CellNEWS

New Strategy for Mending Broken Hearts?

2009-10-14T05:42:00.005+02:00

Patch created to repair damaged heart tissue
Wednesday, 14 October 2009

By mimicking the way embryonic stem cells develop into heart muscle in a lab, Duke University bioengineers believe they have taken an important first step toward growing a living "heart patch" to repair heart tissue damaged by disease.

In a series of experiments using mouse embryonic stem cells, the bioengineers used a novel mould of their own design to fashion a three-dimensional "patch" made up of heart muscle cells, known as cardiomyocytes. The new tissue exhibited the two most important attributes of heart muscle cells – the ability to contract and to conduct electrical impulses. The mould looks much like a piece of Chex cereal in which researchers varied the shape and length of the pores to control the direction and orientation of the growing cells.

The researchers grew the cells in an environment much like that found in natural tissues. They encapsulated the cells within a gel composed of the blood-clotting protein fibrin, which provided mechanical support to the cells, allowing them to form a three-dimensional structure. They also found that the cardiomyocytes flourished only in the presence of a class of "helper" cells known as cardiac fibroblasts, which comprise as much as 60 percent of all cells present in a human heart.

"If you tried to grow cardiomyocytes alone, they develop into an unorganized ball of cells," said Brian Liau, graduate student in biomedical engineering at Duke's Pratt School of Engineering. Liau, who works in the laboratory of assistant professor Nenad Bursac, presented the results of his latest experiments during the annual scientific sessions of the Biomedical Engineering Society in Pittsburgh.

"We found that adding cardiac fibroblasts to the growing cardiomyocytes created a nourishing environment that stimulated the cells to grow as if they were in a developing heart," Liau said.

"When we tested the patch, we found that because the cells aligned themselves in the same direction, they were able to contract like native cells. They were also able to carry the electrical signals that make cardiomyocytes function in a coordinated fashion."

"The addition of fibroblasts in our experiments provided signals that we believe are present in a developing embryo," Liau said. The need for helper cells is not uncommon in mammalian development. For example, he explained, nerve cells need "sheathe" cells known as glia in order to develop and function properly.

Bursac believes that the latest experiments represent a proof-of-principle advance, but said there are still many hurdles to overcome before such patches could be implanted into humans with heart disease.

"While we were able to grow heart muscle cells that were able to contract with strength and carry electric impulses quickly, there are many other factors that need to be considered," Bursac said.

"The use of fibrin as a structural material allowed us to grow thicker, three-dimensional patches, which would be essential for the delivery of therapeutic doses of cells. One of the major challenges then would be establishing a blood vessel supply to sustain the patch."

The researchers plan to test their model using non-embryonic stem cells. For use in humans, this is important for many reasons, both scientifically and ethically, Bursac said. Recent studies have demonstrated that some cells from human adults have the ability to be reprogrammed to become similar to embryonic stem cells.

"Human cardiomyocytes tend to grow a lot slower than those of mice," Bursac said.

"Since it takes nine months for the human heart to complete development, we need to find a way to get the cells to grow faster while maintaining the same essential properties of native cells."

If they could use a patient's own cells, the patch would also evade an immune system reaction, Bursac added.
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ZenMaster

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Liver Cells Grown from Patients' Skin Cells

2009-10-14T05:27:00.003+02:00

Treatment of liver diseases possible
Wednesday, 14 October 2009

Scientists at The Medical College of Wisconsin in Milwaukee have successfully produced liver cells from patients' skin cells opening the possibility of treating a wide range of diseases that affect liver function. The study was led by Stephen A. Duncan, D. Phil., Marcus Professor in Human and Molecular Genetics, and professor of cell biology, neurobiology and anatomy, along with postdoctoral fellow Karim Si-Tayeb, Ph.D., and graduate student Ms. Fallon Noto.

"This is a crucial step forward towards developing therapies that can potentially replace the need for scarce liver transplants, currently the only treatment for most advanced liver disease," says Dr. Duncan.

Liver disease is the fourth leading cause of death among middle aged adults in the United States. Loss of liver function can be caused by several factors, including genetic mutations, infections with hepatitis viruses, by excessive alcohol consumption, or chronic use of some prescription drugs. When liver function goes awry it can result in a wide variety of disorders including diabetes and atherosclerosis and in many cases is fatal.

The Medical College research team generated patient–specific liver cells by first repeating the work of James Thomson and colleagues at University of Wisconsin-Madison who showed that skin cells can be reprogrammed to become cells that resemble embryonic stem cells. They then tricked the skin–derived pluripotent stem cells into forming liver cells by mimicking the normal processes through which liver cells are made during embryonic development. Pluripotent stem cells are so named because of their capacity to develop into any one of eh more than 200 cell types in the human body.

At the end of this process, the researchers found that they were able to very easily produce large numbers of relatively pure liver cells in laboratory culture dishes.

"We were excited to discover that the liver cells produced from human skin cells were able to perform many of the activities associated with healthy adult liver function and that the cells could be injected into mouse livers where they integrated and were capable of making human liver proteins," says Dr. Duncan.

Several studies have shown that liver cells generated from embryonic stem cells could potentially be used for therapy. However, the possible use of such cells is limited by ethical considerations associated with the generation of embryonic stem cells from preimplantation embryos and the fact that embryonic stem cells do not have the same genetic make-up as the patient.

Although the investigations are still at an early stage the researchers believe that the reprogrammed skin cells could be used to investigate and potentially treat metabolic liver disease. The liver may be particularly suitable for stem-cell based therapies because it has a remarkable capacity to regenerate. It is interesting to note that the regenerative nature of the liver was referenced in the ancient Greek tale of Prometheus. When Prometheus was caught stealing the gift of fire from Zeus, he was punished by having his liver eaten daily by an eagle. This provided the eagle with an everlasting meal because each night the liver of Prometheus would re-grow.

The liver is a central regulator of the body's metabolism and is responsible for controlling sugar and cholesterol levels, secretion of a variety of hormones, production of blood clotting factors, and has an essential role in preventing toxins from damaging other organs in the body.

It is possible that in the future a small piece of skin from a patient with loss of liver function could be used to produce healthy liver cells, replacing the diseased liver with normal tissue.

