Silver nanoparticles show 'immense potential' in prevention of blood clots

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Silver nanoparticles (shown) could help prevent blood clots. Credit: The American Chemical Society.
Scientists are reporting discovery of a potential new alternative to aspirin, ReoPro, and other anti-platelet agents used widely to prevent blood clots in coronary artery disease, heart attack and stroke. Their study, scheduled for the June 23 issue of ACS Nano, a monthly journal, involves particles of silver -- 1/50,000th the diameter of a human hair -- that are injected into the bloodstream.

Debabrata Dash and colleagues point out that patients urgently need new anti-thrombotic agents because traditionally prescribed medications too-often cause dangerous bleeding. At the same time, aging of the population, sedentary lifestyle and spiraling rates of certain diseases have increased the use of these drugs. Researchers are seeking treatments that more gently orchestrate activity of platelets, disk-shaped particles in the blood that form clots.
The scientists describe development and lab testing of silver nanoparticles that seem to keep platelets in an inactive state. Low levels of the nanosilver, injected into mice, reduced the ability of platelets to clump together by as much as 40 percent with no apparent harmful side effects.
The nanoparticles “hold immense potential to be promoted as an antiplatelet agent,” the researchers note. “Nanosilver appears to possess dual significant properties critically helpful to the health of mankind — antibacterial and antiplatelet — which together can have unique utilities, for example in coronary stents.”
More information: ACS Nano, Journal Article: “Characterization of Antiplatelet Properties of Silver Nanoparticles”
Provided by American Chemical Society (news : web)

Chemists see first building blocks to life on Earth

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British scientists said on Wednesday that they had figured out key steps in the process by which life on Earth may have emerged from a seething soup of simple chemicals.

Genetic information in living organisms today is held in deoxyribonucleic acid (DNA), the famous "double helix" molecule of sugar, phosphate and a base.
But DNA is too sophisticated to have popped up in an instant, and one avenue of thought says its single-stranded cousin, ribonucleic acid, or RNA, came first.
RNA plays a key role in making proteins and, in viruses, is used to store genetic code.
It is chemically similar to DNA but is simpler and tougher in structure, and thus looks like a good candidate for Earth's first information-coding nucleic acid.
But for all its allure, the "RNA first" theory has run into practical problems.
Its three ingredients -- the base, ribose sugar and phosphate -- must have formed separately and then combined to form the molecule, according to conventional thinking.
Critics, though, say that RNA, while somewhat simpler than DNA, is still a complex molecule and could not have been assembled spontaneously.
These doubters have been comforted by the failure to find any feasible chain of chemical events to explain how the three components all came together.
But a paper published in the British journal Nature by University of Manchester chemists puts forward a different explanation.
The team, led by Professor John Sutherland, venture that an RNA-like synthesis took place through a series of chemical reactions and an important intermediate substance.
Their lab model uses starting materials and environmental conditions that are believed to have been around in early Earth and are also used in the standard "RNA first" scenario.
Their theory starts with a simple sugar called glycolaldehyde, which reacts with cyanmide (a compound of cyanide and ammonia) and phosphate to produce an intermediate compound called 2-aminooxazole.
Gentle warming from the Sun and cooling at night help purify the 2-aminooxazole, turning it into a plentiful precursor which contributes the sugar and base portions of the new ribonucleotide molecule.
The presence of phosphate and ultraviolet light from the Sun complete the synthesis.
In a commentary also published by Nature, US molecular biologist Jack Szostak hailed the research as an elegant explanation as to why the sugar and base would not have to form separately before forming the new molecule.
"It will stand for years as one of the great advances in prebiotic chemistry," the term for the study of the chemical processes that led to life on Earth, he enthused.
Opinions vary as to when the first organisms appeared on Earth.
One estimate, based on fossilised mats of bacteria found in Australia, is that this happened around 3.8 billion years ago, around 700 million years after the planet was formed.
(c) 2009 AFP

Breakthrough in the treatment of bacterial meningitis

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It can take just hours after the symptoms appear for someone to die from bacterial meningitis. Now, after years of research, experts at The University of Nottingham have finally discovered how the deadly meningococcal bacteria is able to break through the body's natural defence mechanism and attack the brain.

