergo routine monitoring for AFP levels as an indicator of whether tumors may have developed in their livers.
But, as Libermann explains, the AFP biomarker has a number of shortcomings, including false positives and false negatives. "AFP not only fails to detect many early tumors, but it also lacks specificity. Consequently, elevated AFP levels could be indicators of not only cancer, but also of other liver diseases or even benign conditions, while on the other hand, many patients with small tumors will test negative for AFP."
The authors, therefore, decided to evaluate the sensitivy and specificity of SELDI-TOF MS for the detection of liver cancer and to compare its effectiveness with AFP.
Examining serum samples of 92 patients - including 51 patients with liver cirrhosis and 41 patients with liver cancer, and among the cancer patients, individuals with both large and small (less than 2 cm) tumors - by SELDI-TOF mass spectrometry, the investigators were able to identify an 11-protein signature that accurately discriminated between the cirrhosis and cancer patients, first in a training set (made up of 26 cirrhosis and 20 liver cancer patients), and then again in an independent validation set (consisting of 25 cirrhosis and 19 liver cancer patients). The resulting diagnostic value - 74 percent sensitivity and 88 percent specificity - compared favorably with the diagnostic accuracy of AFP (73 percent sensitivity and 71 percent specificity) as well as with two other biomarkers currently in clinical development for liver cancer, AFP-L3 and PIVKA-IL.
"Most strikingly," notes Libermann, "in patients with small tumors (less than 2 cm), where AFP identified only three, and AFP-L3 and PIVKA-II only one each, the 11-protein signature correctly identified seven of eight patients at this early stage of disease.
"Biomarkers play a major role in all aspects of personalized medicine, not only in early disease detection, but also in outcome prediction and evaluation of therapeutic responses," he adds. "This study provides strong evidence that serum contains early detection biomarkers and supports the notion that a combination of multiple biomarkers may prove more effective than individual biomarkers for diagnosis of liver cancer, as well as other cancers."
This study was funded by grants from the National Institutes of Health.
In addition to Libermann and Afdhal, study coauthors include BIDMC investigators Noah Zinkin MD, and Franck Grall, PhD, (joint first authors), Killimanagalam Bhaskar, MD, Hasan Otu, PhD, Dimitrios Spentzos, MD, Brett Kalmowitz, MD, Meghan Wells, Manuel Guerrero, BSc, and John Asara, PhD.
Beth Israel Deaconess Medical Center is a patient care, teaching and research affiliate of Harvard Medical School and consistently ranks among the top four in National Institutes of Health funding among independent hospitals nationwide. BIDMC is clinically affiliated with the Joslin Diabetes Center and is a research partner of Dana-Farber/Harvard Cancer Center. BIDMC is the official hospital of the Boston Red Sox. For more information, visit bidmc.harvard/.
Source: Bonnie Prescott
Beth Israel Deaconess Medical Center
"Proteomics represents a potentially powerful tool for the serologic recognition of protein profiles associated with cancer," explains co-senior author Towia Libermann, PhD, Director of the Genomics Center at BIDMC and Associate Professor of Medicine at Harvard Medical School.
"Although this particular proteomics technology, SELDI-TOF MS [surface enhanced laser desorption/ionization time of flight mass spectrometry] had already proven capable of identifying liver cancer in some limited studies, this was the first time that the technology was compared side-by-side with the clinical standard biomarker in a cohort of patients at risk for developing the disease," adds Liebermann, who is also Director of the Dana-Farber/Harvard Cancer Center Proteomics Core in the Division of Interdisciplinary Medicine and Biotechnology at BIDMC.
Over a single decade, the incidence of liver cancer (hepatocellular carcinoma) increased from 1.8 to 2.5 per 100,000 patients, in large part due to a rise in the spread of hepatitis C virus.
"Hepatitis C has become a tremendous public health problem," explains co-senior author Nezam Afdhal, MD, Director of the Liver Center at BIDMC and Associate Professor of Medicine at Harvard Medical School. "And a significant number of hepatitis C-infected patients will go on to develop liver cirrhosis." Cirrhosis results when healthy tissue is replaced by scar tissue, preventing the liver from properly functioning. Cirrhosis itself is responsible for more than 25,000 deaths each year. But, adds Afdhal, secondarily, cirrhosis greatly increases a person's chances of developing liver cancer.
"Each year, cirrhosis patients have a two to five percent chance that their condition will escalate to cancer," he explains. "And the problem is that, right now, there is no reliable means of detecting liver cancer at an early stage, when surgical treatment is an option. Typically by the time the disease is discovered, the cancer has advanced and treatment options become much more limited."
The best hope for early detection is cancer biomarkers, serum proteins found in altered amounts in blood or other body fluids. The current biomarker for liver cancer in clinical use is alpha fetoprotein (AFP). In many cases, patients with hepatitis C undergo routine monitoring for AFP levels as an indicator of whether tumors may have developed in their livers.
But, as Libermann explains, the AFP biomarker has a number of shortcomings, including false positives and false negatives. "AFP not only fails to detect many early tumors, but it also lacks specificity. Consequently, elevated AFP levels could be indicators of not only cancer, but also of other liver diseases or even benign conditions, while on the other hand, many patients with small tumors will test negative for AFP."
The authors, therefore, decided to evaluate the sensitivy and specificity of SELDI-TOF MS for the detection of liver cancer and to compare its effectiveness with AFP.
Examining serum samples of 92 patients - including 51 patients with liver cirrhosis and 41 patients with liver cancer, and among the cancer patients, individuals with both large and small (less than 2 cm) tumors - by SELDI-TOF mass spectrometry, the investigators were able to identify an 11-protein signature that accurately discriminated between the cirrhosis and cancer patients, first in a training set (made up of 26 cirrhosis and 20 liver cancer patients), and then again in an independent validation set (consisting of 25 cirrhosis and 19 liver cancer patients). The resulting diagnostic value - 74 percent sensitivity and 88 percent specificity - compared favorably with the diagnostic accuracy of AFP (73 percent sensitivity and 71 percent specificity) as well as with two other biomarkers currently in clinical development for liver cancer, AFP-L3 and PIVKA-IL.
"Most strikingly," notes Libermann, "in patients with small tumors (less than 2 cm), where AFP identified only three, and AFP-L3 and PIVKA-II only one each, the 11-protein signature correctly identified seven of eight patients at this early stage of disease.
"Biomarkers play a major role in all aspects of personalized medicine, not only in early disease detection, but also in outcome prediction and evaluation of therapeutic responses," he adds. "This study provides strong evidence that serum contains early detection biomarkers and supports the notion that a combination of multiple biomarkers may prove more effective than individual biomarkers for diagnosis of liver cancer, as well as other cancers."
This study was funded by grants from the National Institutes of Health.
In addition to Libermann and Afdhal, study coauthors include BIDMC investigators Noah Zinkin MD, and Franck Grall, PhD, (joint first authors), Killimanagalam Bhaskar, MD, Hasan Otu, PhD, Dimitrios Spentzos, MD, Brett Kalmowitz, MD, Meghan Wells, Manuel Guerrero, BSc, and John Asara, PhD.
Beth Israel Deaconess Medical Center is a patient care, teaching and research affiliate of Harvard Medical School and consistently ranks among the top four in National Institutes of Health funding among independent hospitals nationwide. BIDMC is clinically affiliated with the Joslin Diabetes Center and is a research partner of Dana-Farber/Harvard Cancer Center. BIDMC is the official hospital of the Boston Red Sox. For more information, visit bidmc.harvard/.
Source: Bonnie Prescott
Beth Israel Deaconess Medical Center
New 'clock gene' uncovered
The solving of the human genome sequence was hailed a few years ago as biology's equivalent to landing a man on the moon -- a mammoth milestone of monumental importance.
However, unlike the first moon shot, the real milestone of the human genome project is not a singular event. The genome project's giant leap for mankind is coming not with a single small step taken on one summer's night but with thousands of small steps spread out over the course of several years.
The importance of solving the human genome and the genomes of other species is that those billions of letters of DNA are deposited into databases and become available to scientists everywhere to conduct post-genomic research. Such research includes annotating the human genome -- matching the DNA codes that hold the secret to human life to the genes, proteins, physiologies, and behaviors that define human life. Such work holds great promise for future medicine, and scientists have been investigating how the genes in the human genome actually contribute to the biology of health and disease.
In the last few years, a team of scientists from The Scripps Research Institute and the Genomics Institute of the Novartis Research Foundation (GNF) has been working toward an understanding the biology of circadian rhythms -- the cyclic, clock-like expression of genes in the body.
The team, led by Steve Kay, Ph.D., and John Hogenesch, Ph.D., uses a combination of genomic, biochemical, and behavioral approaches, work that recently revealed a "new genetic component of the mammalian clock -- a protein known as "Rora."
This discovery may someday help people with jet lag, shift workers who feel wiped out after working a night shift, and people with more serious sleep disorders, many of which are related to circadian rhythms, say the scientists, who report their findings in the latest issue of the journal Neuron.
In addition to Kay and Hogenesch, authors on the paper include Satchidananda Panda, Ph.D., a graduate of Scripps Research's Kellogg School of Science and Technology and currently a postdoctoral fellow at GNF, and Trey Sato, Ph.D., currently a postdoctoral fellow at the Scripps Research La Jolla campus who will arrive at Scripps Florida as a staff scientist in December. Kay is a professor of cell biology and the director of the Institute for Childhood and Neglected Diseases at The Scripps Research Institute. Hogenesch is currently a researcher at GNF and will soon become an associate professor and the head of Genome Technology at Scripps Florida.
Circadian Rhythms and Jet Leg: Blame the Liver
Science has known for years that humans, mice, and many other plants and animals possess internal clocks that keep track of time and coordinate physiological, behavioral, and biochemical processes with the rhythm of the 24-hour cycle of day and night.
These so-called circadian rhythms offer distinct advantages to organisms that use them. Plants, for example, shut down photosynthesis at night, and they gear up their photosynthetic machinery and raise their leaves just before dawn. They use their clocks to measure day length and in that way anticipate changes in the seasonsa system that determines when they shed their leaves, produce seeds, and make flowers or fruit.
Humans also have circadian rhythms, and we entrain our internal clocks to the 24-hour day. Under normal conditions, we time our major activities with daylight, we sleep during the nighttime, and some of our vital signs follow this pattern. Our blood pressure fluctuates daily, rising and falling at predictable times of day and night.
Scientists have provided evidence of the existence of internal clock mechanisms by placing organisms like rodents in chambers isolated from day/night cycles. In spite of this, the animals' rhythms still cycle approximately every 24 hours.
"Even if you were to put the lights out on us, we would still [time] our activity to when our body expects light," says Hogenesch.
One of the most intriguing aspects of this is that the mammalian clock is actually composed of many separate clocks that maintain different circadian rhythms specifically adapted to the various tissues of the body.
The liver, the heart, and the kidneys each have their own distinct clocks. The liver, for instance, expresses a number of enzymes that remove toxic substances from the bloodstream during the day, which corresponds with the prime time for food (and toxic compound) intake.
Coordinating the activities of all these different clocks is the job of the master circadian oscillator, or master clock, which in humans and other mammals is the suprachiasmatic nuclei, a small center in the brain's hypothalamus with about 10,000 neurons that sits above the optic chasmthe location where the optic nerves cross each other.
This master clock synchronizes independent clocks that reside in peripheral tissues, and every 24 hours, the master clock cycles. This cycling involves the coordinated expression of many genes involved in feedback loops, in which the expression of one gene turns on the expression of a second gene, which turns off the first gene, which turns off the second gene, which turns the first gene back on, etc., day in and day out.
However, in real life, the situation is not so simple as a loop involving only two genes. Multiple clock genes in mammals are involved in overlapping feedback loops. The clock also keeps time dynamically, constantly shifting to stay in time with environmental changes. And these adjustments vary from tissue to tissue. Different tissues respond to the clock in their own way, and they all reset their own clocks independently of one another.
The heart, for instance, is like an obsessive-compulsive clock watcher. It monitors the master clock closely and trains its circadian rhythms to changes in the master clock rapidly.
The liver, on the other hand, is more slothlike. It is less attentive to the master clock, and it takes several days for the liver to catch up with changes in the master clock. Incidentally, the problems related to jet lag and night shift work are often caused by the liver's inability to respond rapidly to changes in the sleep-wake cycle.
One key to understanding the intricacies of the mammalian clock and to addressing problems with jet lag and certain sleep disorders is discovering the different genes that communicate the timing of the master clock with the circadian rhythms of the various tissues. Not all of these genes are known.
Identifying the Clock Genes
Panda and Sato performed gene array experiments on different samples of mammalian tissue to determine which genes are cycling and which might be components of the clock.
These experiments involve taking cells from the particular tissues, recovering the cells' expressed genes (in the form of messenger RNA, or mRNA), chopping the mRNA into fragments, and plopping the mixture of fragmented mRNA on a gene chip -- a glass or silicon wafer that has thousands of short pieces of RNA attached to it with sequences corresponding to known genes.
These short pieces are laid out in a grid, and genes that are expressed in the tissue will bind to complementary pieces of mRNA on the grid. Then by looking to see which pieces on the grid have RNA bound to them, the scientists are able to determine which genes were being expressed in the sample.
DNA and RNA chips have become a standard tool for genomics research in the last couple of years, and scientists can quite easily put a large number of different oligonucleotide pieces -- conceivably even all the known genes in an organism -- on a single chip.
The Scripps Research and GNF team charted the time course of circadian rhythms by looking at the expression of 10,000 different genes in various murine tissues every three hours over the course of two days. They examined the gene expression in the liver, kidney, aorta, skeletal muscle, and the suprachiasmatic nucleus of the hypothalamus, and they looked for cyclic expression.
What the data showed was that about 10 percent of the genes cycled, but most showed little overlap from tissue to tissue. This type of cycling has to do with local physiology -- for instance, the liver's expression of certain enzymes at certain times of the day. What they were really interested in were the overlapping cyclers -- those genes that cycled in all tissue types. These, they reckoned, would be part of the master clock.
They found 50 genes that cycled at the same time throughout the day across all the various tissues, and they speculated that this collection would include both known and unknown circadian rhythm genes. Indeed, known circadian genes were among the 50, but there were dozens of other cycling genes that had not been previously identified as clock genes.
The scientists speculated that some of these other genes may be part of the mammalian circadian clock, and they reasoned that if they were, they might interact with some of the known clock genes. So they designed an experiment to see if expressing the new genes in a cell could change the expression of known clock genes. To do this, they used a biochemical assay to detect if any of these genes had the ability to control transcription -- that crucial first step in the expression of a gene.
They discovered that a family of genes called the retinoic acid receptor-related orphan receptor-a (Rora) cycled and had the ability to control transcription. These are so named because they are similar in amino acid sequence to the retinoic acid receptor genes, although they do not have the same function and do not bind to retinoic acid. (They are called orphan receptors because it is not known what activates them).
The Flick of a Switch
Rora is a gene that produces a transcription factor -- a type of regulatory protein that binds to DNA and can turn gene expression on or off like the flicking of a switch.