Recently, the National Institutes of Health's National Institute of Diabetes and Digestive and Kidney Diseases through the American Recovery and Reinvestment Act have provided the MCW researchers, in collaboration with Markus Grompe, M.D., at the Oregon Health and Science University, a $1 million research grant to pursue the possibility of using reprogrammed skin cells to study and treat metabolic liver disease. Using this support, as well as donations from individuals throughout Milwaukee, the Medial College researchers are currently producing reprogrammed cells from patients suffering from diabetes, hyperlipidemia, and hypercholesterolemia in an effort to identify new treatments for these diseases.
.........

ZenMaster

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Jumping Genes, Gene Loss and Genome Dark Matter

2009-10-12T23:01:00.004+02:00

New map of copy number variation in the human genome is a resource for human genetics
Monday, 12 October 2009

In research published last week by Nature, an international team describes the finest map of changes to the structure of human genomes and a resource they have developed for researchers worldwide to look at the role of these changes in human disease. They also identify 75 'jumping genes' - regions of our genome that can be found in more than one location in some individuals.

However, the team cautions that they have not found large numbers of candidates that might alter susceptibility to complex diseases such as diabetes or heart disease among the common structural variants. They suggest strategies for finding this 'dark matter' of genetic variation.

Human genomes differ because of single-letter variations in the genetic code and also because whole segments of the code might be deleted or multiplied in different human genomes. These larger, structural differences are called copy number variants (CNVs).

The new research to map and characterize CNVs is of a scale and a power unmatched to date, involving hundreds of human genomes, billions of data points and many thousands of CNVs.

"This study is more than ten times as powerful as our first map, published three years ago," explains Dr Matt Hurles from the Wellcome Trust Sanger Institute and a leader on the project, "and much more detailed than any other. Importantly, we have also assigned the CNVs to a specific genetic background so that they can be readily examined in disease studies carried out by others, such as the Wellcome Trust Case Control Consortium.”

"Nevertheless, we have not found large numbers of common CNVs that we can tie strongly to disease. There remains much to be discovered and much to understand and our freely available genotyped collection will drive that discovery."

The results show that any two genomes differ by more than 1000 CNVs, or around 0.8% of a person's genome sequence. Most of these CNVs are deletions, with a minority being duplications.

Chromosomes are shown colour-coded in the outermost circle. Inside are lines connecting the origin and the new location (where known) of 58 out of 75 putative inter-chromosomal duplications, coloured according to their chromosome of origin. Credit: Jan Aerts, Wellcome Trust Sanger Institute.

Two consequences are particularly striking in this study of apparently healthy people. First, 75 regions have jumped around in the genomes of these samples; second, more than 250 genes can lose one of the two copies in our genome without obvious consequences and a further 56 genes can fuse together potentially to form new composite genes.

"This paper detailing common CNVs in different world populations, and providing the first glimpse into evolutionary biology of such class of human variation, is unquestionably one of the most important advances in human genome research since the completion of a reference human genome," says Professor James R. Lupski, Vice Chair of the department of Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas.

"It complements the cataloguing of single nucleotide variation delineated in the HapMap Project and will both enable some new approaches to, and further augment other studies of, basic human biology relevant to health and disease."

"The genetic 'blueprint' of humans is the human genome," says Sir Mark Walport, Director of the Wellcome Trust.

"But we are each unique as individuals, shaped by variation in both genome and environment. Understanding the variation amongst human genomes is key to understanding the inherited differences between each of us in health and disease. A whole new dimension has been added to our understanding of variation in the human genome by the identification of copy number variants."

The results also give, for the first time, a minimum measure of the rate of CNV mutation: at least one in 17 children will have a new CNV. In many cases, that CNV will have no obvious clinical consequences. However, for some the effects are severe. In those cases the data are captured in the DECIPHER database, a repository of clinical information about CNVs designed to aid the diagnosis of rare disorders in young children.

However, CNVs are not only about here and about now; they are also ancient legacies of how our ancestors adapted to their environments. Among the most impressive variations between populations are CNVs that modify the activity of the immune system, known to be evolving rapidly in human populations, and genes implicated in muscle function. The researchers propose that the consequences of these CNVs can be dissected in population studies.

The team scanned 42 million locations on the genomes of 40 people, half of European ancestry and half of West-African ancestry. The scale of the method meant they could detect CNVs as small as 450 bases occurring in one in 20 individuals.

However, the researchers concede that their map of common variants will not account for much of the 'dark matter' of the genome - the missing heritability where, despite diligent searches, genetic variants have not been found for common disease.

"CNV studies have made huge advances in the past few years, but we are still looking only at the most common CNVs," explains Dr Steve Scherer of the Hospital for Sick Children, Toronto.

"We suspect that there are many CNVs that have real clinical consequences that occur in perhaps one in 50 or one in 100 people - below the level we have detected.”

"Success in the hunt for the missing genetic causes of common disease has become possible in the last few years and we expect to find more as higher resolution searches become possible."

The research group have maximized the value of their research by not only mapping the CNVs, but by also genotyping them - assigning them to a specific genetic background that makes them readily useful in wider genetic studies, such as the Wellcome Trust Case Control Consortium.

"We were determined to develop not only the map, but also to provide the resources that help other researchers and clinical cytogeneticists most rapidly use our CNV results," comments Dr Charles Lee, one of the project leaders from Brigham and Women's Hospital and Harvard Medical School in Boston, USA.

"Already, the data that we have generated is benefiting other large-scale studies such as the 1000 Genomes Projects as well as making an enormous difference in the accurate interpretation of clinical genetic diagnoses.”

"Nonetheless, the human CNV story is far from over."
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ZenMaster

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Major Improvements Made in Engineering Heart Repair Patches from Stem Cells

2009-10-12T22:45:00.003+02:00

Pre-formed blood vessels in patches connect to rodents' heart circulation
Monday, 12 October 2009

University of Washington (UW) researchers have succeeded in engineering human tissue patches free of some problems that have stymied stem-cell repair for damaged hearts.

The disk-shaped patches can be fabricated in sizes ranging from less than a millimetre to a half-inch in diameter. Until now, engineering tissue for heart repair has been hampered by cells dying at the transplant core, because nutrients and oxygen reached the edges of the patch but not the centre. To make matters worse, the scaffolding materials to position the cells often proved to be harmful.

Heart tissue patches composed only of heart muscle cells could not grow big enough or survive long enough to take hold after they were implanted in rodents, the researchers noted in their article, published last month in the Proceedings of the National Academy of Sciences. The researchers decided to look at the possibility of building new tissue with supply lines for the oxygen and nutrients that living cells require.