The discovery could lead to better treatment and vaccines for meningitis and could save the lives of hundreds of children.
Bacterial meningitis in childhood is almost exclusively caused by the respiratory tract pathogens Streptococcus pneumoniae, Neisseria meningitidis, and Haemophilus influenzae. The mechanism used by these lethal germs to break through the blood brain barrier (BBB) has, until now, been unknown.
The team led by Dlawer Ala'Aldeen, Professor of Clinical Microbiology and Head of the Molecular Bacteriology and Immunology Group at the Centre for Biomolecular Sciences, recently discovered that all three pathogens target the same receptor on human cerebrovascular endothelial cells — the specialised filtering system that protects our brain from disease — enabling the organisms to cross the blood-brain barrier.
Their findings, published today in The Journal of Clinical Investigation, suggest that disruption or modulation of this interaction of bacterial adhesins with the receptor might offer unexpectedly broad protection against bacterial meningitis and may provide a therapeutic target for the prevention and treatment of disease.
Professor Ala'Aldeen, who has been studying meningitis and its causes for over 20 years, said: "This is a significant breakthrough which will help us design novel strategies for the prevention and treatment of bacterial meningitis. Identification of the human receptor and bacterial ligands is like identifying a mysterious key and its lock, which will open new doors and pave the way for new discoveries."
The research, carried out in collaboration with the Department of Infectious Diseases at St. Jude Children's Research Hospital in Memphis Tennessee, also involved students from the University who have been regular and willing volunteers in the research programme.
Professor Ala'Aldeen said: "The ultimate aim is to save lives by protecting the healthy and curing the sick. We are one step closer to new breakthroughs that would prevent disease or its complications. There still is a long way to go before we have the ultimate vaccine and the ultimate treatment of bacterial meningitis."
Source: University of Nottingham (news : web)

Developed a human monoclonal antibody that neutralizes the Hepatitis C virus (HCV).

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ScienceDaily (May 11, 2009) — aking aim at a leading cause of liver failure in the United States, a team of scientists at the Massachusetts Biologic Laboratories (MBL) of the University of Massachusetts Medical School (UMMS) has developed a human monoclonal antibody that neutralizes the Hepatitis C virus (HCV). The new antibody effectively neutralized the virus in culture, and then prevented infection by the virus in a pre-clinical animal model of the disease.

Details of the research were presented April 23 in Copenhagen, Denmark at the 44th Annual Meeting of the European Association for the Study of the Liver (EASL). "We are pleased with the progress of this program," said Donna Ambrosino, MD, executive director of the MBL and a professor of pediatrics at the Medical School. "This antibody shows significant efficacy against the virus."
In the current study, MBL scientists injected transgenic mice (HuMAb Mouse® technology, Medarex, Inc.) with elements of HCV and then painstakingly searched for individual human antibodies produced in the mice that would recognize and bind to the HCV's outer coat, known as the glycoprotein. Once they found human antibodies that looked promising, they evaluated in vitro the ability of those antibodies to neutralize the virus and selected a lead candidate antibody for further characterization. Collaborative work with clinical researchers from the Department of Medicine at the Medical School's Worcester campus demonstrated that this antibody, now known as MBL-HCV1, was able to bind tightly with all genotypes of HCV tested from infected patient samples.
MBL-HCV1 was then tested off-site on three non-human primates. In that study, one animal received no antibody, one a low dose of the new antibody, and one a higher dose. Then all three animals were exposed to HCV. The animals with low or no antibody dosages developed HCV infections, but the animal with the higher dose was protected. Subsequently, researchers gave the high-dose of the antibody to the animal that originally received no antibody, and in that case the HCV was cleared from that animal's system. "These results are encouraging as a possible treatment for HCV infected patients, but more work needs to be done before we know how effective it will be in people," Dr. Ambrosino noted.
HCV attacks the liver and can eventually lead to liver failure. According to the U.S. Centers for Disease Control and Prevention, 3.2 million Americans are chronically infected with HCV and some 10,000 die annually of the disease. Globally, as many as 170 million people are estimated to suffer from HCV infection. For the most serious cases of HCV that do not respond to antiviral drugs, liver transplantation is the only option.
Typically 2,000 to 4,000 liver transplants are done each year in the United States (far less than the number of people on the waiting list for available organs). Transplantation can be a life saving treatment; however, in nearly all cases the patient's new liver is eventually infected by HCV because the virus remains in the patient's bloodstream during surgery. The powerful antiviral drugs now used to attack HCV prior to end-stage liver failure are not routinely used during surgery due to the patients' weakened condition and because of the strong medication used to avoid rejection of the new liver. After re-infection with HCV, nearly 40 percent of patients suffer rapid liver failure.
To close that clinical gap, the new antibody developed at MBL is designed to be a therapy shortly before and after transplant surgery. By giving a patient the new antibody before and during the time when the donor liver is implanted, researchers hope the HCV virus left in the bloodstream will be neutralized and rendered unable to infect the new liver. Then, because monoclonal antibodies are highly specific and typically have little or no side-effects, additional dosages of the new antibody could, theoretically, be given immediately after transplant surgery to continue neutralizing any remaining virus.
It is also possible, researchers theorize, that the antibody could be used in combination with new antiviral drugs for treatment in patients with newly diagnosed HCV infection. Use of the new antibody for both liver transplant patients and in newly diagnosed HCV patients will now be further evaluated. A Phase 1 human clinical trial of MBL-HCV1 in healthy subjects is expected to begin later this year.
Adapted from materials provided by University of Massachusetts Medical School, via EurekAlert!, a service of AAAS.

