Fuzzy Logic Predicts Cell Aging

The process of aging disturbs a broad range of cellular mechanisms in a complex fashion and is not well understood. Computer models using fuzzy logic might help to unravel these complexities and predict how aging progresses in cells and organisms, according to a study from Drexel University in Philadelphia and Children's Hospital Boston.

"One important goal of computational approaches in aging is to develop integrated models of a unifying aging theory in order to better understand the progression of aging phenotypes grounded on molecular mechanisms," said Andres Kriete, Associate Professor at Drexel's School of Biomedical Engineering, Science and Health Systems and lead author of the study.

The study, which will appear in the June issue of PLoS Computational Biology, relates progressive damage and dysfunction in aging, dubbed a vicious cycle, to inflammatory and metabolic stress response pathways. Interestingly, the activation of these pathways remodels the inner functioning of the cell in a protective and adaptive manner and thus extends lifespan.

This is the first time that scientists have applied fuzzy logic modeling to the field of aging. "Since cellular biodynamics in aging may be considered a complex control system, a fuzzy logic approach seems to be particularly suitable," said Dr. William Bosl, co-author of this study. Dr. Bosl, a staff scientist in the Informatics Program at Children's Hospital Boston, developed a fuzzy logic modeling platform called Bionet together with a cell biologist, Dr. Rong Li of the Stowers Institute for Medical Research in Kansas City, to study the complex interactions that occur in a cell's machinery using the kind of qualitative information gained from laboratory experiments.

Fuzzy logic can handle imprecise input, but makes precise decisions and has wide industrial applications from air conditioning to anti-lock break systems in cars, using predefined rules. In a similar fashion, the aging model relies on sets of rules drawn from experimental data to describe molecular interactions. "Integration of such data is the declared goal of systems biology, which enables simulation of the response of cells to signaling cues, cell cycling and cell death," said Glenn Booker, who is Faculty at the College of Information Science and Technology at Drexel and co-author on the study.

Applications in aging are currently geared towards deciphering the underlying connections and networks. "We have to realize that the real strength of computational systems biology in aging is to be able to predict and develop strategies to control cellular networks better as they may be related to age related diseases," said Dr. Kriete, "and our approach is just a first step in this direction."

Journal Reference:

1. Kriete A, Bosl WJ, Booker G. Rule-Based Cell Systems Model of Aging using Feedback Loop Motifs Mediated by Stress Responses. PLoS Computational Biology, 2010; 6 (6): e1000820 DOI: 10.1371/journal.pcbi.1000820

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Gut-Residing Bacteria Trigger Arthritis in Genetically Susceptible Individuals

A single species of bacteria that lives in the gut is able to trigger a cascade of immune responses that can ultimately result in the development of arthritis.

Our gut, like that of most mammals, is filled with thousands of species of bacteria, many of which are helpful and aid in the development of a normal, healthy immune system. Gut-residing bacteria can also play a role in disorders of the immune system, especially autoimmune disorders in which the body attacks its own cells.

It turns out that rheumatoid arthritis is one such disorder. Researchers in the laboratories of Christophe Benoist and Diane Mathis at Harvard Medical School and Dan Littman at New York University made this discovery while working in mice prone to arthritis.

"In the absence of all bacteria, these mice didn't develop arthritis, but the introduction of a single bacterium was enough to jump-start the immune process that leads to development of the disease," says Mathis, an HMS professor of pathology.

The findings appear in the June 25 issue of the journal Immunity.

The researchers began by raising arthritis-prone mice in a germ-free environment. The mice had much lower levels of arthritis-causing autoantibodies than mice raised in a non-germ-free facility. The germ-free mice also showed strong attenuation in the onset and severity of clinical arthritis.

At three weeks of age, some mice were transferred to a non-germ-free facility and the researchers introduced segmented filamentous bacteria into their systems. When they introduced this normally-occurring bacteria into the mice, the animals rapidly began producing autoantibodies and developed arthritis within four days.

First author Hsin-Jung Wu emphasizes that these bacteria do not cause the mice to "catch" arthritis. "It's more that they have the genetic susceptibility, and this bacterium creates an environment that allows this genetic susceptibility to play out," says Wu, a postdoctoral researcher at Harvard Medical School. "It's an interaction between genetics and the environment."