The scientists testing this idea are from the UW Center for Cardiovascular Biology and the UW Institute for Stem Cell and Regenerative Medicine, under the guidance of senior author Dr. Charles "Chuck" Murry, professor of pathology and bioengineering. The lead author is Dr. Kelly R. Stevens, a UW doctoral student in bioengineering who came up with solutions to the problems observed in previous grafts. The study is part of a collaborative tissue engineering effort called BEAT (Biological Engineering of Allogeneic Tissue).

Stevens and her fellow researchers added two other types of cells to the heart muscle cell mixture. These were cells similar to those that line the inside of blood vessels and cells that provide the vessel's muscular support. All of the heart muscle cells were derived from embryonic stem cells, while the vascular cells were derived from embryonic stem cells or a variety of more mature sources such as the umbilical cord. The resulting cell mixture began forming a tissue containing tiny blood vessels.

"These were rudimentary blood vessel networks like those seen early in embryonic development," Murry said.

In contrast to the heart muscle cell-only tissue, which failed to survive transplantation and which remained apart from the rat's heart circulatory system, the pre-formed vessels in the mixed-cell tissue joined with the rat's heart circulatory system and delivered rat blood to the transplanted graft.

"The viability of the transplanted graft was remarkably improved," Murry observed.

"We think the gain in viability is due to the ability for the tissue to form blood vessels."

Equally as exciting, the scientists observed that the patches of engineered tissue actively contracted. Moreover, these contractions could be electronically paced, up to what would translate to 120 beats per minute. Beyond that point, the tissue patch did not relax fully and the contractions weakened. However, the average resting adult heart pulses about 70 beats per minute. This suggests that the engineered tissue could, within limits, theoretically keep pace with typical adult heart muscle, according to the study authors.

Another physical quality that made the mixed-cell tissue patches superior to heart muscle-cell patches was their mechanical stiffness, which more closely resembled human heart muscle. This was probably due to the addition of supporting cells, which created connective tissues. Passive stiffness allows the heart to fill properly with blood before it contracts.

When the researchers implanted these mixed celled, pre-vascularised tissue patches into rodents, the patches grew into cell grafts that were ten times larger than the too-small results from tissue composed of heart muscle cells only. The rodents were bred without an immune system that rejects tissue transplants.

Murry noted that these results have significance beyond their contribution to the ongoing search for ways to treat heart attack damage by regenerating heart tissue with stem cells.

The study findings, he observed, suggest that researchers consider including blood vessel-generating and vascular-supporting elements when designing human tissues for certain other types of regenerative therapies unrelated to heart disease.

One of the major obstacles still to be overcome is the likelihood that people's immune systems would reject the stem transplant unless they take medications for the rest of their lives to suppress this reaction. Murry hopes someday that scientists would be able to create new tissues from a person's own cells.

"Researchers can currently turn human skin cells back to stem cells, and then move them forward again into other types of cells, such as heart muscle and blood vessel cells," Murry said.

"We hope this will allow us to build tissues that the body will recognize as 'self.'"

While the clinical application of tissues engineered from stem cells in treating hearts damaged from heart attacks or birth defects is still in the future, the researchers believe progress has been made. This study showed that researchers could create the first entirely human heart tissue patch from human embryonic cell-derived heart muscle cells, blood vessel lining cells and fibre-producing cells, and successfully engraft the tissue into an animal.

Future studies will try to move heart cell regeneration closer toward clinical usefulness, according to Murry and his research team. They forecast that such research would include testing other sources of human cells and developing techniques to create bigger patches for treating larger animals through surgical transplantation or through catheter delivered injections.

Lastly, they concluded, researchers would need to test whether tissue patches actually improve physical functioning after implantation in damaged hearts.
.........

ZenMaster

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Enhanced Stem Cells Promote Tissue Regeneration

2009-10-12T22:37:00.001+02:00

Enhanced Stem Cells Promote Tissue Regeneration
Monday, 12 October 2009

Massachusetts Institute of Technology engineers have boosted stem cells' ability to regenerate vascular tissue (such as blood vessels) by equipping them with genes that produce extra growth factors (naturally occurring compounds that stimulate tissue growth). In a study in mice, the researchers found that the stem cells successfully generated blood vessels near the site of an injury, allowing damaged tissue to survive.

Why it matters: Stem cells hold great potential as a way to promote tissue regeneration. However, this approach has been limited because stem cells don't produce enough growth factors after transplantation. The researchers' new super-charged stem cells could be used to treat an infarction (death of tissue caused by blockage of the blood supply, by a clot or another obstruction), or to induce blood supply for engineered tissues.

Methods: After removing stem cells from mouse bone marrow, the researchers used specially developed nanoparticles to deliver the gene for the growth factor VEGF (vascular endothelial growth factor). The stem cells were then implanted into damaged tissue areas. These nanoparticles, which the MIT team has also tested to deliver cancer treatments, are believed to be safer than the viruses often used for gene delivery.

Next steps: Though the results are promising, the technique needs more improvements before any human trials can begin, says Daniel Anderson, a senior author of the paper.
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ZenMaster

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Human Embryonic Stem Cells Reverse Retinal Degeneration

2009-10-12T04:30:00.001+02:00

Human Embryonic Stem Cells Reverse Retinal Degeneration
Monday, 12 October 2009

A new study reports that transplanted pigment-containing visual cells derived from human embryonic stem cells (hESCs) successfully preserved structure and function of the specialized light-sensitive lining of the eye (known as the retina) in an animal model of retinal degeneration. The findings, published by Cell Press in the October 2nd issue of the journal Cell Stem Cell, represent an exciting step towards the future use of cell replacement therapies to treat devastating degenerative eye diseases that cause millions of people worldwide to lose their sight.

The retinal pigment epithelium (RPE) is a layer of pigmented cells sandwiched between the visual retinal cells, called photoreceptors, and the nourishing blood vessels at the back of the eye. The RPE provides essential support to the retinal photoreceptors and is critical for normal vision. Deterioration of the RPE plays a central role in the progression of diseases such as age-related macular degeneration and sub-types of retinitis pigmentosa. These conditions are associated with a progressive loss of vision that often leads to blindness.

"Although there are a variety of therapeutic approaches under development to delay the degenerative process, the grim reality is that many patients eventually lose their sight," explains Dr. Benjamin Reubinoff, a senior author of the study.