Biotechnology: Engineered Moss Can Produce Human Proteins

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ScienceDaily (May 11, 2009) — ETH Zurich researchers have shown that mosses and humans share unexpected common characteristics. These evolutionary relics could be useful in the production of therapeutic proteins.

At first glance, mosses and human beings have little in common. The moss Physcomitrella patens is small, pale green, immobile, and uses sunlight as its energy source. Humans are large, mobile, and need to obtain energy by eating vegetable or animal foods.
Transferring mammalian genes into moss
This made the result of the experiments carried out by researchers in the group led by Martin Fussenegger, Professor of Chemical and Bioengineering at ETH Zurich, all the more astonishing. In collaboration with researchers at the University of Freiburg im Breisgau, the PhD student Marc Gitzinger carried out tests to see what happens when unmodified human or mammalian genes are inserted into the moss genome. They transferred the foreign, unmodified genes into the moss and discovered that the moss was easily able to manufacture the proteins encoded therein.
This cannot be taken for granted, since the same process does not work when a mammalian gene is implanted into what are known as “higher” flowering plants. The reason is that sections of the start and finish sequences of the genes of animals, plants, fungi and bacteria are considerably different. They are responsible for ensuring that a gene in the organism is recognized as such, and the proteins encoded by it are produced in the correct amount and are released from the cell. The more remote the relationship between living organisms, the greater the difference between these sequences. This is why biotechnologists must normally adapt them to a foreign organism before transplanting a gene into it. The researchers were astonished to find that this was not necessary in the case of the moss.
Moss as a generalist
The explanation given for this by Ralf Reski, Professor of Plant Biotechnology at the University of Freiburg im Breisgau, is that the moss has remained a generalist. It underwent the last major modification about 450 million years ago when it changed from living in water to a life on land, adapting to the new living conditions and then remaining unaltered for millions of years, both in its appearance and at a genetic level.
The process used by the moss to produce its proteins is less sophisticated than in “higher” organisms. In contrast to the moss, these latter organisms underwent major further developments and specializations over the course of 450 million years. On the other hand, the moss clearly retained – for millions of years – the ability to read foreign genes such as those from mammals and thus also from humans, and to translate them into proteins, probably without ever having made any use of this capability during these 450 million years.
A cost-effective alternative to mammalian cells
Today, the moss Physcomitrella patens and its ability to manufacture mammalian proteins could help to satisfy the large worldwide demand for therapeutic proteins. One well-known example is insulin, which enables diabetics to control their blood sugar level.
Nowadays, therapeutic proteins are mainly manufactured in mammalian cells, which are very expensive to culture. They need to be maintained at body temperature with a continuous supply of nutrients and oxygen, and the production process is costly. At present, global production capacity cannot match the demand. Because of the difficulties involved in handling them, production is possible only in industrialized countries.
In contrast, the moss Physcomitrella patens is comparatively undemanding. It needs water, a couple of nutrient salts and some light to allow it to flourish and produce proteins. This makes it convenient and simple to handle in a bioreactor, and, in the future, it might enable even less developed nations to satisfy their requirement for therapeutic proteins. However, further research will be needed before the moss can be used to produce therapeutic proteins on an industrial scale.
Journal reference:
Gitzinger et al. Functional cross-kingdom conservation of mammalian and moss (Physcomitrella patens) transcription, translation and secretion machineries. Plant Biotechnology Journal, 2009; 7 (1): 73 DOI: 10.1111/j.1467-7652.2008.00376.x
Adapted from materials provided by ETH Zurich.