Rora, once it is turned on, activates the transcription of a gene that encodes another transcription activator known as Bmal-1, which is one of the known circadian genes. Bmal-1 drives the transcription of a protein called cryptochrome, which subsequently inhibits the ability of Bmal-1 to activate cryptochrome's own transcription. This feedback loop is what keeps the body entrained to a 24-hour day.
Since Bmal-1 is so crucial to keeping the body's clock entrained, finding something like Rora, which alters Bmal-1's expression, is significant and suggests that Rora is also part of the mammalian clock. But the scientists wanted to go further and prove that Rora protein plays a role in the circadian rhythms inside a living creature.
They observed a mutant murine model that has a defective Rora gene. This murine model is called "staggerer" because its genetic defect causes a characteristic loss of coordination.
As it turns out, the staggerer model with a defective Rora gene also has a defect in its ability to regulate its circadian clock. The team of researchers showed that staggerers have aberrant circadian rhythyms and a shortened clock that is only 23.2 hours long. This situation is sort of like a grandfather clock that cannot keep good time and runs too fast because it has a faulty spring balance.
"What we are showing is that circadian clocks are composed of interlocking feedback loops," says Kay. The overlapping feedback, says Kay, is probably there for a number of reasons. It makes the clock more robust and resilient to change. It means that there is more than one cycle in which changes to clock genes can affect changes to other genes, and therefore the clock can be reset more easily.
Knowing that Rora is a component of the mammalian clock is significant because it may be a valuable target for the development of compounds to correct sleep disorders, many of which are related to circadian rhythms, and for countering the most common circadian problems -- the jet-lag one feels after overseas flights or fatigue when working night shifts.
"Identifying a ligand for Rora might help reduce the effects of jet lag," says Sato.
The article, "A Functional Genomics Strategy Reveals Rora as a Component of the Mammalian Circadian Clock" was authored by Trey K. Sato, Satchidananda Panda, Loren J. Miraglia, Teresa M. Reyes, Radu D. Rudic, Peter McNamara, Kinnery A. Naik, Garret A. FitzGerald, Steve A. Kay, and John B. Hogenesch and appears in the August 19, 2004 issue of the journal Neuron.
This work was supported by the Novartis Research Foundation, the National Institutes of Health, and a Rena and Victor Damone Postdoctoral Fellow Fellowship from the American Cancer Society.
About The Scripps Research Institute
The Scripps Research Institute in La Jolla, California, is one of the world's largest, private, non-profit biomedical research organizations. It stands at the forefront of basic biomedical science that seeks to comprehend the most fundamental processes of life. Scripps Research is internationally recognized for its research into immunology, molecular and cellular biology, chemistry, neurosciences, autoimmune diseases, cardiovascular diseases and synthetic vaccine development.
Contact: Keith McKeown
kmckeownscripps
858-784-8134
Scripps Research Institute
However, unlike the first moon shot, the real milestone of the human genome project is not a singular event. The genome project's giant leap for mankind is coming not with a single small step taken on one summer's night but with thousands of small steps spread out over the course of several years.
The importance of solving the human genome and the genomes of other species is that those billions of letters of DNA are deposited into databases and become available to scientists everywhere to conduct post-genomic research. Such research includes annotating the human genome -- matching the DNA codes that hold the secret to human life to the genes, proteins, physiologies, and behaviors that define human life. Such work holds great promise for future medicine, and scientists have been investigating how the genes in the human genome actually contribute to the biology of health and disease.
In the last few years, a team of scientists from The Scripps Research Institute and the Genomics Institute of the Novartis Research Foundation (GNF) has been working toward an understanding the biology of circadian rhythms -- the cyclic, clock-like expression of genes in the body.
The team, led by Steve Kay, Ph.D., and John Hogenesch, Ph.D., uses a combination of genomic, biochemical, and behavioral approaches, work that recently revealed a "new genetic component of the mammalian clock -- a protein known as "Rora."
This discovery may someday help people with jet lag, shift workers who feel wiped out after working a night shift, and people with more serious sleep disorders, many of which are related to circadian rhythms, say the scientists, who report their findings in the latest issue of the journal Neuron.
In addition to Kay and Hogenesch, authors on the paper include Satchidananda Panda, Ph.D., a graduate of Scripps Research's Kellogg School of Science and Technology and currently a postdoctoral fellow at GNF, and Trey Sato, Ph.D., currently a postdoctoral fellow at the Scripps Research La Jolla campus who will arrive at Scripps Florida as a staff scientist in December. Kay is a professor of cell biology and the director of the Institute for Childhood and Neglected Diseases at The Scripps Research Institute. Hogenesch is currently a researcher at GNF and will soon become an associate professor and the head of Genome Technology at Scripps Florida.
Circadian Rhythms and Jet Leg: Blame the Liver
Science has known for years that humans, mice, and many other plants and animals possess internal clocks that keep track of time and coordinate physiological, behavioral, and biochemical processes with the rhythm of the 24-hour cycle of day and night.
These so-called circadian rhythms offer distinct advantages to organisms that use them. Plants, for example, shut down photosynthesis at night, and they gear up their photosynthetic machinery and raise their leaves just before dawn. They use their clocks to measure day length and in that way anticipate changes in the seasonsa system that determines when they shed their leaves, produce seeds, and make flowers or fruit.
Humans also have circadian rhythms, and we entrain our internal clocks to the 24-hour day. Under normal conditions, we time our major activities with daylight, we sleep during the nighttime, and some of our vital signs follow this pattern. Our blood pressure fluctuates daily, rising and falling at predictable times of day and night.
Scientists have provided evidence of the existence of internal clock mechanisms by placing organisms like rodents in chambers isolated from day/night cycles. In spite of this, the animals' rhythms still cycle approximately every 24 hours.
"Even if you were to put the lights out on us, we would still [time] our activity to when our body expects light," says Hogenesch.
One of the most intriguing aspects of this is that the mammalian clock is actually composed of many separate clocks that maintain different circadian rhythms specifically adapted to the various tissues of the body.
The liver, the heart, and the kidneys each have their own distinct clocks. The liver, for instance, expresses a number of enzymes that remove toxic substances from the bloodstream during the day, which corresponds with the prime time for food (and toxic compound) intake.
Coordinating the activities of all these different clocks is the job of the master circadian oscillator, or master clock, which in humans and other mammals is the suprachiasmatic nuclei, a small center in the brain's hypothalamus with about 10,000 neurons that sits above the optic chasmthe location where the optic nerves cross each other.
This master clock synchronizes independent clocks that reside in peripheral tissues, and every 24 hours, the master clock cycles. This cycling involves the coordinated expression of many genes involved in feedback loops, in which the expression of one gene turns on the expression of a second gene, which turns off the first gene, which turns off the second gene, which turns the first gene back on, etc., day in and day out.
However, in real life, the situation is not so simple as a loop involving only two genes. Multiple clock genes in mammals are involved in overlapping feedback loops. The clock also keeps time dynamically, constantly shifting to stay in time with environmental changes. And these adjustments vary from tissue to tissue. Different tissues respond to the clock in their own way, and they all reset their own clocks independently of one another.
The heart, for instance, is like an obsessive-compulsive clock watcher. It monitors the master clock closely and trains its circadian rhythms to changes in the master clock rapidly.
The liver, on the other hand, is more slothlike. It is less attentive to the master clock, and it takes several days for the liver to catch up with changes in the master clock. Incidentally, the problems related to jet lag and night shift work are often caused by the liver's inability to respond rapidly to changes in the sleep-wake cycle.
One key to understanding the intricacies of the mammalian clock and to addressing problems with jet lag and certain sleep disorders is discovering the different genes that communicate the timing of the master clock with the circadian rhythms of the various tissues. Not all of these genes are known.
Identifying the Clock Genes
Panda and Sato performed gene array experiments on different samples of mammalian tissue to determine which genes are cycling and which might be components of the clock.
These experiments involve taking cells from the particular tissues, recovering the cells' expressed genes (in the form of messenger RNA, or mRNA), chopping the mRNA into fragments, and plopping the mixture of fragmented mRNA on a gene chip -- a glass or silicon wafer that has thousands of short pieces of RNA attached to it with sequences corresponding to known genes.
These short pieces are laid out in a grid, and genes that are expressed in the tissue will bind to complementary pieces of mRNA on the grid. Then by looking to see which pieces on the grid have RNA bound to them, the scientists are able to determine which genes were being expressed in the sample.
DNA and RNA chips have become a standard tool for genomics research in the last couple of years, and scientists can quite easily put a large number of different oligonucleotide pieces -- conceivably even all the known genes in an organism -- on a single chip.
The Scripps Research and GNF team charted the time course of circadian rhythms by looking at the expression of 10,000 different genes in various murine tissues every three hours over the course of two days. They examined the gene expression in the liver, kidney, aorta, skeletal muscle, and the suprachiasmatic nucleus of the hypothalamus, and they looked for cyclic expression.
What the data showed was that about 10 percent of the genes cycled, but most showed little overlap from tissue to tissue. This type of cycling has to do with local physiology -- for instance, the liver's expression of certain enzymes at certain times of the day. What they were really interested in were the overlapping cyclers -- those genes that cycled in all tissue types. These, they reckoned, would be part of the master clock.
They found 50 genes that cycled at the same time throughout the day across all the various tissues, and they speculated that this collection would include both known and unknown circadian rhythm genes. Indeed, known circadian genes were among the 50, but there were dozens of other cycling genes that had not been previously identified as clock genes.
The scientists speculated that some of these other genes may be part of the mammalian circadian clock, and they reasoned that if they were, they might interact with some of the known clock genes. So they designed an experiment to see if expressing the new genes in a cell could change the expression of known clock genes. To do this, they used a biochemical assay to detect if any of these genes had the ability to control transcription -- that crucial first step in the expression of a gene.
They discovered that a family of genes called the retinoic acid receptor-related orphan receptor-a (Rora) cycled and had the ability to control transcription. These are so named because they are similar in amino acid sequence to the retinoic acid receptor genes, although they do not have the same function and do not bind to retinoic acid. (They are called orphan receptors because it is not known what activates them).
The Flick of a Switch
Rora is a gene that produces a transcription factor -- a type of regulatory protein that binds to DNA and can turn gene expression on or off like the flicking of a switch.
Rora, once it is turned on, activates the transcription of a gene that encodes another transcription activator known as Bmal-1, which is one of the known circadian genes. Bmal-1 drives the transcription of a protein called cryptochrome, which subsequently inhibits the ability of Bmal-1 to activate cryptochrome's own transcription. This feedback loop is what keeps the body entrained to a 24-hour day.
Since Bmal-1 is so crucial to keeping the body's clock entrained, finding something like Rora, which alters Bmal-1's expression, is significant and suggests that Rora is also part of the mammalian clock. But the scientists wanted to go further and prove that Rora protein plays a role in the circadian rhythms inside a living creature.
They observed a mutant murine model that has a defective Rora gene. This murine model is called "staggerer" because its genetic defect causes a characteristic loss of coordination.
As it turns out, the staggerer model with a defective Rora gene also has a defect in its ability to regulate its circadian clock. The team of researchers showed that staggerers have aberrant circadian rhythyms and a shortened clock that is only 23.2 hours long. This situation is sort of like a grandfather clock that cannot keep good time and runs too fast because it has a faulty spring balance.
"What we are showing is that circadian clocks are composed of interlocking feedback loops," says Kay. The overlapping feedback, says Kay, is probably there for a number of reasons. It makes the clock more robust and resilient to change. It means that there is more than one cycle in which changes to clock genes can affect changes to other genes, and therefore the clock can be reset more easily.
Knowing that Rora is a component of the mammalian clock is significant because it may be a valuable target for the development of compounds to correct sleep disorders, many of which are related to circadian rhythms, and for countering the most common circadian problems -- the jet-lag one feels after overseas flights or fatigue when working night shifts.
"Identifying a ligand for Rora might help reduce the effects of jet lag," says Sato.
The article, "A Functional Genomics Strategy Reveals Rora as a Component of the Mammalian Circadian Clock" was authored by Trey K. Sato, Satchidananda Panda, Loren J. Miraglia, Teresa M. Reyes, Radu D. Rudic, Peter McNamara, Kinnery A. Naik, Garret A. FitzGerald, Steve A. Kay, and John B. Hogenesch and appears in the August 19, 2004 issue of the journal Neuron.
This work was supported by the Novartis Research Foundation, the National Institutes of Health, and a Rena and Victor Damone Postdoctoral Fellow Fellowship from the American Cancer Society.
About The Scripps Research Institute
The Scripps Research Institute in La Jolla, California, is one of the world's largest, private, non-profit biomedical research organizations. It stands at the forefront of basic biomedical science that seeks to comprehend the most fundamental processes of life. Scripps Research is internationally recognized for its research into immunology, molecular and cellular biology, chemistry, neurosciences, autoimmune diseases, cardiovascular diseases and synthetic vaccine development.
Contact: Keith McKeown
kmckeownscripps
858-784-8134
Scripps Research Institute
Getting Obese Mice Moving And Curing Their Diabetes
Mice lacking the fat hormone leptin or the ability to respond to it become morbidly obese and severely diabetic - not to mention downright sluggish. Now, a new study in the June Cell Metabolism shows that blood sugar control in those animals can be completely restored by returning leptin sensitivity to a single class of neurons in the brain, which account for only a small fraction of those that normally carry the hormone receptors.
"Just the receptors in this little group of neurons are sufficient to do the job," said Christian Bjorbaek of Harvard Medical School.
What's more, animals with leptin receptors only in the so-called pro-opiomelanocortin (POMC) neurons spontaneously increase their physical activity levels despite the fact that they remain profoundly obese. While understanding exactly how the POMC neurons act on other organs remains a future challenge, the discovery suggests that drugs designed to tap into the pathway - turning up or down the dial, so to speak - might have benefit for those with severe diabetes and obesity, according to the researchers.
Such drugs might even encourage obese individuals to get moving. "This gives us the opportunity to search for drugs that might induce the desire or will to voluntarily exercise," Bjorbaek said.
Leptin was first identified 15 years ago and made famous for its ability to curb appetite and lead to weight loss. It is known to play a pivotal role in energy balance through its effects on the central nervous system, specifically by acting on a hypothalamic brain region known as the arcuate nucleus (ARC). The ARC contains two types of leptin-responsive neurons, the POMC neurons, which cause a loss of appetite, and the so-called Agouti-related peptide (AgRP) neurons, which do the opposite.
Studies had also revealed a role for leptin in blood sugar control and activity level, also via effects on the ARC. However, scientists still didn't know which neurons were responsible, until now.
When the researchers restored leptin receptor activity in POMC neurons of otherwise leptin-resistant, obese, and diabetic mice, the animals began eating about 30 percent fewer calories and lost a modest amount of weight. Remarkably, the researchers report, their blood sugar levels returned to normal independently of any change in their eating habits or weight. The animals also doubled their activity levels.