"Cell therapy to replenish the degenerating RPE cells may potentially halt disease progression." Dr. Reubinoff and Dr. Eyal Banin who led the study, with their colleagues from Hadassah-Hebrew University Medical Center in Jerusalem, developed conditions to guide hESCs to differentiate into functional RPE-like cells in the laboratory.

The researchers found that nicotinamide (vitamin B3, NIC) and Activin A, an important growth factor, promoted differentiation of hESCs towards an RPE fate. The hESC-derived RPE-like cells, which could be identified by their characteristic black pigment, exhibited multiple biological properties and genetic markers that define authentic RPE cells. Further, the cells successfully delayed deterioration of retinal structure and function when they were transplanted into an animal model of retinal degeneration caused by RPE dysfunction.

Taken together, the results demonstrate that NIC and Activin A promoted the differentiation of hESCs towards an RPE fate. The hESC-derived cells exhibited the defining characteristics associated with RPE and successfully rescued the retina when transplanted into an animal model of retinal degeneration.

"Our findings are an important step towards the potential future use of hESCs to replenish RPE in blinding diseases," concludes Dr. Banin.
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ZenMaster

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Umbilical Cord Blood Source for Stem Cells

2009-10-12T04:17:00.003+02:00

Readily Available and Patient-specific Stem Cells
Monday, 12 October 2009

Umbilical cord blood cells can successfully be reprogrammed to function like embryonic stem cells, setting the basis for the creation of a comprehensive bank of tissue-matched, cord blood-derived induced pluripotent stem (iPS) cells for off-the-shelf applications, report researchers at the Salk Institute for Biological Studies and the Center for Regenerative Medicine in Barcelona, Spain.

"Cord blood stem cells could serve as a safe, "ready-to-use" source for the generation of iPS cells, since they are easily accessible, immunologically immature and quick to return to an embryonic stem cell-like state," says Juan-Carlos Izpisúa Belmonte, Ph.D., a professor in the Salk's Gene Expression Laboratory, who led the study published in the October issue of the journal Cell Stem Cell.

Worldwide, there are already more than 400,000 cord blood units banked along with immunological information. Due to their early origin, cells found in umbilical cord blood contain a minimal number of somatic mutations and possess the immunological immaturity of newborn cells, allowing the HLA donor-recipient match to be less than perfect without the risk of immune rejection of the transplant.

Human leukocyte antigen (HLA) typing is used to match patients and donors for bone marrow or cord blood transplants. HLAs are special surface markers found on most cells in the body and help the immune system to distinguish between "self" and "non-self."

"Selecting common HLA haplotypes from among already banked cord blood units to create iPS cell would significantly reduce the number of cell lines needed to provide a HLA match for a large percentage of the population," says Izpisúa Belmonte.

The endodermal layer, identified by markers AFP and FoxA2, will give rise to the digestive tract, lungs and bladder. Credit: Courtesy of Juan-Carlos Izpisúa Belmonte, from Cell Stem Cell, Oct. 1, 2009.

Since the first adult cells were converted into iPS cells, they have generated a lot excitement as an uncontroversial alternative to embryonic stem cells and as a potential source for patient-specific stem cells. Unfortunately, taking a patient's cells back in time is not only costly, but could be difficult when those cells are needed right away to mend injured spinal cords or treat acute diseases, and outright impossible when the effects of aging or chronic disease have irrevocably damaged the pool of somatic cells.

"Patient-specific iPS lines have been advocated as a theoretically ideal clinical option to regenerate tissue but from a practical and cost-benefit aspect, this approach may not be feasible," says Izpisúa Belmonte. He hopes that the "large scale production and banking of cord blood-derived iPS lines in a publically available network could be a viable alternative for future clinical applications."

With this in mind, Belmonte and his colleagues set out to transform hematopoietic stem cells isolated from cord blood into iPS cells. They not only successfully converted them using only two out of the four most commonly used factors — Oct4 and Sox2 — but also in less time than any other previously published methodology require. No matter, whether the researchers started with freshly collected cord blood or previously frozen samples, the resulting iPS cells were indistinguishable from human embryonic stem cells.

The mesodermal layer, identified by ASM, will form bones, muscles, connective tissue and the middle layer of the skin. Credit: Courtesy of Juan-Carlos Izpisúa Belmonte , from Cell Stem Cell, Oct. 1, 2009.

"The population of cord blood cells used for reprogramming express reprogramming/stem cell factors at higher levels than those found in other adult somatic cells, which could explain why cord blood cells can be reprogrammed with less factors and in less time," says Izpisúa Belmonte.

"It's almost like they are already half-way there."

In addition, the cord blood-derived iPS cells, CBiPS cells for short, passed all standard tests for pluripotency: The gave rise to stem cell tumours known as teratomas and differentiated into derivatives of the three embryonic tissue layers, including rhythmically beating cardiomyocytes and dopamine-producing neurons.

Izpisúa Belmonte's next goal is to convince cord blood cells to burn back time using methods that are considered safe for clinical applications in humans. The original protocols for producing iPS cells — including the one used by Belmonte and his team — rely on the integration of foreign "reprogramming" genes into the host-cell genome, a process associated with risks including mutation and the development of cancers after iPS-cell transplantation, limiting their therapeutic value.

However, researchers are hard at work to develop alternative methods that allow them to reprogram cells without leaving any genetic traces, such as simply exposing differentiated cells to small molecules.

"Several studies have already shown that this could be possible," says Izpisúa Belmonte.

"If we can show they also work for cord blood cells, this certainly could be a step forward towards the clinical application of iPS cells. We should focus our efforts on this particular cell source, CBiPS cells, at least in the near future."
.........

ZenMaster

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Human Cord Blood Cells Reprogrammed into Embryonic-like Stem Cells

2009-10-12T04:08:00.001+02:00

Human Cord Blood Cells Reprogrammed into Embryonic-like Stem Cells
Monday, 12 October 2009

Human umbilical cord blood cells may be far more versatile than previous research has indicated. Two independent studies, published by Cell Press in the October 2nd issue of the journal Cell Stem Cell, report that they have successfully reprogrammed human umbilical cord blood cells into cells with properties similar to human embryonic stem cells. The results are significant as they identify cord blood as a convenient source for generating cells with a theoretically limitless potential.