How Cells Move: Cooperative Forces Boost Collective Mobility Of Cells

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ScienceDaily (May 11, 2009) — Research by scientists in Spain and their colleagues offers for the first time an experimental answer to the question of how cells move during biological processes as diverse as the development, metastasis, or regeneration of tissues.

The work addresses the issue of collective mobility of cells, that is to say, how cells are moved within tissues, and what is the prevalent form of movement inside living organisms.
"Research into collective cell mobility is very active due to the direct implications it has on fields such as embryologic development, organ regeneration, and cancer. For example, if we could find a way to control cell mobility during metastasis, cancer would be a curable disease in the majority of cases," says Dr. Xavier Trepat, senior researcher of the cellular and respiratory biomechanics group and researcher in the Department of Physiological Sciences at the University of Barcelona, and in the Networking Biomedical Research Centre for respiratory diseases (CIBER).
Up until now, scientists had proposed various mechanisms to explain collective cell migration. One hypothesis for example, suggests that the cells move collectively due to the existence of “leader” cells, which stretch out in the rest of the group, like a train pulls carriages behind it. Another hypothesis suggests that each cell moves independently to those around it, like cars on the motorway during a traffic jam, or like soldiers in a military parade. “We have rejected both these possibilities,” says Trepat.
According to his research, collective cell mobility is the result of a cooperative process in which each cell contributes to the movement of the group, stretching to those around it. “It is a mechanism similar to a tug-of-war game, in which two teams pull a rope by its extremes and the team that pulls the hardest wins. During the game, each player generates force and transmits it to the rope, so that the tension in the rope is the sum of the forces generated by each member of the team. Cells do the same. Each cell generates force to stretch to its neighbours in the direction of the movement» explains the researcher.
Journal reference:
Xavier Trepat, Michael R. Wasserman, Thomas E. Angelini, Emil Millet, David A. Weitz, James P. Butler & Jeffrey J. Fredberg. Physical forces during collective cell migration. Nature Physics, 2009; DOI: 10.1038/nphys1269
Adapted from materials provided by Universidad de Barcelona, via AlphaGalileo.

Sexually Transmitted Infections: Transistors Used To Detect Fungus Candida Albicans

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ScienceDaily (May 11, 2009) — The Nanosensors group from the Universidad Rovira i Virgili has created a biosensor, an electrical and biological device, which is able to selectively detect the Candida albicans yeast in very small quantities of only 50 cfu/ml (colony-forming units per millilitre).

"The technique uses field-effect transistors (electronic devices that contain an electrode source and a draining electrode connected to a transducer) based on carbon nanotubes and with Candida albicans-specific antibodies", Raquel A. Villamizar, lead author of the study said.
The Candida samples, which can be obtained from blood, serum or vaginal secretions, are placed directly on the biosensor, where the interaction between antigens and antibodies changes the electric current of the devices. This change is recorded and makes it possible to measure the amount of yeast present in a sample.
"Thanks to the extraordinary charge transference properties of the carbon nanotubes, the fungus detection process is direct, fast, and does not require the use of any marker", remarks Villamizar, who is co-author of a study that provides details of the biosensor and was published recently in the journal Sensors and Actuators B: Chemical.
To date, conventional diagnosis of Candida has been carried out using microbial cultures, serological tests, PCR molecular biology techniques (polymerase chain reactions used to amplify DNA), or immunoassays such as ELISA (Enzyme Linked Inmunoabsorbent Assay).
These techniques require long analysis times and sometimes give rise to false positives and negatives. ELISA also requires the use of markers (compounds that must be added to detect the presence of yeast by fluorescence and other techniques).
The new carbon nanotubes biosensor, however, "makes it possible to improve some of the quality parameters of the traditional methods, for example the speed and simplicity of measurements, and it is an alternative tool that could be used in routine sample analysis", explains Villamizar.
The researcher adds that by using this biosensor "it will be possible in future to obtain a rapid diagnosis of infection with this pathogen, which will help to ensure administration of the correct prophylactic treatments".
The Candida albicans fungus exists naturally in the skin, mouth, the mucous membranes lining the digestive tract, and the respiratory and genitourinary systems. This yeast can cause anything from simple mycosis of the skin to complicated cases of candidiasis. It is much more commonly found in patients suffering from immunodeficiency, tumours, diabetes and lymphomas, among other diseases.
Journal reference:
Villamizar et al. Improved detection of Candida albicans with carbon nanotube field-effect transistors. Sensors and Actuators B Chemical, 2009; 136 (2): 451 DOI: 10.1016/j.snb.2008.10.013
Adapted from materials provided by Plataforma SINC.