Whether this particular bunch of neurons also plays a similarly important role in animals that are lean remains uncertain, Bjorbaek said. "It may be that in the context of severe obesity and diabetes, these neurons do something they don't normally do," he said. But, he added, even if that were the case, it may not matter when it comes to its potential as a therapeutic target.
The researchers include Lihong Huo, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA, Kevin Gamber, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA, Sarah Greeley, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA, Jose Silva, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA; Nicholas Huntoon, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA, Xing-Hong Leng, Baylor College of Medicine, Houston, TX; and Christian Bjørbæk, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA.
Source:
Cathleen Genova
Cell Press
"Just the receptors in this little group of neurons are sufficient to do the job," said Christian Bjorbaek of Harvard Medical School.
What's more, animals with leptin receptors only in the so-called pro-opiomelanocortin (POMC) neurons spontaneously increase their physical activity levels despite the fact that they remain profoundly obese. While understanding exactly how the POMC neurons act on other organs remains a future challenge, the discovery suggests that drugs designed to tap into the pathway - turning up or down the dial, so to speak - might have benefit for those with severe diabetes and obesity, according to the researchers.
Such drugs might even encourage obese individuals to get moving. "This gives us the opportunity to search for drugs that might induce the desire or will to voluntarily exercise," Bjorbaek said.
Leptin was first identified 15 years ago and made famous for its ability to curb appetite and lead to weight loss. It is known to play a pivotal role in energy balance through its effects on the central nervous system, specifically by acting on a hypothalamic brain region known as the arcuate nucleus (ARC). The ARC contains two types of leptin-responsive neurons, the POMC neurons, which cause a loss of appetite, and the so-called Agouti-related peptide (AgRP) neurons, which do the opposite.
Studies had also revealed a role for leptin in blood sugar control and activity level, also via effects on the ARC. However, scientists still didn't know which neurons were responsible, until now.
When the researchers restored leptin receptor activity in POMC neurons of otherwise leptin-resistant, obese, and diabetic mice, the animals began eating about 30 percent fewer calories and lost a modest amount of weight. Remarkably, the researchers report, their blood sugar levels returned to normal independently of any change in their eating habits or weight. The animals also doubled their activity levels.
Whether this particular bunch of neurons also plays a similarly important role in animals that are lean remains uncertain, Bjorbaek said. "It may be that in the context of severe obesity and diabetes, these neurons do something they don't normally do," he said. But, he added, even if that were the case, it may not matter when it comes to its potential as a therapeutic target.
The researchers include Lihong Huo, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA, Kevin Gamber, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA, Sarah Greeley, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA, Jose Silva, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA; Nicholas Huntoon, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA, Xing-Hong Leng, Baylor College of Medicine, Houston, TX; and Christian Bjørbæk, Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, MA.
Source:
Cathleen Genova
Cell Press
The Molecular Basis Of Detecting Tissue-Damaging Chemicals Goes Back More 500 Million Years
Whenever you choke on acrid cigarette smoke, feel like you're burning up from a mouthful of wasabi-laced sushi, or cry while cutting raw onions and garlic, your response is being triggered by a primordial chemical sensor conserved across some 500 million years of animal evolution, report Brandeis University scientists in a study in Nature this week.
Chemical nociception, the detection of tissue-damaging pungent chemicals like those found in wasabi, tear gas and cigarette smoke, is triggered by a protein receptor known as TRPA1, which is found throughout the human body in the nose, mouth, skin, lungs, and GI tract. Studying the chemical sensors of Drosophila fruit flies, scientists discovered that flies use their ortholog of the human TRPA1 sensor for the same purpose.
Using a combination of behavior, physiology and phylogenetics, the scientists discovered that this defensive response to noxious compounds is an evolutionary stalwart cutting across immense time scales and linking humans, insects and many other animals back to their common ancestor over 500 million years ago, said lead author and biologist Paul Garrity.
The ability to detect such noxious compounds, known as reactive electrophiles, is important for animal survival, prompting them to avoid potentially toxic food or dangerous situations. These receptors give animals a leg-up in survival by acting as a biological warning system, as it were. In humans, chemical nociception causes pain and inflammation.
"What the study, spearheaded by Kyeongjin Kang in my lab, shows, is that this chemical sense is nearly as ancient as vision," said Garrity. "While many aspects of other chemical senses like taste and smell have been independently invented multiple times over the course of animal evolution, the chemical sense that detects these reactive compounds is different. It uses a detector we have inherited in largely unaltered form from an organism that lived a half-billion years ago, an organism that is not only our ancestor, but the ancestor of every vertebrate and invertebrate alive today."
Working with biochemist Doug Theobald, the team reconstructed TRPA1's family tree back some 700 million years using a variety of bioinformatic methods. "We discovered that a new branch split off the tree at least 500 million years ago, and that this new branch, the TRPA1 branch, appeared to have had all the features needed for chemical sensing even back then," said Garrity. "Since that time, it appears that most animals, including humans, have maintained this same ancient system for detecting reactive chemicals."
And therein lies some of the future promise of harnessing TRPA1. Because the receptor is so widely dispersed throughout the animal kingdom, it holds promise both as a target for therapeutics and deterrents. Understanding more about how the receptor works may help lead to important applications in medicine and industry.
"One of the great things about studying TRPA1 is that basic science knowledge, of the kind you get working with fruit flies, can be applied in so many different ways. By learning more about how TRPA1 works, scientists can come up with new ways to turn the receptor off in humans to treat pain and inflammation. And they can also come up with new ways to turn the receptor on in pests like malaria-carrying mosquitoes and aphids to deter them from transmitting disease and destroying crops." said Garrity.
The other authors of the paper are Kyeongjin Kang, Stefan R. Pulver, now at the University of Cambridge, Vincent C. Panzano, Elaine C. Chang, Leslie C. Griffith, and Douglas L. Theobald.
The study was funded by the National Institute of Mental Health and the National Institute of Neurological Disorders and Stroke.
Source:
Laura Gardner
Brandeis University
Chemical nociception, the detection of tissue-damaging pungent chemicals like those found in wasabi, tear gas and cigarette smoke, is triggered by a protein receptor known as TRPA1, which is found throughout the human body in the nose, mouth, skin, lungs, and GI tract. Studying the chemical sensors of Drosophila fruit flies, scientists discovered that flies use their ortholog of the human TRPA1 sensor for the same purpose.
Using a combination of behavior, physiology and phylogenetics, the scientists discovered that this defensive response to noxious compounds is an evolutionary stalwart cutting across immense time scales and linking humans, insects and many other animals back to their common ancestor over 500 million years ago, said lead author and biologist Paul Garrity.
The ability to detect such noxious compounds, known as reactive electrophiles, is important for animal survival, prompting them to avoid potentially toxic food or dangerous situations. These receptors give animals a leg-up in survival by acting as a biological warning system, as it were. In humans, chemical nociception causes pain and inflammation.
"What the study, spearheaded by Kyeongjin Kang in my lab, shows, is that this chemical sense is nearly as ancient as vision," said Garrity. "While many aspects of other chemical senses like taste and smell have been independently invented multiple times over the course of animal evolution, the chemical sense that detects these reactive compounds is different. It uses a detector we have inherited in largely unaltered form from an organism that lived a half-billion years ago, an organism that is not only our ancestor, but the ancestor of every vertebrate and invertebrate alive today."
Working with biochemist Doug Theobald, the team reconstructed TRPA1's family tree back some 700 million years using a variety of bioinformatic methods. "We discovered that a new branch split off the tree at least 500 million years ago, and that this new branch, the TRPA1 branch, appeared to have had all the features needed for chemical sensing even back then," said Garrity. "Since that time, it appears that most animals, including humans, have maintained this same ancient system for detecting reactive chemicals."
And therein lies some of the future promise of harnessing TRPA1. Because the receptor is so widely dispersed throughout the animal kingdom, it holds promise both as a target for therapeutics and deterrents. Understanding more about how the receptor works may help lead to important applications in medicine and industry.
"One of the great things about studying TRPA1 is that basic science knowledge, of the kind you get working with fruit flies, can be applied in so many different ways. By learning more about how TRPA1 works, scientists can come up with new ways to turn the receptor off in humans to treat pain and inflammation. And they can also come up with new ways to turn the receptor on in pests like malaria-carrying mosquitoes and aphids to deter them from transmitting disease and destroying crops." said Garrity.
The other authors of the paper are Kyeongjin Kang, Stefan R. Pulver, now at the University of Cambridge, Vincent C. Panzano, Elaine C. Chang, Leslie C. Griffith, and Douglas L. Theobald.
The study was funded by the National Institute of Mental Health and the National Institute of Neurological Disorders and Stroke.
Source:
Laura Gardner
Brandeis University
Agios Announces Cancer Cell Publication Of Research Illuminating Link Between Cancer Metabolism And Epigenetics
Agios Pharmaceuticals, the leading biopharmaceutical company focused on discovering and developing novel drugs in the rapidly emerging field of cancer metabolism, announced the publication of a paper in Cancer Cell that illuminates the mechanism by which 2-hydroxyglutarate (2HG), and the mutations in the metabolic genes IDH1 and IDH2 that produce it, may be driving the growth of tumors in patients with acute myeloid leukemia (AML). This research was conducted as a collaboration between Weill Cornell Medical College, Memorial Sloan Kettering Cancer Center, the University of Pennsylvania and Agios.
IDH1 and IDH2 are normal metabolic enzymes that are mutated in several forms of cancer, including AML. In a 2009 publication in Nature, Agios researchers previously established that mutated IDH1 and IDH2 have novel enzyme activity (consistent with a cancer-causing gene or oncogene) producing high levels of 2HG in cancer cells with IDH mutations. This new research suggests that high 2HG levels may trigger epigenetic changes within the cells - a known cancer-causing mechanism - thus strengthening the biological link between IDH mutations, 2HG and the development of cancer.
"This study broadens our understanding of the role of IDH1 and IDH2 mutations in AML, where novel therapies are desperately needed," said David Schenkein, M.D., chief executive officer of Agios. "These unique insights will help in the development of programs directed towards finding therapies for AML, and other IDH associated diseases. More broadly, this finding further validates the field of cancer metabolism by demonstrating how altered metabolic processes can cause cancer-related changes in tumors."
In this study, researchers conducted DNA sequencing and DNA methylation analysis for AML-associated recurrent mutations, including IDH1 and IDH2. Specimens were taken from 385 patients with AML less than 60 years of age who are enrolled in a Phase 3 multicenter Eastern Cooperative Oncology Group clinical trial.
The study found that IDH mutations induce DNA hypermethylation, an epigenetic phenomenon noted in many cancer cells, and impair differentiation in hematopoietic cells. Similar DNA hypermethylation effects are caused through the loss of function of TET2, a demethylase enzyme that is also mutated in leukemia. This study supports IDH-mutant and TET2-mutant leukemias as biologically distinct disease subtypes and links cancer metabolism with epigenetic control of gene expression.
"Earlier research about the role of mutant IDH and the discovery of 2HG was very exciting, but until now, we haven't really understood the relationship between 2HG and cancer," said Ari Melnick, M.D., associate professor of medicine, co-director, medical research track at Weill Cornell Medical College. "This study broadens and contextualizes our understanding of the biology of IDH mutations. These findings are meaningful not just for researchers in AML and gliomas, who have historically been interested in IDH, but also for scientists working more broadly in both cancer metabolism and epigenetics."
About Cancer Metabolism
Cancer metabolism is a new and exciting field of biology that provides a novel approach to treating cancer. Cancer cell metabolism is marked by profound changes in nutrient requirements and usage to ensure cell proliferation and survival. Research in the field has demonstrated that cancer cells become addicted to certain fuel sources and metabolic pathways. In cancer, this metabolic reprogramming is coordinated with proliferative signaling and regulated by the same oncogenes and tumor suppressor genes to ensure efficient proliferation. Glycolysis (sugar metabolism), fatty acid metabolism and autophagy (self metabolism) are three pathways shown to play a critical role in cancer metabolism. Identifying and disrupting certain enzymes in these, and perhaps other, metabolic pathways provides a powerful intervention point for discovery and development of cancer therapeutics.
Source:
Agios Pharmaceuticals
IDH1 and IDH2 are normal metabolic enzymes that are mutated in several forms of cancer, including AML. In a 2009 publication in Nature, Agios researchers previously established that mutated IDH1 and IDH2 have novel enzyme activity (consistent with a cancer-causing gene or oncogene) producing high levels of 2HG in cancer cells with IDH mutations. This new research suggests that high 2HG levels may trigger epigenetic changes within the cells - a known cancer-causing mechanism - thus strengthening the biological link between IDH mutations, 2HG and the development of cancer.
"This study broadens our understanding of the role of IDH1 and IDH2 mutations in AML, where novel therapies are desperately needed," said David Schenkein, M.D., chief executive officer of Agios. "These unique insights will help in the development of programs directed towards finding therapies for AML, and other IDH associated diseases. More broadly, this finding further validates the field of cancer metabolism by demonstrating how altered metabolic processes can cause cancer-related changes in tumors."
In this study, researchers conducted DNA sequencing and DNA methylation analysis for AML-associated recurrent mutations, including IDH1 and IDH2. Specimens were taken from 385 patients with AML less than 60 years of age who are enrolled in a Phase 3 multicenter Eastern Cooperative Oncology Group clinical trial.
The study found that IDH mutations induce DNA hypermethylation, an epigenetic phenomenon noted in many cancer cells, and impair differentiation in hematopoietic cells. Similar DNA hypermethylation effects are caused through the loss of function of TET2, a demethylase enzyme that is also mutated in leukemia. This study supports IDH-mutant and TET2-mutant leukemias as biologically distinct disease subtypes and links cancer metabolism with epigenetic control of gene expression.
"Earlier research about the role of mutant IDH and the discovery of 2HG was very exciting, but until now, we haven't really understood the relationship between 2HG and cancer," said Ari Melnick, M.D., associate professor of medicine, co-director, medical research track at Weill Cornell Medical College. "This study broadens and contextualizes our understanding of the biology of IDH mutations. These findings are meaningful not just for researchers in AML and gliomas, who have historically been interested in IDH, but also for scientists working more broadly in both cancer metabolism and epigenetics."
About Cancer Metabolism
Cancer metabolism is a new and exciting field of biology that provides a novel approach to treating cancer. Cancer cell metabolism is marked by profound changes in nutrient requirements and usage to ensure cell proliferation and survival. Research in the field has demonstrated that cancer cells become addicted to certain fuel sources and metabolic pathways. In cancer, this metabolic reprogramming is coordinated with proliferative signaling and regulated by the same oncogenes and tumor suppressor genes to ensure efficient proliferation. Glycolysis (sugar metabolism), fatty acid metabolism and autophagy (self metabolism) are three pathways shown to play a critical role in cancer metabolism. Identifying and disrupting certain enzymes in these, and perhaps other, metabolic pathways provides a powerful intervention point for discovery and development of cancer therapeutics.