Recent research has shown that adult cells can be reprogrammed into cells with characteristics similar to embryonic stem cells by turning on a select set of genes. The cells, called induced pluripotent stem (iPS) cells, have tremendous potential for regenerative medicine. However, issues related to difficulty harvesting adult cells, inefficient reprogramming and the accumulation of genetic errors (mutations) that may contribute to an increased risk for cancer and diminished cellular functionality have presented formidable challenges. Human umbilical cord blood cells have been suggested as an attractive alternative to adult cells for reprogramming.

"Cord blood-derived cells can be collected without any risk for the donor, are young cells expected to carry minimal mutations and possess the immunological immaturity of newborn cells. We believe that cord blood cells could represent, rather than just another cell type that can be reprogrammed, a real alternative for a safer source of iPS cells," explains senior study author Dr. Izpisúa Belmonte from the Center of Regenerative Medicine in Barcelona, Spain and The Salk Institute in La Jolla, California.

Dr. Izpisúa Belmonte and colleagues described a specific process that converted human cord blood cells into embryonic-like stem cells using only two factors.

"From a mechanistic point of view, the fact that cord blood-derived iPS cells could be generated by activating only two genes is a crucial point that offers new possibilities for investigating the molecular basis of the reprogramming process," concludes Dr. Izpisúa Belmonte.

In a separate study, led by Dr. Ulrich Martin from Hannover Medical School in Hannover, Germany, cord blood cells were also used to generate cells that exhibited characteristics typical of embryonic stem cells. Dr. Martin and colleagues demonstrated that the iPS cells had the potential to differentiate into multiple mature cell types, including functional heart muscle cells.

"Our study provides a feasible strategy for the reproducible generation of iPS cells from human cord blood" offers Dr. Martin.

"Importantly, public and commercial cord blood banks may provide a superior and almost unlimited source of for the production of clinically useful iPS cells."

Both research groups highlight the substantial clinical convenience of the existing networks for banking human cord blood and the theoretical advantage of these "young" cells in that they may have a decreased risk of having accumulated damaging genetic mutations associated with adult cells. The successful reprogramming of human cord blood cells into pluripotent stem cells is an important step towards future regenerative therapies. "Our findings should facilitate the clinical translation of iPS cell-based therapies," says Dr. Izpisúa Belmonte.
.........

ZenMaster

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Stem Cell Success Points to Way to Regenerate Parathyroid Glands

2009-10-12T04:01:00.002+02:00

Embryonic stem cells provide model; goal is to prevent bone loss
Monday, 12 October 2009

An early laboratory success is taking University of Michigan Health System researchers a step closer to parathyroid gland transplants that could one day prevent a currently untreatable form of bone loss associated with thyroid surgery.

The scientists were able to induce embryonic stem cells to differentiate into parathyroid cells that produced a hormone essential to maintaining bone density. The laboratory results in live cell cultures, published in Stem Cells and Development, need to be tested in further pre-clinical studies.

Parathyroid glands, four glands each the size of a rice grain that lie next to the thyroid in the neck, are easily damaged when surgeons operate on patients with cancerous or benign thyroid tumours. Without their calcium-regulating hormone, patients can develop osteomalacia, a severe form of bone loss similar to rickets that affects tens of thousands of people in the United States with muscle cramps and numbness in the hands and feet.

"We used human embryonic stem cells as a model for ways to work out the recipe to make parathyroid cells," says Gerard M. Doherty, M.D., chief of endocrine surgery and Norman W. Thompson Professor of Endocrine Surgery at U-M Medical School.

The research illustrates the payoff of rapidly increasing knowledge about how embryonic stem cells give rise to other kinds of cells. That knowledge can be the springboard for influencing other cells to regenerate damaged parts of the body.

Doherty's team used embryonic stem cells from a Bush administration-approved embryonic stem cell line to test a way to produce functioning, differentiated parathyroid cells to transplant into a patient and restore function.

With the recipe worked out, Doherty's team anticipates developing a treatment that does not use embryonic stem cells.

"We anticipate taking a person's own cells and making them into parathyroid cells," Doherty says. Using the patient's own cells should eliminate the risk of rejection.

What's next
Having demonstrated a method for leading embryonic stem cells to produce parathyroid cells, the team hopes to be able to repeat those steps using cells from the patient's own thymus gland. The method involves no genetic modification of cells, a key goal of Doherty's team.

"We want to have a process that will allow us to reintroduce cells into the patient's body safely," Doherty says.

Any successful treatment in people is five to 10 years away.
.........

ZenMaster

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Memories of the Way They Used to Be

2009-09-18T23:32:00.004+02:00

Human iPS Cells Retain Some Gene Expression of Donor Cells
Friday, 18 September 2009

A team of researchers from the University of California, San Diego School of Medicine and the Salk Institute for Biological Studies in La Jolla have developed a safe strategy for reprogramming cells to a pluripotent state without use of viral vectors or genomic insertions. Their studies reveal that these induced pluripotent stem cells (iPSCs) are very similar to human embryonic stem cells, yet maintain a “transcriptional signature.” In essence, these cells retain some memory of the donor cells they once were.

The study, led by UCSD Stem Cell Program researcher Alysson R. Muotri, PhD, assistant professor in the Departments of Pediatrics at UCSD and Rady Children’s Hospital and UCSD’s Department of Cellular and Molecular Medicine, will be published online in PLoS ONE on September 17.

“Working with neural stem cells, we discovered that a single factor can be used to re-program a human cell into a pluripotent state, one with the ability to differentiate into any type of cell in the body” said Muotri. Traditionally, a combination of four factors was used to create iPSCs, in a technology using viral vectors – viruses with the potential to affect the transcriptional profile of cells, sometimes inducing cell death or tumours.

In addition, while both mouse and human iPSCs have been shown to be similar to embryonic stem cells in terms of cell behaviour, gene expression and their potential to differentiate into different types of cells, researchers had not achieved a comprehensive analysis to compare iPSCs and embryonic stem cells.

“One reason is that previous methodologies used to derive iPSCs weren’t ‘footprint free,’” Muotri explained.

“Viruses could integrate into the genome of the cell, possibly affecting or disrupting genes.”

"In order to take full advantage of reprogramming, it is essential to develop methods to induce pluripotency in the absence of permanent changes in the genome," added Fred H. Gage, PhD, a professor in the Laboratory for Genetics at the Salk Institute and the Vi and John Adler Chair for Research on Age-Related Neurodegenerative Diseases.