Deep in the Red: Using Infrared to Watch What Goes On in a Living Body

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An infrared version of the Nobel Prize-winning green fluorescent protein could make the technique even more powerful.

Fluorescent proteins, which are compounds that can absorb and then emit light, have become a powerful instrument in the cell biologist's toolkit—so powerful, in fact, that the discovery and development of green fluorescent proteins from jellyfish earned the 2008 Nobel Prize in Chemistry. (Here's a Q&A with one of the winners, Columbia University's Martin Chalfie, about his work.) These proteins have limitations, however: They need to be excited with the blue to orange part of the visible spectrum, at wavelengths of 495 to 570 nanometers. These wavelengths of light are too short to penetrate tissue very well, and so green fluorescent proteins are mainly used in test tube studies to watch cell division or to label certain cell types.
But one of the 2008 Nobelists, Roger Y. Tsien of the University of California, San Diego, and his U.C.S.D. colleagues report in today's issue of Science that they have developed a new fluorescent protein that could enable scientists to tag and visualize cellular activity as it happens inside a live animal. The protein, after absorbing light from the far-red part of the spectrum, shines in the near-infrared, at wavelengths of around 700 nanometers.
These longer wavelengths can penetrate mammalian tissue and even pass through bone. "Say you label a tumor with a green fluorescent protein, and if this labeled tumor is buried inside the animal, then you barely can get green fluorescence out," says lead researcher Xiaokun Shu. "But if you label this deeply buried tumor by infrared fluorescent proteins, you will get a stronger signal because infrared penetrates tissue more efficiently."
Tsien's group derived the infrared fluorescent protein from a hardy bacterium called Deinococcus radiodurans, famous for its ability to survive extreme environments. Bacteria do not actually use this class of proteins, called bacteriophytochromes, to emit light. "They use these bacteriophytochromes to control gene expression,” Shu says—the proteins convert absorbed light into energy to signal certain genes to turn on or off.
The initial challenge for researchers was to re-engineer the protein so that absorbed light would be re-emitted instead of being used as a source of power. They accomplished the feat by deleting the part of the protein that converts the absorbed light into chemical energy; as a result, this truncated and mutant form gave up its absorbed energy as an infrared glow. The scientists incorporated the engineered bacterial protein into mammalian cells—specifically, into the liver of a live mouse, which lit up with infrared light.
The finding paves the way for in vivo visualization of a wide range of biochemical processes and internal organs in animals. (Its use in humans is unlikely, as it would require gene therapy and the ethically dubious transplantation of bacterial genes into humans.)
“This is so important," comments David James, a cell biologist from Australia's Garvan Institute in Sydney, "because a lot of knowledge at the moment is confined to individual cells grown on a glass coverslip," leaving open the question of whether that knowledge "is transferable to an animal." The infrared version could also solve the problem of naturally occuring fluorescence from other biological molecules, which tend to glow at wavelengths similar to conventional fluorescent protein markers and thereby create a lot of “background noise," James says.

But even greater potential lies in harnessing the bacteriophytochrome's original function, namely, powering gene expression. It should be feasible, Tsien thinks, to put back in the signal-controlling properties of the phytochrome. Then it could be possible with animals to “switch on genes and control biochemistry" with light, he says.
For example, you want to explore the effects on mouse behavior of switching on a particular gene that controls some aspect of brain function, but, thankfully for the mouse, you do not want to open up its skull or stick a needle in its brain. "If the infrared fluorescent protein can be made to turn back into an infrared phytochrome, you could have the switch all ready and just waiting for enough infrared light," Tsien speculates. Because infrared light can penetrate the skull, it can reach the phytochrome and remotely switch the gene on, resulting in observable changes in the mouse's behavior.
It's the next evolutionary step for fluorescent proteins, remarks Tsien, who believes that phytochromes represent a class of proteins with enormous potential. If he's correct, then in the coming years, expect more scientists to see the (infrared) light.

By Bianca Nogrady

Level Of Cellular Stress Determines Longevity Of Retinal Cells

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ScienceDaily (May 9, 2009) — Stress can be adaptive. It can make you sharper, help you focus and it can even improve your performance. But too much of it can tax cells to the point where they can no longer cope and slowly self-destruct.