Source:
Agios Pharmaceuticals
Incomplete Reproductive Isolation Following Host Shift In Brood Parasitic Indigobirds
Distributed across sub-Saharan Africa, ten species of brood parasitic indigobirds reproduce by laying their eggs in the nests of other birds.
Because both male and female indigobirds learn host songs, populations associated with different hosts are behaviourally isolated from one another, allowing for rapid speciation whenever a new host is colonized. We used genetic parentage analyses to test this model for two indigobird host races, recently derived and morphologically indistinguishable populations that mimic the songs of different hosts.
Perhaps due to imperfect fidelity in host choice by the female parasites, reproductive isolation between the two host races is incomplete, suggesting that divergent natural or sexual selection is needed to complete the speciation process in indigobirds.
Proceedings of the Royal Society B: Biological Sciences
Proceedings B is the Royal Society's flagship biological research journal, dedicated to the rapid publication and broad dissemination of high-quality research papers, reviews and comment and reply papers. The scope of journal is diverse and is especially strong in organismal biology.
publishing.royalsociety/proceedingsb
Because both male and female indigobirds learn host songs, populations associated with different hosts are behaviourally isolated from one another, allowing for rapid speciation whenever a new host is colonized. We used genetic parentage analyses to test this model for two indigobird host races, recently derived and morphologically indistinguishable populations that mimic the songs of different hosts.
Perhaps due to imperfect fidelity in host choice by the female parasites, reproductive isolation between the two host races is incomplete, suggesting that divergent natural or sexual selection is needed to complete the speciation process in indigobirds.
Proceedings of the Royal Society B: Biological Sciences
Proceedings B is the Royal Society's flagship biological research journal, dedicated to the rapid publication and broad dissemination of high-quality research papers, reviews and comment and reply papers. The scope of journal is diverse and is especially strong in organismal biology.
publishing.royalsociety/proceedingsb
Potential Treatment For Bone Death In The Hip From Osteonecrosis
Researchers at Mount Sinai School of Medicine have found a potential new treatment for osteonecrosis, or the death of bone tissue, in people who are treated with steroids for several common medical conditions. There are currently no treatment options for people with this debilitating disease. The research is published in the April 27 issue of Proceedings of the National Academy of Sciences.
Glucocorticoids are a class of steroids used to treat several common diseases, including asthma, ulcerative colitis, kidney diseases, and rheumatologic disorders. These steroids cause bone loss, and can eventually cause severe osteoporosis and fracture, as well as osteonecrosis. The Mount Sinai team, led by Mone Zaidi, MD, PhD, FRCP, Professor of Medicine and Physiology and Director of The Mount Sinai Bone Program at Mount Sinai School of Medicine, discovered that injecting the naturally-produced hormone adrenocorticotropic hormone (ACTH) in rabbits with osteonecrosis caused by treatment with glucocorticoids significantly reduced bone death in the hip.
"Osteonecrosis is a very painful condition that has the potential to affect hundreds of thousands of Americans who are treated with steroids, with no treatment option until now except hip replacement," said Dr. Zaidi. "Our research is the first to show the therapeutic benefit of ACTH in experimental osteonecrosis, providing the first treatment option for these patients."
Glucocorticoids cause reduced blood flow to bone cells in the hip, resulting in cell death, and ACTH reduces these devastating side effects. However, research indicates that osteonecrosis is not significant in people in which steroid levels are high in the blood. Dr. Zaidi's team knew that these tumors produce excess ACTH, and this spurred the team to evaluate the ACTH's potential therapeutic effect.
The researchers injected one group of rabbits with depomedrol, a type of steroid, and another group with depomedrol plus ACTH. Osteonecrosis was dramatically reduced in the rabbits that were treated with ACTH. Dr. Zaidi's team found that ACTH stimulates the vascular endothelial growth factor (VEGF), a protein that signals for the growth of new blood vessels. The stimulation of VEGF results in increased blood flow to the bone cells, preventing cell death.
"The results confirm that ACTH may be of value as a drug to prevent osteonecrosis," said Dr. Zaidi. "While more research is required, we hope to someday evaluate the efficacy of ACTH in treating osteoporosis as well."
Source:
Mount Sinai Press Office
The Mount Sinai Hospital / Mount Sinai School of Medicine
Glucocorticoids are a class of steroids used to treat several common diseases, including asthma, ulcerative colitis, kidney diseases, and rheumatologic disorders. These steroids cause bone loss, and can eventually cause severe osteoporosis and fracture, as well as osteonecrosis. The Mount Sinai team, led by Mone Zaidi, MD, PhD, FRCP, Professor of Medicine and Physiology and Director of The Mount Sinai Bone Program at Mount Sinai School of Medicine, discovered that injecting the naturally-produced hormone adrenocorticotropic hormone (ACTH) in rabbits with osteonecrosis caused by treatment with glucocorticoids significantly reduced bone death in the hip.
"Osteonecrosis is a very painful condition that has the potential to affect hundreds of thousands of Americans who are treated with steroids, with no treatment option until now except hip replacement," said Dr. Zaidi. "Our research is the first to show the therapeutic benefit of ACTH in experimental osteonecrosis, providing the first treatment option for these patients."
Glucocorticoids cause reduced blood flow to bone cells in the hip, resulting in cell death, and ACTH reduces these devastating side effects. However, research indicates that osteonecrosis is not significant in people in which steroid levels are high in the blood. Dr. Zaidi's team knew that these tumors produce excess ACTH, and this spurred the team to evaluate the ACTH's potential therapeutic effect.
The researchers injected one group of rabbits with depomedrol, a type of steroid, and another group with depomedrol plus ACTH. Osteonecrosis was dramatically reduced in the rabbits that were treated with ACTH. Dr. Zaidi's team found that ACTH stimulates the vascular endothelial growth factor (VEGF), a protein that signals for the growth of new blood vessels. The stimulation of VEGF results in increased blood flow to the bone cells, preventing cell death.
"The results confirm that ACTH may be of value as a drug to prevent osteonecrosis," said Dr. Zaidi. "While more research is required, we hope to someday evaluate the efficacy of ACTH in treating osteoporosis as well."
Source:
Mount Sinai Press Office
The Mount Sinai Hospital / Mount Sinai School of Medicine
'2-Faced' Particles Act Like Tiny Submarines
For the first time, researchers at North Carolina State University have demonstrated that microscopic "two-faced" spheres whose halves are physically or chemically different - so-called Janus particles - will move like stealthy submarines when an alternating electrical field is applied to liquid surrounding the particles.
A paper describing the research, published in the Feb. 8, 2008, edition of Physical Review Letters, advances knowledge about how potential "smart" materials - think of tiny engines or sensors - can move around and respond to changes in their environment. Janus particles could be used as microscopic mixers, molecular "shuttles," self-propelling microsensors or means of targeted drug delivery.
The researchers - Dr. Orlin Velev, associate professor of chemical and biomolecular engineering at NC State and lead author of the paper; Sumit Gangwal, an NC State graduate student; Dr. Olivier Cayre, a post-doctoral researcher in Velev's lab; and Dr. Martin Bazant from Massachusetts Institute of Technology - created tiny two-faced gold and plastic particles and applied low frequency alternating current to the water containing the particles. The electric field was of voltage and frequency similar to the ones you'd get if you plugged a device into a socket in your home or office.
Velev says the micrometer-sized particles convert the electrical field into liquid motion around them and then unexpectedly propel themselves perpendicular to the direction of the powered electrodes - not in the direction of the electrical field, as would be expected. The particles always travel in the same orientation: with the plastic "face" as the front of the mini-submarine and the metallic "face" in the rear, Velev added.
The phenomenon - called "induced-charge electrophoresis," which had been predicted in a theoretical model by the MIT collaborator - had not been demonstrated previously.
The term "Janus particle" comes from the name of a Roman god with two faces. Velev says that these materials have the potential to perform a variety of applications.
"You can imagine other types of Janus particles comprising a 'smart gel' that responds to a change in its environment and then releases drugs, for example," Velev says. Fabricating these responsive materials on the microscale and nanoscale is an exciting and rapidly developing area of science, he adds.
"We are able to create tiny Janus particles of the same size and shape and are beginning to learn how to give them functionality," Velev said. "The next step is to create more complex particles that are able to perform more specialized functions in addition to propelling themselves around."
The research is funded by the National Science Foundation and a Camile and Henry Dreyfus Teacher-Scholar grant.
The abstract of the paper follows.
"Induced Charge Electrophoresis of Metallodielectric Particles"
Authors: Sumit Gangwal, Olivier J. Cayre and Dr. Orlin D. Velev, NC State University; Dr. Martin Z. Bazant,
Massachusetts Institute of Technology
Published: Feb. 4, 2008, in Physical Review Letters
Abstract: The application of ac electric fields in aqueous suspensions of anisotropic particles leads to unbalanced liquid flows and nonlinear, induced-charge electrophoretic motion. We report experimental observations of the motion of Janus microparticles with one dielectric and one metal-coated hemisphere induced by uniform fields of frequency 100Hz-10kHz in NaCl solutions. The motion is perpendicular to the field axis and persists after particles are attracted to a glass wall. This phenomenon may find application in microactuators, microsensors and microfluidic devices.
Source: Dr. Orlin Velev
North Carolina State University
A paper describing the research, published in the Feb. 8, 2008, edition of Physical Review Letters, advances knowledge about how potential "smart" materials - think of tiny engines or sensors - can move around and respond to changes in their environment. Janus particles could be used as microscopic mixers, molecular "shuttles," self-propelling microsensors or means of targeted drug delivery.
The researchers - Dr. Orlin Velev, associate professor of chemical and biomolecular engineering at NC State and lead author of the paper; Sumit Gangwal, an NC State graduate student; Dr. Olivier Cayre, a post-doctoral researcher in Velev's lab; and Dr. Martin Bazant from Massachusetts Institute of Technology - created tiny two-faced gold and plastic particles and applied low frequency alternating current to the water containing the particles. The electric field was of voltage and frequency similar to the ones you'd get if you plugged a device into a socket in your home or office.
Velev says the micrometer-sized particles convert the electrical field into liquid motion around them and then unexpectedly propel themselves perpendicular to the direction of the powered electrodes - not in the direction of the electrical field, as would be expected. The particles always travel in the same orientation: with the plastic "face" as the front of the mini-submarine and the metallic "face" in the rear, Velev added.
The phenomenon - called "induced-charge electrophoresis," which had been predicted in a theoretical model by the MIT collaborator - had not been demonstrated previously.
The term "Janus particle" comes from the name of a Roman god with two faces. Velev says that these materials have the potential to perform a variety of applications.
"You can imagine other types of Janus particles comprising a 'smart gel' that responds to a change in its environment and then releases drugs, for example," Velev says. Fabricating these responsive materials on the microscale and nanoscale is an exciting and rapidly developing area of science, he adds.
"We are able to create tiny Janus particles of the same size and shape and are beginning to learn how to give them functionality," Velev said. "The next step is to create more complex particles that are able to perform more specialized functions in addition to propelling themselves around."
The research is funded by the National Science Foundation and a Camile and Henry Dreyfus Teacher-Scholar grant.
The abstract of the paper follows.
"Induced Charge Electrophoresis of Metallodielectric Particles"
Authors: Sumit Gangwal, Olivier J. Cayre and Dr. Orlin D. Velev, NC State University; Dr. Martin Z. Bazant,
Massachusetts Institute of Technology
Published: Feb. 4, 2008, in Physical Review Letters
Abstract: The application of ac electric fields in aqueous suspensions of anisotropic particles leads to unbalanced liquid flows and nonlinear, induced-charge electrophoretic motion. We report experimental observations of the motion of Janus microparticles with one dielectric and one metal-coated hemisphere induced by uniform fields of frequency 100Hz-10kHz in NaCl solutions. The motion is perpendicular to the field axis and persists after particles are attracted to a glass wall. This phenomenon may find application in microactuators, microsensors and microfluidic devices.
Source: Dr. Orlin Velev
North Carolina State University
2008 Joint Biophysical Society Annual Meeting And IUPAB International Biophysics Congress Awards Ceremony
The Biophysical Society is pleased to announce the recipients of its 2008 Society awards. The fourteen recipients will receive their awards at the Joint Biophysical Society Annual Meeting and IUPAB International Biophysics Congress Awards Ceremony on Monday February 4, 2008 at the Convention Center in Long Beach, California. The awardees are:
Ben de Kruijff of Utrecht University will receive the Avanti Award in Lipids for his excellent and high-impact contributions to the field of lipids and membrane biology;
Robert Callender of Albert Einstein College of Medicine will receive the Distinguished Service Award for his service and remarkable commitment to the Biophysical Society Journal during his tenure as Editor-in-Chief;
David S. Eisenberg of the University of California, Los Angeles and Donald M. Crothers of Yale Univeristy will share the Emily M. Gray Award for their significant contributions to education through creating rigorous, ground-breaking text enriching generations of biophysicists;
Peter G. Wolynes of the Univeristy of California, San Diego will receive the Founders Award for his exceptional intellectual contributions in advancing biophysical theory and physical sciences.
Sergei Sukharev of the University of Maryland will receive the Michael and Kate Barany Award for Young Investigators for his outstanding and creative contributions to membrane biophysics;
Steven M. Block of Stanford University will receive the U.S. Genomics Award for Outstanding Investigator in the Field of Single Molecule Biology for his contributions, leadership, and creativity in advancing the field of single molecule biology;
Judith Klein-Seetharaman of the University of Pittsburgh School of Medicine will receive the Margaret Oakley Dayhoff Award for her remarkable work in computational biology embracing the full spectrum of experimental biophysics. This award is given to a junior woman scientist of promise in the field of biophysics, who has not yet reached a position of high recognition within the structures of academic society; and
H. Ronald Kaback of the University of California, Los Angeles will receive the Anatrace Membrane Protein Award for his outstanding contributions to unraveling the structure and mechanism of action of E. coli lactose permease.
In addition, five Biophysical Society members have been named to the 2007 class of Society Fellows. They are:
Timothy A. Cross of Florida State University for his scientific accomplishments and leadership in the field of solid state NMR methods to the biophysical characterization of membrane proteins, and for service to the scientific community;
Eve E. Marder of Brandeis University for her seminal discoveries in the field of neuroscience and elegantly combining both experiment and theoretical work in an innovative manner to advance the field of neurobiology;
Ivan Rayment of the University of Wisconsin for his work in protein crystallography demonstrating the essential function it plays in the modern biophysics;
Stephen G. Sligar of the University of Illinois, Urbana-Champaign for advancing our knowledge of biological functions through the concerted application of numerous biophysical methods; and
Attila Szabo of NIDDK, NIH for developing novel theoretical analyses for a wide variety of experiments and bringing leadership to the service of biological physics.
The Biophysical Society, founded in 1956, is a professional, scientific society established to encourage development and dissemination of knowledge in biophysics. The Society promotes growth in this expanding field through its annual meeting, monthly journal, and committee and outreach activities. Its nearly 8000 members are located throughout the U.S. and the world, where they teach and conduct research in colleges, universities, laboratories, government agencies, and industry. For more information on the society or the 2007 annual meeting, visit biophysics/.