The team mapped out the complex chain of events leading to arthritis. The segmented filamentous bacteria cause the animals to produce more of a particular subset of T cells. The immune system reacts to the activity of the T cells as if to a foreign threat and produces autoantibodies that trigger the devastating disease.

One surprising finding was that bacteria in the gut could influence the development of an autoimmune disease affecting tissues distant from the gut. Diseases such as irritable bowel syndrome have been linked to gut-residing bacteria, but this study is unique in showing the mechanism by which a bacterium in the gut can influence the development of an autoimmune response that ends in inflammation and pain in the joints.

The team will continue to use this mouse model of arthritis to answer questions about the link between the disease and autoimmune response. Next, they plan on tackling the molecular explanation of how these bacteria promote the development of this particular subset of T cells and to explore connections with other autoimmune diseases, in particular type-1 diabetes.

This research was funded by the National Institutes of Health.

Story Source:

The above story is reprinted (with editorial adaptations by ScienceDaily staff) from materials provided by Harvard Medical School. The original article was written by Mary Bates.

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Malaria Threat Is as Old as Humanity, New Research Shows

New research by scientists funded by the Biotechnology and Biological Sciences Research Council (BBSRC) shows that malaria is tens of thousands of years older than previously thought. An international team, led by researchers at Imperial College London, have found that the potentially deadly tropical disease evolved alongside anatomically modern humans and moved with our ancestors as they migrated out of Africa around 60-80,000 years ago.

The research is published in the journal Current Biology.

The findings and the techniques in the study could be important in informing current control strategies aimed at reducing the prevalence of malaria. There are an estimated 230 million cases each year, causing between 1 and 3 million deaths, and around 1.4bn people are considered to be at risk of infection.

Dr Francois Balloux from the Medical Research Council (MRC) Centre for Outbreak Analysis and Modelling at Imperial College London was lead researcher on the project. He said: "Most recent work to understand how malaria has spread across the tropics has worked on the premise that the disease arose alongside the development of agriculture around 10,000 years ago. Our research shows that the malaria parasite has evolved and spread alongside humans and is at least as old as the event of the human expansion out of Africa 60-80,000 years ago."

The international team worked on the largest collection of malaria parasites ever assembled. By characterising them by DNA sequencing they were able to track the progress of malaria across the tropics and to calculate the age of the parasite. The scientists discovered clear correlation of decreasing genetic diversity with distance from sub-Saharan Africa. This accurately mirrored the same data for humans suggesting strong evidence of co-evolution and migration.

Dr Balloux said: "The genetic sequencing of the malaria parasite shows a geographic spread pattern with striking similarities to studies on humans. This points to a shared geographic origin, age and route of spread around the world. This understanding is important because despite the prevalence and deadly impact of malaria little research has previously been done to understand the genetic variation of the parasite. The genetic diversity of malaria parasites is central to their threat as it helps them to overcome the immune system and to develop drug resistance, making this research vital in informing new and more effective control strategies."

Journal Reference:

1. Tanabe et al. Plasmodium falciparum Accompanied the Human Expansion out of Africa. Current Biology, 2010; DOI: 10.1016/j.cub.2010.05.053

Courtesy: ScienceDaily

How DNA Is Copied Onto RNA Revealed Through Three-Dimensional Transcription Film

Gene expression takes place in two stages: the transcription of DNA to RNA by an enzyme called RNA polymerase, , followed by the translation of this RNA into proteins, whose behaviour affects the characteristics of each individual.

Transcription: a mechanism controlled in time and space

Transcription involves about fifty regulatory molecules that interact with each other to begin reading the gene at the right place and the right time. The slightest irregularity of one of these molecules disturbs the transcription. An understanding of the initiation and regulation mechanisms is essential in order to understand gene expression. The structural biology researchers at IGBMC are studying molecular structures to gain a better understanding of how they function. Patrick Schultz's team is particularly focusing on the architecture of the molecules involved in transcription and attempting to decode the mechanisms of their interactions.

An 'image-by-image' analysis

An analysis of the transcription complexes by electron cryomicroscopy allows a molecule to be observed in a hydrated state close to its natural state. Each photograph, taken using a microscope, shows thousands of specimens of the same molecule from different angles and at different instants in their reaction cycle. The statistical analysis of these images performed by Patrick Schultz's team revealed different conformations in three dimensions, which correspond to different stages of transcription initiation. 'We performed image-by-image sequencing and made a film of the initial stages of transcription,' says Schultz.