By creating iPSCs from human neural stem cells without the use of viruses, the scientists learned something new. While the genetic transcriptional profile of the new iPSCs was closer to that of embryonic stem cells than to human neural stem cells, the iPSCs still carried a transcriptional “signature” of the original neural cell.

“While most of the original genetic memory was erased when the cells were reprogrammed, some were retained,” said Muotri.

He added that, in the past, it wasn’t known if this was caused by the use of viral vectors.

“By using a footprint-free methodology, we have shown a safe way to generate human iPSCs for clinical purposes and basic research. We’ve also raised an interesting question about what, if any, effect the ‘memory retention’ of these cells might have.”

The research was supported by start-up funds from the UCSD Stem Cell Research Program, and by grants from the California Institute of Regenerative Medicine and The Lookout Fund Foundation.

Reference:
Transcriptional Signature and Memory Retention of Human-Induced Pluripotent Stem Cells
Maria C. N. Marchetto, Gene W. Yeo, Osamu Kainohana, Martin Marsala, Fred H. Gage, Alysson R. Muotri
PLoS ONE 4(9): e7076. doi:10.1371/journal.pone.0007076
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Rare Genetic Disease Successfully Reversed using Stem Cell Transplantation

2009-09-17T21:22:00.002+02:00

Scripps Research scientists correct gene defect in mice that causes lethal symptoms in children
Thursday, 17 September 2009

A recent study by Scripps Research Institute scientists offers good news for families of children afflicted with the rare genetic disorder, cystinosis. In research that holds out hope for one day developing a potential therapy to treat the fatal disorder, the study shows that the genetic defect in mice can be corrected with stem cell transplantation.

"After meeting the children who suffer from this disease, like an 18-year-old who has already had three kidney transplants, and the families who are desperately searching for help, our team is committed to moving toward a cure for cystinosis, a lysosomal storage disorder," says principal investigator Stephanie Cherqui, assistant professor in the Department of Molecular and Experimental Medicine.

"This study is an important step toward that goal."

In the study, which is published in the September 17, 2009 print edition of the journal Blood, the Scripps Research team used bone marrow stem cell transplantation to address symptoms of cystinosis in a mouse model. The procedure virtually halted the cystine accumulation responsible for the disease and the cascade of cell death that follows.

Cystine is a by-product of the break down of cellular components the body no longer needs in the cell's "housekeeping" organelles, called lysosomes. Normally, cystine is shunted out of cells, but in cystinosis a gene defect of the lysosomal cystine transporter causes it to build up, forming crystals that are especially damaging to the kidneys and eyes.

A Rare But Devastating Disease
While cystinosis is rare — affecting an estimated 500 people in the United States and 2,000 worldwide — it is devastating. Three types of cystinosis have been described based on the age at diagnosis and the amount of cystine in cells: infantile onset, adolescent onset, and adult onset. Children as young as six months can begin to suffer renal dysfunction, which grows progressively worse with time. Other symptoms include diabetes, muscular disease, neurological dysfunction, and retinopathy. Infantile onset is the most common, as well as the most severe, form of the disease.

The only available drug to treat cystinosis, cysteamine, while slowing the progression of kidney degradation, does not prevent it, and end-stage kidney failure is inevitable.

"Cysteamine must be given every six hours, so children have to be woken up each night to take this drug, which has unpleasant side effects, and many others to treat various symptoms," Cherqui says.

"So although there is treatment, it is difficult treatment that does not cure the disease."

"Surprised and Encouraged"
In the new study, the researchers found that transplanted bone marrow stem cells carrying the normal lysosomal cystine transporter gene abundantly engrafted into every tissue of the experimental mice. This led to an average drop in cystine levels of about 80 percent in every organ. In addition to preventing kidney dysfunction, there was less deposition of cystine crystals in the cornea, less bone demineralization, and an improvement in motor function.

"The results really surprised and encouraged us," says Cherqui, who as a doctoral student in France in 1998 helped discover the gene involved in cystinosis.

"Because the defect is present in every cell of the body, we did not expect a bone marrow stem cell transplant to be so widespread and effective."

Cherqui, who generated the mouse model in 2000 that is currently used to study cystinosis, says that adult bone marrow stem cell therapy is particularly well suited as a potential treatment for cystinosis because these cells target all types of tissues. In addition, stem cells reside in the bone marrow for the duration of a patient's life, becoming active as needed, a particular benefit for a progressive disease like cystinosis.

The work of Cherqui and her colleagues may have wider applications for other genetic diseases, providing proof of principle that adult stem cell transplants may be successful in humans for genetic diseases with systemic defects, especially those of a progressive nature.

Cherqui expects to spend the next several years analyzing the safety of genetically modified autologous (obtained from the same individual) bone marrow transplants in the cystinosis mouse and other models before moving on to human clinical trials.

This work was funded by the Cystinosis Research Foundation.

Reference:
Successful treatment of the murine model of cystinosis using bone marrow cell transplantation
Kimberly Syres, Frank Harrison, Matthew Tadlock, James V. Jester, Jennifer Simpson, Subhojit Roy, Daniel R. Salomon, and Stephanie Cherqui
Blood 2009 114: 2542-2552, DOI 10.1182/blood-2009-03-213934
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How microRNAs Drive Tumour Progression

2009-09-17T21:02:00.001+02:00

How microRNAs Drive Tumour Progression
Thursday, 17 September 2009

UCSF researchers have identified collections of tiny molecules known as microRNAs that affect distinct processes critical for the progression of cancer. The findings, they say, expand researchers' understanding of the important regulatory function of microRNAs in tumour biology and point to new directions for future study and potential treatments.

The researchers refer to these microRNA collections as signatures, and their study results are reported in the September 15 issue of "Genes & Development.'' The study was led by the laboratory of Douglas Hanahan, PhD, an American Cancer Society Research Professor in the Department of Biochemistry and Biophysics at UCSF.

Approximately five percent of all known human genes encode, or produce, microRNAs, yet scientists are only now — nearly a decade after their discovery — beginning to unlock the mystery of their functions.

MicroRNAs are snippets of single-stranded RNAs that prevent a gene's code from being translated from messenger RNA into proteins, which are essential for cell growth and development. Produced in the nucleus and released into the cytoplasm, they home in on messenger RNAs that possess a stretch that is complementary to their genetic sequence. When they locate them, they latch on, preventing the messenger RNA from being processed by the protein-making machines known as ribosomes. As such, microRNAs are able to ratchet down a cell's production of a given protein.