Scientists at Rockefeller University now show that when the protein-making factory of the cell is exposed to moderate stress, neurons in the fruit fly retina and other cells not only resist death but also shore up their defenses against damaging free radicals and ultraviolet radiation.
The finding sheds light on the molecular mechanism by which cells compute their fate, and may point to therapeutic targets that protect against or delay the onset of neurodegeneration.
In their work, Hermann Steller, head of the Strang Laboratory of Apoptosis and Cancer Biology, and César Mendes, a former graduate student in the lab, genetically turned on three known cell death signals in fruit fly retinal cells, each of which directs the cell to undergo a process of controlled suicide. But when they knocked out a gene called NinaA, they saw that the cells halt their descent toward death. “The loss of NinaA tosses the cell a lifeline,” says Steller, who is also a Howard Hughes Medical Institute investigator and Strang Professor at Rockefeller. “It can send a pro-life signal that tells the neuron, ‘give repair a chance.’”
NinaA encodes for a protein that folds rhodopsin, the light-absorbing molecule that allows us to see color, into its proper shape. In cells that lack NinaA, rhodopsin doesn’t fold properly and starts to accumulate in the endoplasmic reticulum (ER), the cellular factory where proteins are modified, packaged and shipped to their proper destinations. In response to this accumulation, called ER stress, the cells activate a repair pathway to fix the problem, either derailing or halting the cell’s death cascade.
“Unlike other studies that use pharmacology and overexpression systems that quickly overwhelm cells and drive them to death, we managed to induce a more physiological and nonlethal level of ER stress by removing one or both copies of the NinaA gene,” says Mendes, who is now a postdoc at Columbia University. “This is one of the beauties of Drosophila as a model system — the capability to finely tune genetic dosage.”
The team, including Bertrand Mollereau, who is now a professor at the École Normale Supérieure de Lyon in France, believes that a mechanism underlying this protection may involve antioxidant genes that protect retinal neurons from ultraviolet radiation and free radicals. When these neurons are exposed to mild ER stress, the team showed that they upregulate genes that shield them from the substances’ harmful effects. “As in neurodegenerative diseases, when photoreceptor neurons die, they may never be replaced,” explains graduate student Alexis Gambis, who also worked on the project. “The antioxidant upregulation is one way neurons have evolved to protect themselves from exogenous stress and it’s especially important in the eye, which receives damaging UV energy from the sun.”
But while the loss of NinaA delays the cell death cascade, this protection is lost when rhodopsin is absent from retinal cells, suggesting that it’s actually the loss of NinaA and the resulting ER stress, and not the loss of rhodopsin’s function, that makes the cells live longer. The finding further disentangles the molecular decision points at which cells choose between life and death under ER stress, which has been linked to a host of human diseases, including Alzheimer’s, diabetes and cancer. “Cells don’t make these decisions lightly,” says Steller. “They have had millions of years to figure how to direct their fate.”
Journal reference:
Mendes et al. ER stress protects from retinal degeneration. The EMBO Journal, April 2, 2009; DOI: 10.1038/emboj.2009.76
Adapted from materials provided by Rockefeller University.

Scientists Shed Light On Inner Workings Of Human Embryonic Stem Cells

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ScienceDaily (May 7, 2009) — Scientists at UC Santa Barbara have made a significant discovery in understanding the way human embryonic stem cells function.