The factor TFIID, the main player in the transcription process

Patrick Schultz's team is interested in a complex protein that acts as an assembly platform in the initiation phase of transcription: the factor TFIID. Through interaction with the activator Rap1, bound upstream from the gene to be transcribed, it is attracted to the DNA and binds onto it. Combined with another factor, TFIIA, it changes conformation and allows the RNA polymerase to initiate transcription. The original aspect of this mechanism is based on the formation of a DNA loop, which allows the RNA polymerase to be positioned exactly at the start of the sequence of the gene to be transcribed.

The structure of the transcription factor TFIID obtained after image analysis is represented in yellow on an electron cryomicroscopy image background, showing the frozen hydrated molecules in dark grey. The transcription activator Rap1 (red) interacts with the factor TFIIA (blue) and contributes to forming a DNA loop (green).

What is electron cryomicroscopy?

The biological molecules in living organisms exist in an aqueous environment, which must be preserved whilst observing the molecules. In order to be 'seen', however, molecules must be placed in an electron microscope, which operates in a vacuum and dehydrates the sample. The solution, developed in the 1980s, is to use refrigeration to keep the specimen hydrated and to examine it by electron cryomicroscopy. A very thin film (approximately 100 nm, or one ten-thousandth of a millimetre thick) of the suspension containing the sample to be analysed must be created in order to be transparent to electrons. (Thin film shown in light blue in Figure A.) This film is cooled very rapidly (at a rate of approximately 10,000°C per second) by plunging it into liquid ethane cooled to -170°C. This freezing speed prevents the formation of ice crystals, and the sample (yellow in Figure A) is trapped in a layer of vitrified water. The cold chain must be maintained throughout the observation period using a cold plate. The molecules (dark grey in Figure B) are hydrated and observed without contrast agent.

Journal Reference:

1. Papai et al. TFIIA and the transactivator Rap1 cooperate to commit TFIID for transcription initiation. Nature, 2010; 465 (7300): 956 DOI: 10.1038/nature09080

Courtesy: ScienceDaily

Single-Molecule Devices Can Serve as Powerful New Science Tools

With controlled stretching of molecules, Cornell researchers have demonstrated that single-molecule devices can serve as powerful new tools for fundamental science experiments. Their work has resulted in detailed tests of long-existing theories on how electrons interact at the nanoscale.

The work, led by professor of physics Dan Ralph, is published in the June 10 online edition of the journal Science. First author is J.J. Parks, a former graduate student in Ralph's lab.

The scientists studied particular cobalt-based molecules with so-called intrinsic spin -- a quantized amount of angular momentum.

Theories first postulated in the 1980s predicted that molecular spin would alter the interaction between electrons in the molecule and conduction electrons surrounding it, and that this interaction would determine how easily electrons flow through the molecule. Before now, these theories had not been tested in detail because of the difficulties involved in making devices with controlled spins.

Understanding single-molecule electronics requires expertise in both chemistry and physics, and Cornell's team has specialists in both.

"People know about high-spin molecules, but no one has been able to bring together the chemistry and physics to make controlled contact with these high-spin molecules," Ralph said.

The researchers made their observations by stretching individual spin-containing molecules between two electrodes and analyzing their electrical properties. They watched electrons flow through the cobalt complex, cooled to extremely low temperatures, while slowly pulling on the ends to stretch it. At a particular point, it became more difficult to pass current through the molecule. The researchers had subtly changed the magnetic properties of the molecule by making it less symmetric.

After releasing the tension, the molecule returned to its original shape and began passing current more easily -- thus showing the molecule had not been harmed. Measurements as a function of temperature, magnetic field and the extent of stretching gave the team new insights into exactly what is the influence of molecular spin on the electron interactions and electron flow.

The effects of high spin on the electrical properties of nanoscale devices were entirely theoretical issues before the Cornell work, Ralph said. By making devices containing individual high-spin molecules and using stretching to control the spin, the Cornell team proved that such devices can serve as a powerful laboratory for addressing these fundamental scientific questions.