Over the last several years, several groups have identified hundreds of microRNAs that are deregulated between normal tissue and tumours, however researchers only understand what a handful of these powerful regulators are doing to drive tumour formation.

"Virtually all cancers acquire approximately six distinct capabilities en route to tumour formation," said lead author Peter Olson, PhD, a postdoctoral fellow in the Diabetes Center and Helen Diller Family Comprehensive Cancer Center at UCSF.

"When a cancer researcher observes a gene or microRNA go awry, it can be challenging to understand how that microRNA impacts tumourigenesis."

To home in on the question, the authors turned to a mouse model of pancreatic neuroendocrine tumours in which lesions go through discrete stages before culminating in invasive and metastatic carcinomas. In the three-year microRNA study, they found that cells in the mouse model developed and functioned normally but started to replicate uncontrollably at five weeks. Several weeks later, some pancreatic islets had become angiogenic (forming new blood vessels) — a step in the journey from a dormant state to a malignant state — though had not yet formed a tumour. By 10 weeks, a subset of angiogenic lesions had progressed to the tumour stage, and by week 16, a small percentage of mice had developed liver metastasis.

"This represents the spectrum of stages that we think are important for all tumours, including human disease," said Olson.

By measuring the expression level of all known microRNA in pre-tumour stages, tumours and metastases, the authors were able to associate deregulated microRNAs with processes such as hyper-proliferation, angiogenesis and metastasis.

Focusing on the metastatic signature, researchers found — in one of the most striking observations of the project — that tumours bore a startlingly divergent microRNA expression pattern compared to primary tumours. Moreover, a subset of primary tumours showed more similarity to metastases than to other primary tumours.

"If you can identify tumours that have an increased propensity to metastasize, then it would have a very important clinical application," said Olson.

"A lively debate in metastatic research has centred around whether primary tumour cells must suffer an additional mutation that endows that cell with a metastatic capability, or whether certain mutational combinations that are responsible for primary tumour formation also significantly increase the propensity of that cell to metastasize. These data provide evidence for the latter.''

Reference:
MicroRNA dynamics in the stages of tumorigenesis correlate with hallmark capabilities of cancer
Peter Olson, Jun Lu, Hao Zhang, Anny Shai, Matthew G. Chun, Yucheng Wang, Steven K. Libutti, Eric K. Nakakura, Todd R. Golub, and Douglas Hanahan
Genes Dev. September 15, 2009 23: 2152-2165; doi:10.1101/gad.1820109
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Cure for Colour Blindness in Monkeys

2009-09-16T20:49:00.003+02:00

Gene therapy used to treat adult vision disorders involving cone cells
Wednesday, 16 September 2009

Researchers from the University of Washington and the University of Florida used gene therapy to cure two squirrel monkeys of colour blindness — the most common genetic disorder in people.

Writing online Wednesday in the journal Nature, scientists cast a rosy light on the potential for gene therapy to treat adult vision disorders involving cone cells — the most important cells for vision in people.

"We've added red sensitivity to cone cells in animals that are born with a condition that is exactly like human colour blindness," said William W. Hauswirth, Ph.D., a professor of ophthalmic molecular genetics at the UF College of Medicine and a member of the UF Genetics Institute and the Powell Gene Therapy Center.

"Although colour blindness is only moderately life-altering, we've shown we can cure a cone disease in a primate, and that it can be done very safely. That's extremely encouraging for the development of therapies for human cone diseases that really are blinding."

The finding is also likely to intrigue millions of people around the world who are colour-blind, including about 3.5 million people in the United States, more than 13 million in India and more than 16 million in China. The problem mostly affects men, leaving about 8 percent of Caucasian men in the United States incapable of discerning red and green hues that are important for everyday things like recognizing traffic lights.

"People who are colour-blind feel that they are missing out," said Jay Neitz, Ph.D., a professor of ophthalmology at the University of Washington.

"If we could find a way to do this with complete safety in human eyes, as we did with monkeys, I think there would be a lot of people who would want it. Beyond that, we hope this technology will be useful in correcting lots of different vision disorders."

The discovery comes about 10 years after Neitz and his wife Maureen Neitz, Ph.D., a professor of ophthalmology at the University of Washington and senior author of the study, began training two squirrel monkeys named Dalton and Sam.

In addition to teaching the animals, the Neitz research group worked with the makers of a standard vision-testing technique called the Cambridge Colour Test to perfect a way the monkeys could "tell" them which colours they were seeing.

The tests are similar to ones given to elementary children the world over, in which students are asked to identify a specific pattern of coloured dots among a field of dots that vary in size, colour and intensity. The researchers devised a computer touch screen the monkeys could use to trace the colour patterns. When the animals chose correctly, they received a reward of grape juice.

Likewise, decades were spent by Hauswirth and colleagues at the University of Florida, to develop the gene-transfer technique that uses a harmless adeno-associated virus to deliver corrective genes to produce a desired protein.

In this case, researchers wanted to produce a substance called long-wavelength opsin in the retinas of the monkeys. This particular form of opsin is a colourless protein that works in the retina to make pigments that are sensitive to red and green.

"We used human DNA’s, so we won't have to switch to human genes as we move toward clinical treatments," said Hauswirth, who is also involved in a clinical trial with human patients to test gene therapy for the treatment of Leber congenital amaurosis, a form of blindness that strikes children.

About five weeks after the treatment, the monkeys began to acquire colour vision, almost as if it occurred overnight.

"Nothing happened for the first 20 weeks," Neitz said.

"But we knew right away when it began to work. It was if they woke up and saw these new colours. The treated animals unquestionably responded to colours that had been invisible to them."

It took more than a year and a half to test the monkeys' ability to discern 16 hues, with some of the hues varying as much as 11-fold in intensity.

Dalton is named for John Dalton, an English chemist who realized he was colour-blind and published the first paper about the condition in 1798.

"We've had Dalton and Sam for 10 years. They are like our children," Neitz said.

"This species are friendly, docile monkeys that we just love. We think it is useful to continue to follow them — it's been two years now that they've been seeing in colour, and continuing to check their vision and allowing them to play with the computer is part of their enrichment."

With the discovery, the researchers are the first to address a vision disorder in primates in which all photoreceptors is intact and healthy, providing a hint of gene therapy's full potential to restore vision.