They explain nature's way of controlling whether these cells will renew, or will transform to become part of an ear, a liver, or any other part of the human body. The study is reported in the May 1 issue of the journal Cell.
The scientists say the finding bodes well for cancer research, since tumor stem cells are the engines responsible for the growth of tumors. The discovery is also expected to help with other diseases and injuries. The study describes nature's negative feedback loop in cell biology.
"We have found an element in the cell that controls 'pluripotency,' that is the ability of the human embryonic stem cell to differentiate or become almost any cell in the body," said senior author Kenneth S. Kosik, professor in the Department of Molecular, Cellular & Developmental Biology. Kosik is also co-director and Harriman Chair in Neuroscience Research of UCSB's Neuroscience Research Institute.
"The beauty and elegance of stem cells is that they have these dual properties," said Kosik. "On the one hand, they can proliferate –– they can divide and renew. On the other hand, they can also transform themselves into any tissue in the body, any type of cell in the body."
The research team includes James Thomson, who provided an important proof to the research effort. Thomson, an adjunct professor at UCSB, is considered the "father of stem cell biology." Thomson pioneered work in the isolation and culture of non-human primate and human embryonic stem cells. These cells provide researchers with unprecedented access to the cellular components of the human body, with applications in basic research, drug discovery, and transplantation medicine.
With regard to human embryonic stem cells, Kosik explained that for some time he and his team have been studying a set of control genes called microRNAs. "To really understand microRNAs, the first step is to remember the central dogma of biology ––DNA is the template for RNA and RNA is translated to protein. But microRNAs stop at the RNA step and never go on to make a protein.
"The heart of the matter is that before this paper, we knew that if you want to maintain a pluripotent state and allow self-renewal of embryonic stem cells, you have to sustain levels of transcription factors," said Kosik. "We also knew that stem cells transition to a differentiated state when you decrease those factors. Now we know how that happens a little better."
The new research shows that a microRNA –– a single-stranded RNA whose function is to decrease gene expression –– lowers the activity of three key ingredients in the recipe for embryonic stem cells. This microRNA is known as miR-145. The discovery may have implications for improving the efficiency of methods designed to reprogram differentiated cells into embryonic stem cell-like cells.
As few as three or four genes can make cells pluripotent. "We know what these genes are," Kosik said. That information was used recently for one of the most astounding breakthroughs of biology of the last couple of years –– the discovery of induced pluripotent skin cells.
"You can take a cell, a skin cell, or possibly any cell of the body, and revert it back to a stem cell," Kosik said. "The way it's done, is that you take the transcription factors that are required for the pluripotent state, and you get them to express themselves in the skin cells; that's how you can restore the embryonic stem cell state. You clone a gene, you put it into what's called a vector, which means you put it into a little bit of housing that allows those genes to get into a cell, then you shoot them into a stem cell. Next, when those genes –– those very critical pluripotent cell genes –– get turned on, the skin cell starts to change, it goes back to the embryonic pluripotent stem cell state."
The researchers explained that a rise in miR-145 prevents human embryonic stem cells' self-renewal and lowers the activity of genes that lend stem cells the capacity to produce other cell types. It also sends the cells on a path toward differentiation. In contrast, when miR-145 is lost, the embryonic stem cells are prevented from differentiating as the concentrations of transcription factors rise.
They also show that the control between miR-145 and the "reprogramming factors" goes both ways. The promoter for miR-145 is bound and repressed by a transcription factor known as OCT4, they found.
"It's a beautiful double negative feedback loop," Kosik said. "They control each other. That is the essence of this work."
Because there is typically less "wiggle room" in the levels of microRNA compared to mRNA, further studies are needed to quantify more precisely the copy numbers of miR-145 and its targets, to figure out exactly how this layer of control really works, Kosik said.
Kosik credits the lion's share of this discovery to first author Na Xu, a postdoctoral fellow who is also supported by the California Institute for Regenerative Medicine (CIRM). "Na Xu deserves enormous credit for this work," said Kosik. "She performed nearly every experiment in the paper and was the major contributor to the ideas in the paper." Meanwhile, Thales Papagiannakopoulos, a graduate student working in the Kosik lab, was very generous in helping Na Xu with one of the experiments. He helped with one of several proofs that showed that the targets of miR-145 are the three transcription factors that are being reported, explained Kosik.
Thomson provided one of several proofs for the control point of miR-145 expression, said Kosik.
Journal reference:
Na Xu, Thales Papagiannakopoulos, Guangjin Pan, James A. Thomson, and Kenneth S. Kosik. MicroRNA-145 Regulates OCT4, SOX2, and KLF4 and Represses Pluripotency in Human Embryonic Stem Cells. Cell, 2009; DOI: 10.1016/j.cell.2009.02.038
Adapted from materials provided by University of California - Santa Barbara.

Building The Lymphatic Drainage System

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ScienceDaily (May 6, 2009) — Our bodies' tissues need continuous irrigation and drainage. Blood vessels feeding the tissues bring in the fluids, and drainage occurs via the lymphatic system. While much is known about how blood vessels are built, the same was not true for lymph vessels. Now though, Norrmén et al. have identified two of the lead engineers that direct drainage construction in the mouse embryo.