The study was funded primarily by the National Science Foundation.

Journal Reference:

1. J. J. Parks, A. R. Champagne, T. A. Costi, W. W. Shum, A. N. Pasupathy, E. Neuscamman, S. Flores-Torres, P. S. Cornaglia, A. A. Aligia, C. A. Balseiro, G. K.-L. Chan, H. D. Abruña, and D. C. Ralph. Mechanical Control of Spin States in Spin-1 Molecules and the Underscreened Kondo Effect. Science, June 10, 2010 DOI: 10.1126/science.1186874

Courtesy: ScienceDaily

Plastic Antibody Works in First Tests in Living Animals

Scientists are reporting the first evidence that a plastic antibody -- an artificial version of the proteins produced by the body's immune system to recognize and fight infections and foreign substances -- works in the bloodstream of a living animal.

The discovery, they suggest in a report in the Journal of the American Chemical Society, is an advance toward medical use of simple plastic particles custom tailored to fight an array of troublesome "antigens."

Those antigens include everything from disease-causing viruses and bacteria to the troublesome proteins that cause allergic reactions to plant pollen, house dust, certain foods, poison ivy, bee stings and other substances.

In the report, Kenneth Shea, Yu Hosino, and colleagues refer to previous research in which they developed a method for making plastic nanoparticles, barely 1/50,000th the width of a human hair, that mimic natural antibodies in their ability to latch onto an antigen. That antigen was melittin, the main toxin in bee venom. They make the antibody with molecular imprinting, a process similar to leaving a footprint in wet concrete. The scientists mixed melittin with small molecules called monomers, and then started a chemical reaction that links those building blocks into long chains, and makes them solidify. When the plastic dots hardened, the researchers leached the poison out. That left the nanoparticles with tiny toxin-shaped craters.

Their new research, together with Naoto Oku's group of the University Shizuoka Japan, established that the plastic melittin antibodies worked like natural antibodies. The scientists gave lab mice lethal injections of melittin, which breaks open and kills cells. Animals that then immediately received an injection of the melittin-targeting plastic antibody showed a significantly higher survival rate than those that did not receive the nanoparticles. Such nanoparticles could be fabricated for a variety of targets, Shea says. "This opens the door to serious consideration for these nanoparticles in all applications where antibodies are used," he adds.

Journal Reference:

1. Hoshino et al. Recognition, Neutralization, and Clearance of Target Peptides in the Bloodstream of Living Mice by Molecularly Imprinted Polymer Nanoparticles: A Plastic Antibody. Journal of the American Chemical Society, 2010; 132 (19): 6644 DOI: 10.1021/ja102148f

Courtesy: ScienceDaily

New Strain of Bacteria Discovered That Could Aid in Oil Spill, Other Environmental Cleanup

Researchers have discovered a new strain of bacteria that can produce non-toxic, comparatively inexpensive "rhamnolipids," and effectively help degrade polycyclic aromatic hydrocarbons, or PAHs -- environmental pollutants that are one of the most harmful aspects of oil spills.

Because of its unique characteristics, this new bacterial strain could be of considerable value in the long-term cleanup of the massive Gulf Coast oil spill, scientists say.

More research to further reduce costs and scale up production would be needed before its commercial use, they added.

The findings on this new bacterial strain that degrades the PAHs in oil and other hydrocarbons were just published in a professional journal, Biotechnology Advances, by researchers from Oregon State University and two collaborating universities in China. OSU is filing for a patent on the discovery.

"PAHs are a widespread group of toxic, carcinogenic and mutagenic compounds, but also one of the biggest concerns about oil spills," said Xihou Yin, a research assistant professor in the OSU College of Pharmacy.

"Some of the most toxic aspects of oil to fish, wildlife and humans are from PAHs," Yin said. "They can cause cancer, suppress immune system function, cause reproductive problems, nervous system effects and other health issues. This particular strain of bacteria appears to break up and degrade PAHs better than other approaches we have available."

The discovery is strain "NY3" of a common bacterium that has been known of for decades, called Pseudomonas aeruginosa. It was isolated from a site in Shaanxi Province in China, where soils had been contaminated by oil.