Source: Ellen R. Weiss
Biophysical Society
Ben de Kruijff of Utrecht University will receive the Avanti Award in Lipids for his excellent and high-impact contributions to the field of lipids and membrane biology;
Robert Callender of Albert Einstein College of Medicine will receive the Distinguished Service Award for his service and remarkable commitment to the Biophysical Society Journal during his tenure as Editor-in-Chief;
David S. Eisenberg of the University of California, Los Angeles and Donald M. Crothers of Yale Univeristy will share the Emily M. Gray Award for their significant contributions to education through creating rigorous, ground-breaking text enriching generations of biophysicists;
Peter G. Wolynes of the Univeristy of California, San Diego will receive the Founders Award for his exceptional intellectual contributions in advancing biophysical theory and physical sciences.
Sergei Sukharev of the University of Maryland will receive the Michael and Kate Barany Award for Young Investigators for his outstanding and creative contributions to membrane biophysics;
Steven M. Block of Stanford University will receive the U.S. Genomics Award for Outstanding Investigator in the Field of Single Molecule Biology for his contributions, leadership, and creativity in advancing the field of single molecule biology;
Judith Klein-Seetharaman of the University of Pittsburgh School of Medicine will receive the Margaret Oakley Dayhoff Award for her remarkable work in computational biology embracing the full spectrum of experimental biophysics. This award is given to a junior woman scientist of promise in the field of biophysics, who has not yet reached a position of high recognition within the structures of academic society; and
H. Ronald Kaback of the University of California, Los Angeles will receive the Anatrace Membrane Protein Award for his outstanding contributions to unraveling the structure and mechanism of action of E. coli lactose permease.
In addition, five Biophysical Society members have been named to the 2007 class of Society Fellows. They are:
Timothy A. Cross of Florida State University for his scientific accomplishments and leadership in the field of solid state NMR methods to the biophysical characterization of membrane proteins, and for service to the scientific community;
Eve E. Marder of Brandeis University for her seminal discoveries in the field of neuroscience and elegantly combining both experiment and theoretical work in an innovative manner to advance the field of neurobiology;
Ivan Rayment of the University of Wisconsin for his work in protein crystallography demonstrating the essential function it plays in the modern biophysics;
Stephen G. Sligar of the University of Illinois, Urbana-Champaign for advancing our knowledge of biological functions through the concerted application of numerous biophysical methods; and
Attila Szabo of NIDDK, NIH for developing novel theoretical analyses for a wide variety of experiments and bringing leadership to the service of biological physics.
The Biophysical Society, founded in 1956, is a professional, scientific society established to encourage development and dissemination of knowledge in biophysics. The Society promotes growth in this expanding field through its annual meeting, monthly journal, and committee and outreach activities. Its nearly 8000 members are located throughout the U.S. and the world, where they teach and conduct research in colleges, universities, laboratories, government agencies, and industry. For more information on the society or the 2007 annual meeting, visit biophysics/.
Source: Ellen R. Weiss
Biophysical Society
Argonne's Joachimiak And Rosenbaum Honored With The 2007 Compton Award
The Department of Energy's Advanced Photon Source (APS) and the APS Users Organization have announced that the 2007 Arthur H. Compton Award will be presented jointly to Andrzej Joachimiak and Gerold Rosenbaum of Argonne National Laboratory for pioneering advances and leadership that helped to establish the APS as a premier location worldwide for protein crystallography research.
"Andrzej and Gerd were nominated for the Compton Award because of their individual, key contributions to protein crystallography research at the APS. The award underscores the worldwide stature of protein crystallography conducted at the APS and the importance of this field of research," said Murray Gibson, Argonne associate laboratory director for scientific user facilities. "It is a reflection of the quality of their work, dedication and the major role they both play in protein structure determination."
Andrzej Joachimiak is recognized both for his talents as a prolific crystallographer and methodological innovator working with difficult structures. He has been instrumental in establishing structural genomics as a part of modern biology and serves as director of both the Structural Biology Center (SBC) and the Midwest Center for Structural Genomics (MCSG), each within the Biosciences Division at Argonne.
Gerold Rosenbaum is recognized for his pioneering demonstration in 1970 that synchrotron radiation could be a source for biological X-ray diffraction as well as his innovative designs, that have set the world standard for biological diffraction. Currently Rosenbaum is a senior beamline scientist at the Southeast Regional Collaborative Access Team, operated at the APS by the University of Georgia, where he holds an appointment with the Department of Biochemistry and Molecular Biology.
The award was presented at the first science session of 2007 Users Week at Argonne.
The Arthur H. Compton award was established in 1995 by the APS Users Organization to recognize an important scientific or technical accomplishment at, or beneficial to, the Advanced Photon Source.
Compton was an American physicist who won the Nobel Prize for Physics in 1927 for discovering and explaining changes in x-ray wavelengths resulting from x-ray collisions with electrons, the so-called Compton effect. This important discovery in 1922 confirmed the dual nature (wave and particle) of electromagnetic radiation.
About Argonne
The nation's first national laboratory, Argonne conducts basic and applied scientific research across a wide spectrum of disciplines, ranging from high-energy physics to climatology and biotechnology. It is the home of the Advanced Photon Source which is the Western Hemisphere's most powerful source of x-rays for research. More than 3,000 users from industry, academia and government laboratories around the world use the APS each year for research in materials science, chemistry, biology, physics, earth and planetary science, and environmental science. Since 1990, Argonne has worked with more than 600 companies and numerous federal agencies and other organizations to help advance America's scientific leadership and prepare the nation for the future. Argonne is managed by UChicago Argonne, LLC for the U.S. Department of Energy's Office of Science.
Contact: Eleanor Taylor
DOE/Argonne National Laboratory
"Andrzej and Gerd were nominated for the Compton Award because of their individual, key contributions to protein crystallography research at the APS. The award underscores the worldwide stature of protein crystallography conducted at the APS and the importance of this field of research," said Murray Gibson, Argonne associate laboratory director for scientific user facilities. "It is a reflection of the quality of their work, dedication and the major role they both play in protein structure determination."
Andrzej Joachimiak is recognized both for his talents as a prolific crystallographer and methodological innovator working with difficult structures. He has been instrumental in establishing structural genomics as a part of modern biology and serves as director of both the Structural Biology Center (SBC) and the Midwest Center for Structural Genomics (MCSG), each within the Biosciences Division at Argonne.
Gerold Rosenbaum is recognized for his pioneering demonstration in 1970 that synchrotron radiation could be a source for biological X-ray diffraction as well as his innovative designs, that have set the world standard for biological diffraction. Currently Rosenbaum is a senior beamline scientist at the Southeast Regional Collaborative Access Team, operated at the APS by the University of Georgia, where he holds an appointment with the Department of Biochemistry and Molecular Biology.
The award was presented at the first science session of 2007 Users Week at Argonne.
The Arthur H. Compton award was established in 1995 by the APS Users Organization to recognize an important scientific or technical accomplishment at, or beneficial to, the Advanced Photon Source.
Compton was an American physicist who won the Nobel Prize for Physics in 1927 for discovering and explaining changes in x-ray wavelengths resulting from x-ray collisions with electrons, the so-called Compton effect. This important discovery in 1922 confirmed the dual nature (wave and particle) of electromagnetic radiation.
About Argonne
The nation's first national laboratory, Argonne conducts basic and applied scientific research across a wide spectrum of disciplines, ranging from high-energy physics to climatology and biotechnology. It is the home of the Advanced Photon Source which is the Western Hemisphere's most powerful source of x-rays for research. More than 3,000 users from industry, academia and government laboratories around the world use the APS each year for research in materials science, chemistry, biology, physics, earth and planetary science, and environmental science. Since 1990, Argonne has worked with more than 600 companies and numerous federal agencies and other organizations to help advance America's scientific leadership and prepare the nation for the future. Argonne is managed by UChicago Argonne, LLC for the U.S. Department of Energy's Office of Science.
Contact: Eleanor Taylor
DOE/Argonne National Laboratory
New Imaging Technologies To Stop A Deadly Killer
To develop new strategies to control tuberculosis (TB), a contagious disease that infects one-third of the world's population and kills almost two million people every year, the University of Pittsburgh Center for Vaccine Research has received an $11.4 million grant from the Bill & Melinda Gates Foundation. The grant will enable Pitt researchers to use new imaging technologies to study TB to shorten and simplify its course of treatment, potentially improving survival and curtailing the global TB epidemic.
"One of the most challenging issues in treating TB and stopping its spread is the length of time it takes to adequately stem the infection," said JoAnne Flynn, Ph.D., principal investigator of the grant and professor of microbiology and molecular genetics, University of Pittsburgh School of Medicine. "Current drugs are available, but we don't fully understand how or why they work. TB treatment must be continued for at least six months to be effective, placing an undue burden on those who are infected - often from the poorest and most disadvantaged countries."
According to Dr. Flynn, TB is difficult to control because the germs that cause the infection hide from the immune system in small tissue nodules called granulomas, enabling the infection to reactivate years, and even decades, later. Although for the most part TB is a curable disease, patients must adhere to treatment long after symptoms have faded. This proves challenging in many regions of the world where medication is not readily accessible. Indeed, an inadequate or incomplete course of treatment is the major factor that causes drug-resistant TB strains to develop. These strains are alarmingly high in many countries around the world.
"Current medications for TB were developed more than three decades ago," said Dr. Flynn. "To create significantly shorter and simplified approaches to treatment, we must improve our understanding of this disease and how current drugs are localized at the site of infection."
To understand more about the basic biology of TB, Dr. Flynn and colleagues are using the grant to develop positron emission tomography (PET) and computed tomography (CT) imaging studies in non-human primates. By using combined PET/CT, the researchers will be able to follow the progression of the disease in animals over time and analyze changes in tissue and responses to particular drugs. They will be using three imaging technologies - radionuclides, fluorescence and mass spectrometry - in combination to develop imaging probes and techniques to precisely locate bacteria associated with TB and to explore the underlying factors responsible for slow drug metabolism.
"By applying the tools of modern medicine to TB, we hope to lay the groundwork for real-time measurements of TB drug efficacy in clinical trials and develop new targeted therapies that will considerably shorten the length of treatment," said Dr. Flynn.
Tuberculosis is a bacterial disease usually affecting the lungs. Called pulmonary TB, the disease is characterized by a persistent cough, shortness of breath, weight loss and chest pain. Left untreated, one person with active pulmonary TB will infect on average between 10 and 15 other people every year. The bacteria associated with the disease also can infect nearly any part of the body, such as the lymph nodes, the spine or bones. TB is deadly if left untreated.
Co-investigators on the grant include Clifton Barry III, Ph.D., National Institute of Allergy and Infectious Diseases; Richard Caprioli, Ph.D., and Michelle Reyzer, Ph.D., Vanderbilt University; David Russell, Ph.D., and Warren Zipfel, Ph.D., Cornell University; Kim Janda, Ph.D., and Tobin Dickerson, Ph.D., The Scripps Research Institute; Benjamin Davis, Ph.D., Oxford University; Chet Mathis, Ph.D., Jonathan P. Carney, Ph.D., and Brian J. Lopresti, B.S., University of Pittsburgh; and Veronique Dartois, Ph.D., Novartis Institute of Tropical Disease.
The Center for Vaccine Research (CVR) at the University of Pittsburgh houses both the Regional Biocontainment Laboratory and the Vaccine Research Laboratory. Researchers at the CVR, directed by Donald S. Burke, M.D., dean of the University of Pittsburgh Graduate School of Public Health and UPMC Jonas Salk Professor of Global Health, develop new methods and strategies to prevent and treat infectious diseases, potentially improving and protecting global health.
Source: Clare Collins
University of Pittsburgh Schools of the Health Sciences
"One of the most challenging issues in treating TB and stopping its spread is the length of time it takes to adequately stem the infection," said JoAnne Flynn, Ph.D., principal investigator of the grant and professor of microbiology and molecular genetics, University of Pittsburgh School of Medicine. "Current drugs are available, but we don't fully understand how or why they work. TB treatment must be continued for at least six months to be effective, placing an undue burden on those who are infected - often from the poorest and most disadvantaged countries."
According to Dr. Flynn, TB is difficult to control because the germs that cause the infection hide from the immune system in small tissue nodules called granulomas, enabling the infection to reactivate years, and even decades, later. Although for the most part TB is a curable disease, patients must adhere to treatment long after symptoms have faded. This proves challenging in many regions of the world where medication is not readily accessible. Indeed, an inadequate or incomplete course of treatment is the major factor that causes drug-resistant TB strains to develop. These strains are alarmingly high in many countries around the world.
"Current medications for TB were developed more than three decades ago," said Dr. Flynn. "To create significantly shorter and simplified approaches to treatment, we must improve our understanding of this disease and how current drugs are localized at the site of infection."
To understand more about the basic biology of TB, Dr. Flynn and colleagues are using the grant to develop positron emission tomography (PET) and computed tomography (CT) imaging studies in non-human primates. By using combined PET/CT, the researchers will be able to follow the progression of the disease in animals over time and analyze changes in tissue and responses to particular drugs. They will be using three imaging technologies - radionuclides, fluorescence and mass spectrometry - in combination to develop imaging probes and techniques to precisely locate bacteria associated with TB and to explore the underlying factors responsible for slow drug metabolism.
"By applying the tools of modern medicine to TB, we hope to lay the groundwork for real-time measurements of TB drug efficacy in clinical trials and develop new targeted therapies that will considerably shorten the length of treatment," said Dr. Flynn.
Tuberculosis is a bacterial disease usually affecting the lungs. Called pulmonary TB, the disease is characterized by a persistent cough, shortness of breath, weight loss and chest pain. Left untreated, one person with active pulmonary TB will infect on average between 10 and 15 other people every year. The bacteria associated with the disease also can infect nearly any part of the body, such as the lymph nodes, the spine or bones. TB is deadly if left untreated.
Co-investigators on the grant include Clifton Barry III, Ph.D., National Institute of Allergy and Infectious Diseases; Richard Caprioli, Ph.D., and Michelle Reyzer, Ph.D., Vanderbilt University; David Russell, Ph.D., and Warren Zipfel, Ph.D., Cornell University; Kim Janda, Ph.D., and Tobin Dickerson, Ph.D., The Scripps Research Institute; Benjamin Davis, Ph.D., Oxford University; Chet Mathis, Ph.D., Jonathan P. Carney, Ph.D., and Brian J. Lopresti, B.S., University of Pittsburgh; and Veronique Dartois, Ph.D., Novartis Institute of Tropical Disease.
The Center for Vaccine Research (CVR) at the University of Pittsburgh houses both the Regional Biocontainment Laboratory and the Vaccine Research Laboratory. Researchers at the CVR, directed by Donald S. Burke, M.D., dean of the University of Pittsburgh Graduate School of Public Health and UPMC Jonas Salk Professor of Global Health, develop new methods and strategies to prevent and treat infectious diseases, potentially improving and protecting global health.