The engineers are the transcription factors, Foxc2 and NFATc1. Foxc2 had been implicated in lymph vessel development already, but Norrmén and colleagues have now found that the factor specifically regulates a late stage of lymph development when large, valve-containing vessels arise from more primitive capillaries.
Foxc2 built the lymph vessel valves with the help of NFATc1, which was a known heart valve engineer. Norrmén and colleagues also showed that Foxc2 and NFATc1 physically interact and that many DNA binding sites for the two transcription factors are closely linked. This latter finding generated a long list of target genes that might be controlled by the two factors.
The team now plans to investigate these targets as well as to work out the upstream molecular pathways controlling Foxc2 and NFATc1. Whatever the mechanisms, if the team can show that Foxc2 and NFATc1 also prompt lymph vessel regeneration in adults, boosting these factors could help patients with lymph drainage problems – including those that have suffered extensive tissue injuries, or have had lymph nodes removed as part of cancer treatment.
The study will be published online April 27 and will appear in the May 4 print issue of the Journal of Cell Biology.
Journal reference:
Norrmen et al. FOXC2 controls formation and maturation of lymphatic collecting vessels through cooperation with NFATc1. The Journal of Cell Biology, 2009; 185 (3): 439 DOI: 10.1083/jcb.200901104
Adapted from materials provided by Rockefeller University Press, via EurekAlert!, a service of AAAS.

Nanoneedle Is Small In Size, But Huge In Applications

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ScienceDaily (May 6, 2009) — Researchers at the University of Illinois have developed a membrane-penetrating nanoneedle for the targeted delivery of one or more molecules into the cytoplasm or the nucleus of living cells. In addition to ferrying tiny amounts of cargo, the nanoneedle can also be used as an electrochemical probe and as an optical biosensor.

"Nanoneedle-based delivery is a powerful new tool for studying biological processes and biophysical properties at the molecular level inside living cells," said
Min-Feng Yu, a professor of mechanical science and engineering and corresponding author of a paper accepted for publication in Nano Letters, and posted on the journal's Web site.
In the paper, Yu and collaborators describe how they deliver, detect and track individual fluorescent quantum dots in a cell's cytoplasm and nucleus. The quantum dots can be used for studying molecular mechanics and physical properties inside cells.
To create a nanoneedle, the researchers begin with a rigid but resilient boron-nitride nanotube. The nanotube is then attached to one end of a glass pipette for easy handling, and coated with a thin layer of gold. Molecular cargo is then attached to the gold surface via "linker" molecules. When placed in a cell's cytoplasm or nucleus, the bonds with the linker molecules break, freeing the cargo.
With a diameter of approximately 50 nanometers, the nanoneedle introduces minimal intrusiveness in penetrating cell membranes and accessing the interiors of live cells.
The delivery process can be precisely controlled, monitored and recorded – goals that have not been achieved in prior studies.
"The nanoneedle provides a mechanism by which we can quantitatively examine biological processes occurring within a cell's nucleus or cytoplasm," said Yang Xiang, a professor of molecular and integrative physiology and a co-author of the paper. "By studying how individual proteins and molecules of DNA or RNA mobilize, we can better understand how the system functions as a whole."
The ability to deliver a small number of molecules or nanoparticles into living cells with spatial and temporal precision may make feasible numerous new strategies for biological studies at the single-molecule level, which would otherwise be technically challenging or even impossible, the researchers report.
"Combined with molecular targeting strategies using quantum dots and magnetic nanoparticles as molecular probes, the nanoneedle delivery method can potentially enable the simultaneous observation and manipulation of individual molecules," said Ning Wang, a professor of mechanical science and engineering and a co-author of the paper.
Beyond delivery, the nanoneedle-based approach can also be extended in many ways for single-cell studies, said Yu, who also is a researcher at the Center for Nanoscale Chemical-Electrical-Mechanical Manufacturing Systems. "Nanoneedles can be used as electrochemical probes and as optical biosensors to study cellular environments, stimulate certain types of biological sequences, and examine the effect of nanoparticles on cellular physiology."
With Wang, Xiang and Yu, co-authors of the paper are graduate student Kyungsuk Yum and postdoctoral research associate Sungsoo Na. Yu and Wang are affiliated with the university's Beckman Institute. Wang is also affiliated with the department of bioengineering and with the university's Micro and Nanotechnology Laboratory.
The Grainger Foundation, National Science Foundation and National Institutes of Health funded the work.
Journal reference:
Kyungsuk Yum, Sungsoo Na, Yang Xiang, Ning Wang, Min-Feng Yu. Mechanochemical Delivery and Dynamic Tracking of Fluorescent Quantum Dots in the Cytoplasm and Nucleus of Living Cells. Nano Letters, 2009; 090414110255042 DOI: 10.1021/nl901047u
Adapted from materials provided by University of Illinois at Urbana-Champaign.