P. aeruginosa is widespread in the environment and can cause serious infections, but usually in people with health problems or compromised immune systems. However, some strains also have useful properties, including the ability to produce a group of "biosurfactants" called rhamnolipids.

A "surfactant," technically, is a type of wetting agent that lowers surface tension between liquids -- but we recognize surfactants more commonly in such products as dishwashing detergent or shampoo. Biosurfactants are produced by living cells such as bacteria, fungi and yeast, and are generally non-toxic, environmentally benign and biodegradable. By comparison, chemical surfactants, which are usually derived from petroleum, are commonly toxic to health and ecosystems, and resist complete degradation.

Biosurfactants of various types are already used in a wide range of applications, from food processing to productions of paints, cosmetics, household products and pharmaceuticals. But they also have uses in decontamination of water and soils, with abilities to degrade such toxic compounds as heavy metals, carcinogenic pesticides and hydrocarbons.

Although the type of biosurfactant called "rhamnolipids" have been used for many years, the newly discovered strain, NY3, stands out for some important reasons. Researchers said in the new study that it has an "extraordinary capacity" to produce rhamnolipids that could help break down oil, and then degrade some of its most serious toxic compounds, the PAHs.

Rhamnolipids are not toxic to microbial flora, human beings and animals, and they are completely biodegradable. These are compelling advantages over their synthetic chemical counterparts made from petroleum. Even at a very low concentration, rhamnolipids could remarkably increase the mobility, solubility and bioavailability of PAHs, and strain NY3 of P. aeruginosa has a strong capability of then degrading and decontaminating the PAHs.

"The real bottleneck to replacing synthetic chemicals with biosurfactants like rhamnolipid is the high cost of production," Yin said. "Most of the strains of P. aeruginosa now being used have a low yield of rhamnolipid. But strain NY3 has been optimized to produce a very high yield of 12 grams per liter, from initial production levels of 20 milligrams per liter."

By using low-cost sources of carbon or genetic engineering techniques, it may be possible to reduce costs even further and scale up production at very cost-effective levels, researchers said.

The rhamnolipids produced by NY3 strain appear to be stable in a wide range of temperature, pH and salinity conditions, and strain NY3 aggressively and efficiently degrades at least five PAH compounds of concern, the study showed. It's easy to grow and cultivate in many routine laboratory media, and might be available for commercial use in a fairly short time. Further support to develop the technology is going to be sought from the National Science Foundation.

"Compared to their chemically synthesized counterparts, microbial surfactants show great potential for useful activity with less environmental risk," the researchers wrote in their report. "The search for safe and efficient methods to remove environmental pollutants is a major impetus in the search for novel biosurfactant-producing and PAH-degrading microorganisms."

Collaborating on this research were scientists from Xi'an University of Architecture and Technology and Nanjing Agricultural University in China.

Journal Reference:

1. Maiqian Nie, Xihou Yin, Chunyan Ren, Yang Wang, Feng Xu, Qirong Shen. Novel rhamnolipid biosurfactants produced by a polycyclic aromatic hydrocarbon-degrading bacterium Pseudomonas aeruginosa strain NY3. Biotechnology Advances, 2010; DOI: 10.1016/j.biotechadv.2010.05.013

Courtesy: ScienceDaily

New Technique Turns Proteins Into Glass: Could Lead to New Ways to Deliver Medication

Duke University researchers have devised a method to dry and preserve proteins in a glassified form that seems to retain the molecules' properties as workhorses of biology.

They are exploring whether their glassification technique could bring about protein-based drugs that are cheaper to make and easier to deliver than current techniques which render proteins into freeze dried powders to preserve them.

Duke engineer and chemist David Needham describes this glassification process as "molecular water surgery" because it removes virtually all the water from around a dissolved protein by almost magically pulling the water into a second solvent.

"It's like a sponge sucking water off a counter," said Needham, a professor of mechanical engineering and materials science at Duke's Pratt School of Engineering, who has formed a company called Biogyali ("gyali" means glass in Greek) to develop the innovation. That firm has also applied to patent the idea of turning proteins into tiny glass beads at room temperature for drug delivery systems.

A report by Needham, graduate student Deborah Rickard and former graduate student P. Brent Duncan online in the Biophysical Journal describes how his team carefully controlled water removal during glassification by releasing single tiny droplets of water-dissolved protein into the organic solvent decanol with a micropipette.