Source: Clare Collins
University of Pittsburgh Schools of the Health Sciences
Scientists Devise Way To Link Complex Traits With Underlying Genes
Princeton University scientists have developed a new way to identify the hidden genetic material responsible for complex traits, a breakthrough they believe ultimately could lead to a deeper understanding of how multiple genes interact to produce everything from blue eyes to blood pressure problems.
Writing in the April 15 edition of Nature, scientists led by Leonid Kruglyak, a professor in Princeton's Department of Ecology and Evolutionary Biology and Lewis-Sigler Institute for Integrative Genomics, report that they developed a straightforward method for studying millions of yeast cells at the same time.
Their method allows them to identify regions of the genome that cause a specific trait in the offspring of two yeast strains that have been mated. In using such a large group, the scientists have been able to identify subtle patterns that could not be detected before.
"One of the important insights gained from research enabled by the sequencing of the human genome is that, rather than being obvious, the connections between genes and most traits are very complicated," Kruglyak said. "Our results show, however, that it is possible to identify many of the factors underlying complex traits using straightforward techniques."
The Princeton team's finding could help illuminate the answers to the current difficulties inherent in tying traits to genes, known as the "missing heritability problem," Kruglyak said.
There are some cases, he said, where scientists have identified mutations in single genes that produce a specific trait, such as a susceptibility to cystic fibrosis or Huntington's disease. In most cases, however, scientists believe that large numbers of genes working in concert produce trait variation. Some genes play a major role while others are more "quiet" but still are important. Scientists want to know all of the genes involved in producing a given complex trait, but they have not been able to find these groupings, leading to the "missing" problem.
Kruglyak was part of an expert panel the National Human Genome Research Institute convened last year on the missing heritability problem. When the Human Genome Project was completed in 2003, it provided an entire sequence of human DNA. The panel discussions centered upon the idea that, despite major technology advances made possible by the project and studies of tens of thousands of individuals, the great majority of the genetic factors responsible for differences between individuals have not yet been found.
"In many cases, the effects of genes are so small that detecting them is extremely difficult," said Ian Ehrenreich, a postdoctoral research fellow who is the first author on the Nature paper. "Under conventional methods, we just don't have the power to identify many of these genes. We knew we had to find a different way."
The method described in the paper is "a creative adaptation of existing family-based, genome-wide methodology," said Yin Yao, who is chief of the molecular and genetic epidemiology program in the division of neuroscience and basic behavioral science at the National Institute of Mental Health. She and Thomas Lehner, chief of the genomic research branch at the institute, said Kruglyak's work is highly valued and described him as a pioneer in the field of statistical genomics.
Scientists in Kruglyak's lab conduct experiments on organisms like yeast, as well as perform computational analyses, aimed at understanding how changes in DNA are shaped by molecular and evolutionary forces. They also study how these changes lead to the observable differences among individuals within a species. For this research project, the team looked to develop a process that would identify genetic associations with observable traits.
"We know in the human genome there are 20,000 genes, but I can't ask someone to point out to me which genes account for most of the variation in human height, for example, because we just don't know," Kruglyak said. "The underlying goal of what we are trying to do is both understand how complicated these patterns are and try to come up with some concrete examples where we can take some traits and nail down most of the variations, as opposed to only finding a small percentage."
Studies in model organisms like yeast -- just as in humans -- have failed to detect a large fraction of the genes believed to underlie most complex traits. So Kruglyak and his team developed a method using a sample size of yeast that went beyond the scale of any human studies. They crossed two strains of yeast, generating about 10 million offspring. Each of these progeny was genetically distinct as opposed to being a clone.
To find a subset offspring that shared a trait, the scientists grew the progeny on a chemical that causes breaks in DNA, killing most of them. They then sequenced the genomes of the few thousand yeast that survived, looking to see what genes they inherited from each parent.
Mendel's laws, which explain the principles of heredity, state that most of the genetic material should be randomly inherited from one or the other parent in a manner equivalent to a coin flip - half of the cells should have one parent's genes and half the other parent's. But, at the locations of the genes that protect yeast from the chemical, most of the cells should have genetic material from the parent with the version of the gene that produces greater resistance.
The surviving yeast cells' genomes were placed on tiny chips and scanned on automated laboratory machines, a process known as genotyping. The machines surveyed each yeast cell's genome for strategically selected markers of genetic variation. They found certain genetic variations to be significantly more frequent in the surviving yeast, serving as a powerful pointer to the regions of the genome where the genes underlying the trait resided.
The scientists repeated this experiment with other chemicals that were toxic to most of the yeast, looking again and again for skewed genetic patterns of inheritance. Each time, they were able to locate gene regions pointing to specific traits, confirming that the method worked.
Viewing their success, Ehrenreich said, "It's really been a combination of having the technology to do this genotyping precisely and also being able to survey such a large number of individuals simultaneously."
From here, the team intends to use the information it has on gene regions and markers to fine tune the method and identify the specific genes associated with each trait, and to extend the method to many other yeast strains.
Detlef Weigel, director of the Max Planck Institute for Developmental Biology in Germany, already sees additional applications for the technique. "The new work by Dr. Kruglyak and colleagues beautifully showcases how new sequencing technologies are revolutionizing genetics," he said. "While the work was carried out with yeast, I am convinced that it can be easily extended to any other genetically tractable organism, including crop plants."
Other Princeton scientists on the paper were: Noorossadat Torabi, a graduate student; Amy Caudy, a Lewis-Sigler fellow; Joshua Shapiro, a postdoctoral research fellow; Yue Jia, a research specialist; and Jonathan Kent and Stephen Martis, undergraduate students. Another author, David Gresham, who participated in the effort as a postdoctoral research fellow, is now at the Center for Genomics and Systems Biology at New York University.
The research was supported by the National Institutes of Health, a James S. McDonnell Centennial Fellowship and the Howard Hughes Medical Institute.
Source:
Kitta MacPherson
Princeton University
Writing in the April 15 edition of Nature, scientists led by Leonid Kruglyak, a professor in Princeton's Department of Ecology and Evolutionary Biology and Lewis-Sigler Institute for Integrative Genomics, report that they developed a straightforward method for studying millions of yeast cells at the same time.
Their method allows them to identify regions of the genome that cause a specific trait in the offspring of two yeast strains that have been mated. In using such a large group, the scientists have been able to identify subtle patterns that could not be detected before.
"One of the important insights gained from research enabled by the sequencing of the human genome is that, rather than being obvious, the connections between genes and most traits are very complicated," Kruglyak said. "Our results show, however, that it is possible to identify many of the factors underlying complex traits using straightforward techniques."
The Princeton team's finding could help illuminate the answers to the current difficulties inherent in tying traits to genes, known as the "missing heritability problem," Kruglyak said.
There are some cases, he said, where scientists have identified mutations in single genes that produce a specific trait, such as a susceptibility to cystic fibrosis or Huntington's disease. In most cases, however, scientists believe that large numbers of genes working in concert produce trait variation. Some genes play a major role while others are more "quiet" but still are important. Scientists want to know all of the genes involved in producing a given complex trait, but they have not been able to find these groupings, leading to the "missing" problem.
Kruglyak was part of an expert panel the National Human Genome Research Institute convened last year on the missing heritability problem. When the Human Genome Project was completed in 2003, it provided an entire sequence of human DNA. The panel discussions centered upon the idea that, despite major technology advances made possible by the project and studies of tens of thousands of individuals, the great majority of the genetic factors responsible for differences between individuals have not yet been found.
"In many cases, the effects of genes are so small that detecting them is extremely difficult," said Ian Ehrenreich, a postdoctoral research fellow who is the first author on the Nature paper. "Under conventional methods, we just don't have the power to identify many of these genes. We knew we had to find a different way."
The method described in the paper is "a creative adaptation of existing family-based, genome-wide methodology," said Yin Yao, who is chief of the molecular and genetic epidemiology program in the division of neuroscience and basic behavioral science at the National Institute of Mental Health. She and Thomas Lehner, chief of the genomic research branch at the institute, said Kruglyak's work is highly valued and described him as a pioneer in the field of statistical genomics.
Scientists in Kruglyak's lab conduct experiments on organisms like yeast, as well as perform computational analyses, aimed at understanding how changes in DNA are shaped by molecular and evolutionary forces. They also study how these changes lead to the observable differences among individuals within a species. For this research project, the team looked to develop a process that would identify genetic associations with observable traits.
"We know in the human genome there are 20,000 genes, but I can't ask someone to point out to me which genes account for most of the variation in human height, for example, because we just don't know," Kruglyak said. "The underlying goal of what we are trying to do is both understand how complicated these patterns are and try to come up with some concrete examples where we can take some traits and nail down most of the variations, as opposed to only finding a small percentage."
Studies in model organisms like yeast -- just as in humans -- have failed to detect a large fraction of the genes believed to underlie most complex traits. So Kruglyak and his team developed a method using a sample size of yeast that went beyond the scale of any human studies. They crossed two strains of yeast, generating about 10 million offspring. Each of these progeny was genetically distinct as opposed to being a clone.
To find a subset offspring that shared a trait, the scientists grew the progeny on a chemical that causes breaks in DNA, killing most of them. They then sequenced the genomes of the few thousand yeast that survived, looking to see what genes they inherited from each parent.
Mendel's laws, which explain the principles of heredity, state that most of the genetic material should be randomly inherited from one or the other parent in a manner equivalent to a coin flip - half of the cells should have one parent's genes and half the other parent's. But, at the locations of the genes that protect yeast from the chemical, most of the cells should have genetic material from the parent with the version of the gene that produces greater resistance.
The surviving yeast cells' genomes were placed on tiny chips and scanned on automated laboratory machines, a process known as genotyping. The machines surveyed each yeast cell's genome for strategically selected markers of genetic variation. They found certain genetic variations to be significantly more frequent in the surviving yeast, serving as a powerful pointer to the regions of the genome where the genes underlying the trait resided.
The scientists repeated this experiment with other chemicals that were toxic to most of the yeast, looking again and again for skewed genetic patterns of inheritance. Each time, they were able to locate gene regions pointing to specific traits, confirming that the method worked.
Viewing their success, Ehrenreich said, "It's really been a combination of having the technology to do this genotyping precisely and also being able to survey such a large number of individuals simultaneously."
From here, the team intends to use the information it has on gene regions and markers to fine tune the method and identify the specific genes associated with each trait, and to extend the method to many other yeast strains.
Detlef Weigel, director of the Max Planck Institute for Developmental Biology in Germany, already sees additional applications for the technique. "The new work by Dr. Kruglyak and colleagues beautifully showcases how new sequencing technologies are revolutionizing genetics," he said. "While the work was carried out with yeast, I am convinced that it can be easily extended to any other genetically tractable organism, including crop plants."
Other Princeton scientists on the paper were: Noorossadat Torabi, a graduate student; Amy Caudy, a Lewis-Sigler fellow; Joshua Shapiro, a postdoctoral research fellow; Yue Jia, a research specialist; and Jonathan Kent and Stephen Martis, undergraduate students. Another author, David Gresham, who participated in the effort as a postdoctoral research fellow, is now at the Center for Genomics and Systems Biology at New York University.
The research was supported by the National Institutes of Health, a James S. McDonnell Centennial Fellowship and the Howard Hughes Medical Institute.
Source:
Kitta MacPherson
Princeton University
Revue De Synthese To Be Published By Springer As Of 2007
Springer will publish Revue de synthese on behalf of the Foundation "pour la science" starting in January 2007. Revue de synthese was created in 1910 by Henri Berr as a respected journal bridging multidisciplinary topics such as philosophy, sociology and history. During the past century, the journal has established itself as an important forum for intellectual debate, drawing interest from authors and readers worldwide.
"I am very proud of the partnership between Springer and the Fondation "pour la science", as it confirms the excellent match between the requirements of a prestigious institution and the objectives of an innovative publisher," said Guido Zosimo-Landolfo, Managing Director of Springer France.
Professor Eric Brian, Director of the Fondation "pour la science" and Editor-in-Chief of the Revue de synthese, added, "Publishing the Revue de synthese with a renowned partner such as Springer is, as Henri Berr said over a century ago, 'to realize the ideal of a publication which is constantly renewing itself and always aware of its effect'. Today, it is important not only to renew the dialogue between disciplines, but also to revitalize the way scholars communicate among each other."
Springer will publish the quarterly French-language journal Revue de synthese, with English abstracts, in both print and electronic versions. English-language abstracts are included. As of 2007, it will be available via SpringerLink, Springer's online information platform, and will include Online FirstTM, a feature where articles are published online before they appear in print. In addition, it will offer all authors, via the Springer Open Choice program, the option of publishing their articles using the open access publishing model.
About Foundation "pour la science"
The Fondation "pour la science" was established in 1924 by Henri Berr, and has been officially recognized as an organization operating in the public interest. Its goal is to "develop and coordinate pure scientific research, and thus counteract over-specialization." Its activities focus on new approaches to the historiography, history and philosophy of science, from the era between the two world wars to our own time.
About Springer
Springer is the second-largest publisher worldwide in the science, technology, and medicine (STM) sector. Springer is part of Springer Science+Business Media, one of the world's leading suppliers of scientific and specialist literature. The group owns 70 publishing houses, together publishing a total of 1,450 journals and more than 5,000 new books a year. The group operates in over 20 countries in Europe, the USA, and Asia, and has some 5,000 employees. In 2005, it generated annual sales of around EUR 838 million.
Contact: Joan Robinson
Springer
"I am very proud of the partnership between Springer and the Fondation "pour la science", as it confirms the excellent match between the requirements of a prestigious institution and the objectives of an innovative publisher," said Guido Zosimo-Landolfo, Managing Director of Springer France.
Professor Eric Brian, Director of the Fondation "pour la science" and Editor-in-Chief of the Revue de synthese, added, "Publishing the Revue de synthese with a renowned partner such as Springer is, as Henri Berr said over a century ago, 'to realize the ideal of a publication which is constantly renewing itself and always aware of its effect'. Today, it is important not only to renew the dialogue between disciplines, but also to revitalize the way scholars communicate among each other."
Springer will publish the quarterly French-language journal Revue de synthese, with English abstracts, in both print and electronic versions. English-language abstracts are included. As of 2007, it will be available via SpringerLink, Springer's online information platform, and will include Online FirstTM, a feature where articles are published online before they appear in print. In addition, it will offer all authors, via the Springer Open Choice program, the option of publishing their articles using the open access publishing model.
About Foundation "pour la science"
The Fondation "pour la science" was established in 1924 by Henri Berr, and has been officially recognized as an organization operating in the public interest. Its goal is to "develop and coordinate pure scientific research, and thus counteract over-specialization." Its activities focus on new approaches to the historiography, history and philosophy of science, from the era between the two world wars to our own time.
About Springer
Springer is the second-largest publisher worldwide in the science, technology, and medicine (STM) sector. Springer is part of Springer Science+Business Media, one of the world's leading suppliers of scientific and specialist literature. The group owns 70 publishing houses, together publishing a total of 1,450 journals and more than 5,000 new books a year. The group operates in over 20 countries in Europe, the USA, and Asia, and has some 5,000 employees. In 2005, it generated annual sales of around EUR 838 million.