Preliminary evaluations by his senior scientist David Gaul and a team of undergraduate students showed that four test proteins undergoing such procedures retained all or most of their original activity when water was restored. His group has received about $1 million from the National Institutes of Health grants for the research.

Having devised a way to turn proteins into glassy microbeads measuring only about 26 millionths of a meter in diameter, Needham hopes those can be directly injected into the body for use as "biologic" drugs.

His group's early research shows high concentrations of such tiny beadlets would not be as viscous as proteins dehydrated into the normal powder form, which tend to clog up syringes, he said.

These microbeads might also be packaged for slow time-release by surrounding them with a polymer that would biodegrade over time, though how to do that has not been resolved yet, he added.

In collaborations with Duke's Brain Tumor Center and Comprehensive Cancer Center, the researchers are seeking additional funding to do initial evaluations on glassified forms of three molecules with drug potential.

One, known as O6-AMBG, can help the cancer drug Temozolomide work better when infused into brain tumors. A second, Lapatinib, is designed to knock out other molecules that help cancer cells grow in the breast and elsewhere. The third, shepherdin, also targets breast cancers.

Their discovery of protein glassification grew out of a basic exploration of a general question: What can dissolve in what?

Needham's research group found, for example, that air and the organic liquid chloroform will both dissolve in water at about the same rate. It also found that water will dissolve in decanol, a substance it cannot even mix with in large quantities.

These experiments, and the theory underlying them, are described in a second report led by Needhams's graduate student Jonathan Su, now published online in the Journal of Chemical Physics ( http://link.aip.org/link/?JCP/132/044506 ).

"Mixing" and "dissolving" are not the same thing, Needham said. "A good example of a suspended mixture is salad dressing, where oil and water are mixed but oil does not appreciably dissolve in water, nor water in oil."

They next tried a more complex variation of a familiar high school experiment which dissolves so much salt in water that some begins coming back out of the solution as a crystal.

In this case, after dissolving the salt in water, Needham's group then inserted a microbubble of that solution into immiscible decanol in a microscopic chamber. The water itself then dissolved into the decanol and left behind the salt, which also crystallized.

According to his group's Biophysical Journal report, while decanol has practically no tendency to dissolve in water, water has a high probability of dissolving in decanol, allowing the latter to be used as a "drying" agent to remove the former.

"So then we asked: what if we did the same thing with the protein albumin?" Needham said. "I expected to maybe get crystallized albumin," Needham recalled. "But, in just a few minutes, we instead formed a glassified microbead of protein on the tip of a micropipette, at a high density just a bit more dense than water itself. That protein glass is not a crystal. It's really a solid liquid."

Many proteins can be coaxed into forming crystals, solids created by repeating three dimensional patterns of atoms as surrounding water is removed. On the other hand, Needham said he was not really surprised that his protein samples instead formed into glasses, which are more unorganized assemblage of molecules that can still "flow" over very long time scales.

The water loss in his process is apparently too rapid for the molecules of big and irregular proteins to reorganize into a crystal form in such a short time, he explained.

Careful studies by his graduate student Rickard found that the decanol removed all the water that is not bound up in the proteins' molecular structures. And the remaining "bound" water was insufficient to support the growth of bacteria and fungi. Storing proteins as microbeads could thus preserve them.

Proteins are currently dried into clumpy, irregular powders by several industrial processes -- usually freeze-drying -- to protect them from such microbe damage. Drying also avoids the chemical breakdowns that can also occur when proteins are kept in solution. "But in the freeze-drying process itself, some very sensitive biologic drugs can also get damaged," Needham said.

Freeze-drying proteins into solids is also slower and more expensive than glassifying them, he added. And the resulting "flaky" powder is harder to handle than glassified beads.

Glassification "is a fast process," said Gaul, a senior research scientist in Needham's lab. Unlike freeze-drying, "we can dry particles within minutes, if not seconds, and don't need any specialized equipment."

Journal Reference:

1. Deborah L. Rickard, P. Brent Duncan, and David Needham. Hydration Potential of Lysozyme: Protein Dehydration Using a Single Microparticle Technique. Biophysical Journal, 2010; 98 (6): 1075-1084 DOI: 10.1016/j.bpj.2009.11.043