Contact: Joan Robinson
Springer
Segregating Out UbcH10's Role In Tumor Formation
A ubiquitin-conjugating enzyme that regulates the cell cycle promotes chromosome missegregation and tumor formation, according to van Ree et al. in the January 11 issue of the Journal of Cell Biology.
The mitotic E2 enzyme UbcH10 partners with the anaphase-promoting complex/cyclosome (APC/C) to ubiquitinate cell cycle regulators, targeting them for proteasomal destruction, and ensuring progression through mitosis. UbcH10 is overexpressed in a variety of human cancers, but whether it causes tumors or is simply up-regulated due to the increased number of proliferating cancer cells is unknown.
van Ree et al. generated mice expressing high levels of UbcH10 and found that they formed tumors in a broad range of tissues. Many of these tumors displayed aneuploidy - abnormal numbers of chromosomes resulting from errors in cell division. Live microscopy showed that cells expressing high amounts of UbcH10 had problems segregating sister chromatids correctly, possibly because the cells contained extra numbers of centrosomes that might complicate formation of a normal mitotic spindle. UbcH10 overexpression also reduced levels of the mitotic regulator cyclinB - a substrate of the APC/C - though it remains to be seen if this contributes directly to centrosome amplification and aneuploidy.
The same research group recently demonstrated that chromosome segregation defects drive tumorigenesis by promoting the loss of tumor suppressor genes like p53. Senior author Jan van Deursen now wants to investigate whether UbcH10 synergizes with other factors to promote chromosome instability in human cancers.
Source:
Rita Sullivan
Rockefeller University Press
The mitotic E2 enzyme UbcH10 partners with the anaphase-promoting complex/cyclosome (APC/C) to ubiquitinate cell cycle regulators, targeting them for proteasomal destruction, and ensuring progression through mitosis. UbcH10 is overexpressed in a variety of human cancers, but whether it causes tumors or is simply up-regulated due to the increased number of proliferating cancer cells is unknown.
van Ree et al. generated mice expressing high levels of UbcH10 and found that they formed tumors in a broad range of tissues. Many of these tumors displayed aneuploidy - abnormal numbers of chromosomes resulting from errors in cell division. Live microscopy showed that cells expressing high amounts of UbcH10 had problems segregating sister chromatids correctly, possibly because the cells contained extra numbers of centrosomes that might complicate formation of a normal mitotic spindle. UbcH10 overexpression also reduced levels of the mitotic regulator cyclinB - a substrate of the APC/C - though it remains to be seen if this contributes directly to centrosome amplification and aneuploidy.
The same research group recently demonstrated that chromosome segregation defects drive tumorigenesis by promoting the loss of tumor suppressor genes like p53. Senior author Jan van Deursen now wants to investigate whether UbcH10 synergizes with other factors to promote chromosome instability in human cancers.
Source:
Rita Sullivan
Rockefeller University Press
New Animal Model Furthers Research
Two neuroscientists at the University of Wisconsin-Milwaukee (UWM) are working with local company PhysioGenix to investigate a novel animal model the company has developed for researching diseases like depression, anxiety, schizophrenia and ADHD.
The models are genetically altered rats, originally created by researchers at the Medical College of Wisconsin (MCW) as a way to clone genes related to human diseases. Called consomic rats, they were produced by replacing a single chromosome from the genetic background of a "diseased" rat with the same chromosome from a "normal" rat.
In theory, if the new strain is "cured" of a disease, then the genes responsible for the disease are either on the transferred chromosome or somehow related to it. Consomic rats can be used to rapidly identify new genes and cellular targets associated with certain diseases and to develop and test the efficacy of new drug therapies.
In fact, the consomic rats have already proved useful in studies of cardiovascular disease and hypertension -- the main reason for their development.
The process of identifying which genes are the players in certain complex diseases or behaviors is much quicker using consomic rats than by traditional gene-hunting methods, says Steven Nye, director of genomics at PhysioGenix and principal investigator of the National Institutes of Health grant for commercializing the consomic rats.
PhysioGenix, a spin-off company founded by researchers at MCW, has contracted with UWM Psychology Professors Rodney Swain and Fred Helmstetter to characterize the rats' behaviors in a battery of psychological tests to confirm whether chromosome substitution improves their conditions.
The task is daunting, considering there are 44 strains of consomic rats and potentially hundreds of psychological tests to choose from.
Currently, Swain and Helmstetter are probing their efficacy in identifying the genetic roots of psychological disorders related to learning and memory. Insights from the research could lead to a wide spectrum of related research, he says.
"The impairments that we saw in one consomic strain are similar to some of the symptoms that you see in human children with autism or attention deficit hyperactivity disorder (ADHD)," Swain says.
Test results so far have shown that some of the consomic rats exhibited increased depressive-like behavior, increased pain sensitivity, lower expression of anxiety and enhanced learning in spatial navigation tasks.
The researchers say their findings illustrate the role of genetics on behavior.
"Since my lab has been studying how angiogenesis (the creation of new blood vessels) contributes to better learning, we were very interested to see how this consomic strain performed on a variety of learning tasks," he says.
PhysioGenix, which opened its lab six years ago, licensed the intellectual property from MCW that allows it to commercialize the consomic rats, and the company is finding new applications for them. Howard Jacob, one of the founders of PhysioGenix, is also director of the Human Molecular Genetics Center at MCW. He was instrumental in creating the consomic rats, and was a leader in the rat genome sequencing project funded through the National Institutes of Health. Richard Roman, director of the Kidney Disease Institute at MCW, also is a co-founder of PhysioGenix.
Together, researchers at PhysioGenix, along with their UWM partners, will be presenting three papers at the annual meeting of the Society for Neuroscience in November.
Ultimately, PhysioGenix plans to distribute the consomic rats to academic researchers for studying human neurological diseases and to pharmaceutical researchers for developing new drug therapies.
Milwaukee-based PhysioGenix, Inc. applies the fields of physiology, genetics, genomics and bioinformatics to enhance research models for studying human diseases and for drug development including target validation, drug screening, and predictive toxicology. PhysioGenix provides contract research services to pharmaceutical companies to accelerate their product development processes.
Serving 28,000 students in 12 schools and colleges, UW-Milwaukee offers 155 academic degree programs and is the second largest research university in the state and one of only two doctoral degree-granting institutions in the UW System.
Source: Steven Nye
University of Wisconsin - Milwaukee
The models are genetically altered rats, originally created by researchers at the Medical College of Wisconsin (MCW) as a way to clone genes related to human diseases. Called consomic rats, they were produced by replacing a single chromosome from the genetic background of a "diseased" rat with the same chromosome from a "normal" rat.
In theory, if the new strain is "cured" of a disease, then the genes responsible for the disease are either on the transferred chromosome or somehow related to it. Consomic rats can be used to rapidly identify new genes and cellular targets associated with certain diseases and to develop and test the efficacy of new drug therapies.
In fact, the consomic rats have already proved useful in studies of cardiovascular disease and hypertension -- the main reason for their development.
The process of identifying which genes are the players in certain complex diseases or behaviors is much quicker using consomic rats than by traditional gene-hunting methods, says Steven Nye, director of genomics at PhysioGenix and principal investigator of the National Institutes of Health grant for commercializing the consomic rats.
PhysioGenix, a spin-off company founded by researchers at MCW, has contracted with UWM Psychology Professors Rodney Swain and Fred Helmstetter to characterize the rats' behaviors in a battery of psychological tests to confirm whether chromosome substitution improves their conditions.
The task is daunting, considering there are 44 strains of consomic rats and potentially hundreds of psychological tests to choose from.
Currently, Swain and Helmstetter are probing their efficacy in identifying the genetic roots of psychological disorders related to learning and memory. Insights from the research could lead to a wide spectrum of related research, he says.
"The impairments that we saw in one consomic strain are similar to some of the symptoms that you see in human children with autism or attention deficit hyperactivity disorder (ADHD)," Swain says.
Test results so far have shown that some of the consomic rats exhibited increased depressive-like behavior, increased pain sensitivity, lower expression of anxiety and enhanced learning in spatial navigation tasks.
The researchers say their findings illustrate the role of genetics on behavior.
"Since my lab has been studying how angiogenesis (the creation of new blood vessels) contributes to better learning, we were very interested to see how this consomic strain performed on a variety of learning tasks," he says.
PhysioGenix, which opened its lab six years ago, licensed the intellectual property from MCW that allows it to commercialize the consomic rats, and the company is finding new applications for them. Howard Jacob, one of the founders of PhysioGenix, is also director of the Human Molecular Genetics Center at MCW. He was instrumental in creating the consomic rats, and was a leader in the rat genome sequencing project funded through the National Institutes of Health. Richard Roman, director of the Kidney Disease Institute at MCW, also is a co-founder of PhysioGenix.
Together, researchers at PhysioGenix, along with their UWM partners, will be presenting three papers at the annual meeting of the Society for Neuroscience in November.
Ultimately, PhysioGenix plans to distribute the consomic rats to academic researchers for studying human neurological diseases and to pharmaceutical researchers for developing new drug therapies.
Milwaukee-based PhysioGenix, Inc. applies the fields of physiology, genetics, genomics and bioinformatics to enhance research models for studying human diseases and for drug development including target validation, drug screening, and predictive toxicology. PhysioGenix provides contract research services to pharmaceutical companies to accelerate their product development processes.
Serving 28,000 students in 12 schools and colleges, UW-Milwaukee offers 155 academic degree programs and is the second largest research university in the state and one of only two doctoral degree-granting institutions in the UW System.
Source: Steven Nye
University of Wisconsin - Milwaukee
World's Smallest Periscopes Invented By Vanderbilt Scientists
A team of Vanderbilt scientists have invented the world's smallest version of the periscope and are using it to look at cells and other micro-organisms from several sides at once.
"With an off-the-shelf laboratory microscope you only see cells from one side, the top," says team member Chris Janetopoulos, assistant professor of biological sciences. "Not only can we see the tops of cells, we can view their sides as well - something biologists almost never see."
The researchers have dubbed their devices "mirrored pyramidal wells." As the name implies, they consist of pyramidal-shaped cavities molded into silicon whose interior surfaces are coated with a reflective layer of gold or platinum. They are microscopic in dimension - about the width of a human hair - and can be made in a range of sizes to view different-sized objects. When a cell is placed in such a well and viewed with a regular optical microscope, the researcher can see several sides simultaneously.
"This technology is exciting because these mirrored wells can be made at very low cost, unlike other, more complex methods for 3D microscopy," says Assistant Professor of the Practice of Biomedical Engineering Kevin Seale.
According to Ron Reiserer, "This could easily become as ubiquitous as the microscope slide and could replace more expensive methods currently used to position individual cells." Reiserer is a lab manager at the Vanderbilt Institute for Integrative Biosystems Research and Education (VIIBRE) who helped design the protocol used to make the micropyramids.
The Vanderbilt group is not the first to make microscopic pyramidal wells, but it is the first to apply them to make 3D images of microorganisms. In 2006, a group of scientists in England created pyramidal micromirrors and applied them to trapping atoms. And last spring researchers at the National Institute of Standards and Technology used similar structures to track nanoparticles.
The Vanderbilt researchers reported their achievement last September in the Journal of Microscopy. Dmitry A. Markov and Igor Ges, research associates in biomedical engineering; undergraduate researcher Charlie Wright and John P. Wikswo, Gordon A. Cain University Professor and Director of VIIBRE, participated in the development with Janetopoulos, Seale and Reiserer.
So far, the researchers have used the mirrored wells to examine how protozoa swim and cells divide. "The method is particularly well suited for studying dynamic processes within cells because it can follow them in three dimensions," says Janetopoulos. Researchers in his lab have used the wells to track the 3D position of the centrosome - the specialized region of a cell next to the nucleus that is the assembly point where the microscopic polymer tubes that serve as part of the cell's cytoskeleton are assembled before cell division and broken down afterwards.
The mirrored pyramidal wells provide a high resolution, multi-vantage-point form of microscopy that also makes it easier for researchers to measure a number of important cell properties. For his senior thesis, for example, Wright explored how the technique can be used to measure the volume of individual yeast cells with unprecedented accuracy. In addition, Wikswo and Markov plan to create mirrored microchannels to measure how cells are deformed under stress induced by fluid flowing through hair-width channels in order to determine how fluid flow affects cell behavior and attachment.
A popular method for studying biological processes uses genetic engineering to attach genes that produce fluorescent molecules to different cell structures such as specific surface receptors. This procedure makes the targeted cell structures light up when illuminated by ultraviolet light, but strong UV light also has the potential to damage the structures. If the engineered cell structures are put in a micropyramidal well, the fluorescent light that is emitted toward the mirrored sides is reflected upward toward the microscope, allowing the researchers to reduce the intensity of the UV light and its potential for damaging the engineered cells.
According to Janetopoulos, the micropyramids also have a major advantage for single molecule studies. Optical noise is a constant problem when working at the low light levels involved. Being able to pinpoint actual light sources in two or three dimensions allows the researchers to reject spurious signals. This should be useful in quantitative fluorescence or bioluminescence studies: Cells can be genetically modified to glow in the dark to provide a measure of cellular metabolic activity or the expression of a specific gene.
The research was funded in part by a grant from the Air Force Office of Scientific Research. Vanderbilt University has applied for a patent on the use of the pyramidal mirrored wells for simultaneous, multi-vantage-point imaging.
[Note: A multimedia version of this story is available on Exploration, Vanderbilt's online research magazine, at vanderbilt/exploration/stories/micropyramids.html]
Contact: David F. Salisbury
Vanderbilt University
"With an off-the-shelf laboratory microscope you only see cells from one side, the top," says team member Chris Janetopoulos, assistant professor of biological sciences. "Not only can we see the tops of cells, we can view their sides as well - something biologists almost never see."
The researchers have dubbed their devices "mirrored pyramidal wells." As the name implies, they consist of pyramidal-shaped cavities molded into silicon whose interior surfaces are coated with a reflective layer of gold or platinum. They are microscopic in dimension - about the width of a human hair - and can be made in a range of sizes to view different-sized objects. When a cell is placed in such a well and viewed with a regular optical microscope, the researcher can see several sides simultaneously.
"This technology is exciting because these mirrored wells can be made at very low cost, unlike other, more complex methods for 3D microscopy," says Assistant Professor of the Practice of Biomedical Engineering Kevin Seale.
According to Ron Reiserer, "This could easily become as ubiquitous as the microscope slide and could replace more expensive methods currently used to position individual cells." Reiserer is a lab manager at the Vanderbilt Institute for Integrative Biosystems Research and Education (VIIBRE) who helped design the protocol used to make the micropyramids.
The Vanderbilt group is not the first to make microscopic pyramidal wells, but it is the first to apply them to make 3D images of microorganisms. In 2006, a group of scientists in England created pyramidal micromirrors and applied them to trapping atoms. And last spring researchers at the National Institute of Standards and Technology used similar structures to track nanoparticles.
The Vanderbilt researchers reported their achievement last September in the Journal of Microscopy. Dmitry A. Markov and Igor Ges, research associates in biomedical engineering; undergraduate researcher Charlie Wright and John P. Wikswo, Gordon A. Cain University Professor and Director of VIIBRE, participated in the development with Janetopoulos, Seale and Reiserer.
So far, the researchers have used the mirrored wells to examine how protozoa swim and cells divide. "The method is particularly well suited for studying dynamic processes within cells because it can follow them in three dimensions," says Janetopoulos. Researchers in his lab have used the wells to track the 3D position of the centrosome - the specialized region of a cell next to the nucleus that is the assembly point where the microscopic polymer tubes that serve as part of the cell's cytoskeleton are assembled before cell division and broken down afterwards.
The mirrored pyramidal wells provide a high resolution, multi-vantage-point form of microscopy that also makes it easier for researchers to measure a number of important cell properties. For his senior thesis, for example, Wright explored how the technique can be used to measure the volume of individual yeast cells with unprecedented accuracy. In addition, Wikswo and Markov plan to create mirrored microchannels to measure how cells are deformed under stress induced by fluid flowing through hair-width channels in order to determine how fluid flow affects cell behavior and attachment.
A popular method for studying biological processes uses genetic engineering to attach genes that produce fluorescent molecules to different cell structures such as specific surface receptors. This procedure makes the targeted cell structures light up when illuminated by ultraviolet light, but strong UV light also has the potential to damage the structures. If the engineered cell structures are put in a micropyramidal well, the fluorescent light that is emitted toward the mirrored sides is reflected upward toward the microscope, allowing the researchers to reduce the intensity of the UV light and its potential for damaging the engineered cells.
According to Janetopoulos, the micropyramids also have a major advantage for single molecule studies. Optical noise is a constant problem when working at the low light levels involved. Being able to pinpoint actual light sources in two or three dimensions allows the researchers to reject spurious signals. This should be useful in quantitative fluorescence or bioluminescence studies: Cells can be genetically modified to glow in the dark to provide a measure of cellular metabolic activity or the expression of a specific gene.
The research was funded in part by a grant from the Air Force Office of Scientific Research. Vanderbilt University has applied for a patent on the use of the pyramidal mirrored wells for simultaneous, multi-vantage-point imaging.
[Note: A multimedia version of this story is available on Exploration, Vanderbilt's online research magazine, at vanderbilt/exploration/stories/micropyramids.html]
Contact: David F. Salisbury
Vanderbilt University
CLSI Publishes Updated Guidelines For Clinical Evaluation Of Immunoassays And Methods For Utilizing Flow Cytometry And Solid Phase Assays
Clinical and Laboratory Standards Institute (CLSI) has recently published guidelines in the area of immunology and ligand assay.
Clinical Evaluation of Immunoassays; Approved Guideline-Second Edition (I/LA21-A2) provides guidance for assessing analytical performance, methods comparison, and clinical accuracy of laboratory tests. This document focuses on unique characteristics of immunoassays, and provides a guide to designing, executing, and analyzing a clinical evaluation.
Marilyn M. Lightfoote, MD, PhD, Food and Drug Administration (FDA) Center for Devices and Radiological Health, and chairholder of the working group that developed the guideline says, "The updated information provides a must-have resource for specialty laboratories, industry, and developers of assays. It is a terrific reference document."
The elements of this guideline include:
- a development plan for an effective analysis and evaluation;
- a discussion of the planning and design considerations that are necessary for a successful evaluation;
- a description of requirements for conducting the evaluation through monitoring and database management; and
- a brief review of the analytical performance measures that must be in place before testing clinical specimens.
This document replaces the first edition of the approved guideline, I/LA21-A, which was published in 2002. It includes the following updates:
- specific details on selection and use of test specimen panels;
- specimen library collections;
- reference panels including specimen commutability issues;
- sample size considerations for evaluation studies; and
- an appendix to guide the user in sample size selections.
This document will aid developers of "in-house" assays for institutional use, developers of assays used for monitoring pharmacologic effects of new drugs or biologics, and clinical and regulatory personnel responsible for commercializing products.
CLSI has also published a new document, Detection of HLA-Specific Alloantibody by Flow Cytometry and Solid Phase Assays; Approved Guideline (I/LA29-A), which describes criteria for optimizing flow cytometry crossmatching and the detection of human leukocyte antigen (HLA) alloantibody by solid-phase methods in conventional and multiplex platforms.
The specific areas addressed in the guideline include:
- technical consideration for instrument setup and staining procedures;
- screening methods;
- single-antigen and multiantigen approaches;
- reporting formats;
- clinical interpretation; and
- multicenter quality assurance.
This guideline is intended for solid organ and stem cell transplant laboratories, manufacturers of systems for histocompatability testing, and organizations that manage organ sharing.
CLSI is a global, nonprofit, membership-based organization dedicated to developing standards and guidelines for the health care and medical-testing community. CLSI's unique consensus process facilitates the creation of standards and guidelines that are reliable, practical, and achievable for an effective quality system.
CLSI
Clinical Evaluation of Immunoassays; Approved Guideline-Second Edition (I/LA21-A2) provides guidance for assessing analytical performance, methods comparison, and clinical accuracy of laboratory tests. This document focuses on unique characteristics of immunoassays, and provides a guide to designing, executing, and analyzing a clinical evaluation.
Marilyn M. Lightfoote, MD, PhD, Food and Drug Administration (FDA) Center for Devices and Radiological Health, and chairholder of the working group that developed the guideline says, "The updated information provides a must-have resource for specialty laboratories, industry, and developers of assays. It is a terrific reference document."
The elements of this guideline include:
- a development plan for an effective analysis and evaluation;
- a discussion of the planning and design considerations that are necessary for a successful evaluation;
- a description of requirements for conducting the evaluation through monitoring and database management; and
- a brief review of the analytical performance measures that must be in place before testing clinical specimens.
This document replaces the first edition of the approved guideline, I/LA21-A, which was published in 2002. It includes the following updates:
- specific details on selection and use of test specimen panels;
- specimen library collections;
- reference panels including specimen commutability issues;
- sample size considerations for evaluation studies; and
- an appendix to guide the user in sample size selections.
This document will aid developers of "in-house" assays for institutional use, developers of assays used for monitoring pharmacologic effects of new drugs or biologics, and clinical and regulatory personnel responsible for commercializing products.
CLSI has also published a new document, Detection of HLA-Specific Alloantibody by Flow Cytometry and Solid Phase Assays; Approved Guideline (I/LA29-A), which describes criteria for optimizing flow cytometry crossmatching and the detection of human leukocyte antigen (HLA) alloantibody by solid-phase methods in conventional and multiplex platforms.
The specific areas addressed in the guideline include:
- technical consideration for instrument setup and staining procedures;
- screening methods;
- single-antigen and multiantigen approaches;
- reporting formats;
- clinical interpretation; and
- multicenter quality assurance.
This guideline is intended for solid organ and stem cell transplant laboratories, manufacturers of systems for histocompatability testing, and organizations that manage organ sharing.
CLSI is a global, nonprofit, membership-based organization dedicated to developing standards and guidelines for the health care and medical-testing community. CLSI's unique consensus process facilitates the creation of standards and guidelines that are reliable, practical, and achievable for an effective quality system.
CLSI
New Method Enabling Routine Targeted Gene Modification Developed By Consortium
A multi-institutional team led by Massachusetts General Hospital (MGH) investigators has developed a powerful new tool for genomic research and medicine - a robust method for generating synthetic enzymes that can target particular DNA sequences for inactivation or repair. In the July 25 issue of Molecular Cell, the researchers describe an efficient, publicly available method to engineer customized zinc-finger nucleases (ZFNs), which can be used to induce specific genomic modifications in many types of cells.
"Recent work has shown that ZFNs can alter genes with high efficiency in cells from plants or model organisms like fruitflies, roundworms and zebrafish, and in human cells," says J. Keith Joung, MD, PhD, of the MGH Molecular Pathology Unit, the paper's senior author. "However, a significant bottleneck has been the lack of access to an effective method for generating the customized DNA-binding domains needed to guide ZFNs to their target sites. Our method will enable academic researchers to rapidly create high quality ZFNs for genes of interest and will stimulate use of this technology in biological research and potentially gene therapy."
Zinc-finger peptides, which bind to DNA, occur naturally in many important proteins that regulate or otherwise interact with DNA. Zinc-finger nucleases are constructed from synthetic "designer" zinc-finger domains targeted to a specific genetic sequence and another protein segment that breaks both DNA strands within the binding site. Currently available methods for generating ZFNs are either inefficient or involve constructing and analyzing huge libraries of zinc-finger peptides, a task that exceeds the capabilities of all but a handful of laboratories in the world.
First author Morgan L. Maeder of the Joung lab led an effort by researchers from six institutions that demonstrated how this new method (termed OPEN for Oligomerized Pool ENgineering) can rapidly generate ZFNs that induce alterations at sites in three biologically important human genes and a plant gene. ZFNs made by the new OPEN method - which utilizes a new archive of reagents that will be made publicly available by the Zinc Finger Consortium - were so efficient that they could modify as many as four copies of a gene in human cells and two copies in plant cells.
"Our study provides the first evidence that ZFNs can make specific changes in plant genes with high efficiency and opens a new avenue for plant genetic modification," says Daniel Voytas, PhD, of the University of Minnesota, whose lab conducted the plant cell experiments. Recently relocated from Iowa State University, Voytas and his team are interested in modifying plant genes for crop improvement.
"With the development of OPEN, many more academic labs will be able to construct, test and use ZFNs in their biological research projects," adds Joung. "OPEN should also stimulate additional research into the potential application of ZFNs for gene therapy of single-gene disorders, such as sickle cell anemia and cystic fibrosis." Joung's lab has already begun to explore ways to further simplify the OPEN method so that it can be performed more quickly and for a larger number of gene targets at once. He is an assistant Professor of Pathology at Harvard Medical School and director of the Molecular Pathology Unit at MGH.
The Joung and Voytas teams worked jointly with labs from Charite Medical School in Berlin, the University of Iowa, Iowa State University, and the University of Texas Southwestern Medical Center to develop and validate this new technology. The participating teams are members of the Zinc Finger Consortium (zincfingers/), an international group of investigators committed to the development of engineered zinc-finger nuclease technology.
The study was supported by organizations including the National Institutes of Health, the National Science Foundation, the Cystic Fibrosis Foundation, the European Commission's 6th Framework Programme, and the Roy Carver Charitable Trust.
Massachusetts General Hospital (massgeneral/), established in 1811, is the original and largest teaching hospital of Harvard Medical School. The MGH conducts the largest hospital-based research program in the United States, with an annual research budget of more than $500 million and major research centers in AIDS, cardiovascular research, cancer, computational and integrative biology, cutaneous biology, human genetics, medical imaging, neurodegenerative disorders, regenerative medicine, systems biology, transplantation biology and photomedicine.
The University of Minnesota's Academic Health Center (ahc.umn/) is home to six health professional schools and colleges as well as several health-related centers and institutes. Founded in 1851, the University of Minnesota is one of the oldest and largest land grant institutions in the country. The Academic Health Center prepares the new health professionals who improve the health of communities, discover and deliver new treatments and cures, and strengthen the health economy.
Source: Sue McGreevey
Massachusetts General Hospital
"Recent work has shown that ZFNs can alter genes with high efficiency in cells from plants or model organisms like fruitflies, roundworms and zebrafish, and in human cells," says J. Keith Joung, MD, PhD, of the MGH Molecular Pathology Unit, the paper's senior author. "However, a significant bottleneck has been the lack of access to an effective method for generating the customized DNA-binding domains needed to guide ZFNs to their target sites. Our method will enable academic researchers to rapidly create high quality ZFNs for genes of interest and will stimulate use of this technology in biological research and potentially gene therapy."
Zinc-finger peptides, which bind to DNA, occur naturally in many important proteins that regulate or otherwise interact with DNA. Zinc-finger nucleases are constructed from synthetic "designer" zinc-finger domains targeted to a specific genetic sequence and another protein segment that breaks both DNA strands within the binding site. Currently available methods for generating ZFNs are either inefficient or involve constructing and analyzing huge libraries of zinc-finger peptides, a task that exceeds the capabilities of all but a handful of laboratories in the world.
First author Morgan L. Maeder of the Joung lab led an effort by researchers from six institutions that demonstrated how this new method (termed OPEN for Oligomerized Pool ENgineering) can rapidly generate ZFNs that induce alterations at sites in three biologically important human genes and a plant gene. ZFNs made by the new OPEN method - which utilizes a new archive of reagents that will be made publicly available by the Zinc Finger Consortium - were so efficient that they could modify as many as four copies of a gene in human cells and two copies in plant cells.
"Our study provides the first evidence that ZFNs can make specific changes in plant genes with high efficiency and opens a new avenue for plant genetic modification," says Daniel Voytas, PhD, of the University of Minnesota, whose lab conducted the plant cell experiments. Recently relocated from Iowa State University, Voytas and his team are interested in modifying plant genes for crop improvement.
"With the development of OPEN, many more academic labs will be able to construct, test and use ZFNs in their biological research projects," adds Joung. "OPEN should also stimulate additional research into the potential application of ZFNs for gene therapy of single-gene disorders, such as sickle cell anemia and cystic fibrosis." Joung's lab has already begun to explore ways to further simplify the OPEN method so that it can be performed more quickly and for a larger number of gene targets at once. He is an assistant Professor of Pathology at Harvard Medical School and director of the Molecular Pathology Unit at MGH.
The Joung and Voytas teams worked jointly with labs from Charite Medical School in Berlin, the University of Iowa, Iowa State University, and the University of Texas Southwestern Medical Center to develop and validate this new technology. The participating teams are members of the Zinc Finger Consortium (zincfingers/), an international group of investigators committed to the development of engineered zinc-finger nuclease technology.
The study was supported by organizations including the National Institutes of Health, the National Science Foundation, the Cystic Fibrosis Foundation, the European Commission's 6th Framework Programme, and the Roy Carver Charitable Trust.
Massachusetts General Hospital (massgeneral/), established in 1811, is the original and largest teaching hospital of Harvard Medical School. The MGH conducts the largest hospital-based research program in the United States, with an annual research budget of more than $500 million and major research centers in AIDS, cardiovascular research, cancer, computational and integrative biology, cutaneous biology, human genetics, medical imaging, neurodegenerative disorders, regenerative medicine, systems biology, transplantation biology and photomedicine.
The University of Minnesota's Academic Health Center (ahc.umn/) is home to six health professional schools and colleges as well as several health-related centers and institutes. Founded in 1851, the University of Minnesota is one of the oldest and largest land grant institutions in the country. The Academic Health Center prepares the new health professionals who improve the health of communities, discover and deliver new treatments and cures, and strengthen the health economy.
Source: Sue McGreevey
Massachusetts General Hospital
Подписаться на:
Сообщения (Atom)