About 1 in 30,000 Americans have a hereditary form of blindness called achromatopsia, which causes nearly complete colour blindness and extremely poor central vision.

"Those patients would be targets for almost exactly the same treatment," Hauswirth said.

Even in common types of blindness such as age-related macular degeneration and diabetic retinopathy, vision could potentially be rescued by targeting cone cells, he said.

"The major thrust of the study is you can ameliorate if not cure colour blindness with gene therapy," said Gerald H. Jacobs, Ph.D., a research professor of psychology at the University of California, Santa Barbara, who was not involved in the research.

"There are still questions about safety, but in these monkeys at least, there were no untoward effects. Those who are motivated to ameliorate their colour defect might take some hope from the findings.”

"This is also another example of how utterly plastic the visual system is to change," Jacobs said.

"The nervous system can extract information from alterations to photo-pigments and make use of it almost instantaneously."

Reference:
Gene therapy for red–green colour blindness in adult primates
Katherine Mancuso, William W. Hauswirth, Qiuhong Li, Thomas B. Connor, James A. Kuchenbecker, Matthew C. Mauck, Jay Neitz & Maureen Neitz
Nature advance online publication 16 September 2009, doi:10.1038/nature08401
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Benefits and Limits of Personal Genetics

2009-09-15T20:35:00.001+02:00

Dartmouth researchers get personal with genetics
Tuesday, 15 September 2009

Two recent studies by Dartmouth College researchers use individual genetic data to reveal the powers and limits of our current understanding of how the genome influences human health and what genes can reveal about the ancestry of the people of New Hampshire.

Published in the Sept. 11 issue of the American Journal of Human Genetics, Dartmouth Professor Jason Moore and Vanderbilt Professor Scott Williams analyzed how personal genetic testing companies are using still-nascent genome data to judge the health of their customers.

People can now buy inexpensive kits, submit a DNA sample (often a swab from the inside of a cheek or a little bit of saliva), and receive data about their susceptibility to a number of gene-influenced ailments, such as prostate cancer, Alzheimer's, or type II diabetes. Moore and Williams argue that our knowledge of the human genome and its relationship to human health, while growing by leaps and bounds, is still in its infancy.

"The relationship between health and genetics is very complex," says Moore, professor of genetics and of community and family medicine at Dartmouth Medical School (DMS).

"It's often a combination of multiple genes and multiple environmental factors that work together to increase or decrease your risk of disease. I don't think the knowledge base is sufficient to put genetics in the hands of the public quite yet." Moore is also the Frank Lane Research Scholar in Computational Genetics and Director of Bioinformatics at DMS

The authors admit that genetic research is progressing, and they cite the example of the discovery of the BRCA1 and BRCA2 genes and their role in breast cancer. However, the authors caution that, while there is no question these genes are involved in breast cancer, the underlying mechanisms behind the genetic risk are still being worked out.

"There is a perception that these tests can provide answers," says Moore.

"I used my own genetic material for this study, and my results didn't really tell me anything I didn't know, based on family history."

Moore and Williams call for refocusing and stepping up the research on gene-to-gene and gene-to-environment interactions. They explain that for many years, researchers have focused on single genes and clinical endpoints. The time has come, they say, to embrace rather than ignore the complexity of human traits as they are expressed by the whole genome working in concert.

"Although genetic testing for common human diseases is not yet useful, using genetic testing results to reveal an individual's ancestry is increasingly reliable," says Moore. He and PhD candidate Chantel Sloan recently mined some genetic data for a study that examined the population structure of New Hampshire residents.

Published in the September issue of PLoS ONE (a journal of the Public Library of Science), they study by Sloan and Moore and their colleagues analyzed more than 1,000 genetic markers from 864 people in New Hampshire. They discovered six subgroups of people with distinct genetic backgrounds including a group of Finnish and Russian/Polish/Lithuanian ancestry.

"I knew that people would be primarily European," says Sloan.

"What I didn't expect was the strong connection between genetic structure and people of Eastern European ancestry, which I learned is consistent with New Hampshire census and immigration data from 1870 to 1930."

Sloan used data initially compiled for a cancer study, so the genetic markers were cancer susceptibility genes rather than known ancestral genes, and the population being analyzed was not racially or geographically distinct. The results challenge the assumption that large numbers of special genetic markers are needed to discover genetically distinct groups of people.

"This is an example of how personal genetic data can be used to help inform people of their ancestry," says Moore.

"Informing people of their future health is still out of reach, though."

References:
Epistasis and Its Implications for Personal Genetics
Jason H. Moore, Scott M. Williams
AJHG, Volume 85, Issue 3, 309-320, 11 September 2009, doi:10.1016/j.ajhg.2009.08.006

Genetic Population Structure Analysis in New Hampshire Reveals Eastern European Ancestry
Chantel D. Sloan, Angeline D. Andrew, Eric J. Duell, Scott M. Williams, Margaret R. Karagas, Jason H. Moore
PLoS ONE 4(9): e6928. doi:10.1371/journal.pone.0006928
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How Stem Cells Make Skin

2009-09-13T22:17:00.004+02:00

EMBL scientists come a step closer to understanding skin, breast and other cancers.
Sunday, 13 September 2009

Stem cells have a unique ability: when they divide, they can either give rise to more stem cells, or to a variety of specialised cell types. In both mice and humans, a layer of cells at the base of the skin contains stem cells that can develop into the specialised cells in the layers above. Scientists at the European Molecular Biology Laboratory (EMBL) in Monterotondo, in collaboration with colleagues at the Centro de Investigaciones Energéticas, Medioambientales y Tecnologicas (CIEMAT) in Madrid, have discovered two proteins that control when and how these stem cells switch to being skin cells. The findings, published online today in Nature Cell Biology, shed light on the basic mechanisms involved not only in formation of skin, but also on skin cancer and other epithelial cancers.

At some point in their lives, the stem cells at the base of the skin stop proliferating and start differentiating into the cells that form the skin itself. To do so, they must turn off the 'stem cell programme' in their genes and turn on the 'skin cell programme'. Researchers suspected that a family of proteins called C/EBPs might be involved in this process, as they were known to regulate it in other types of stem cell, but had so far failed to identify which C/EBP protein controlled the switch in skin. Claus Nerlov and his group at EMBL Monterotondo discovered it was not one protein, but two: C/EBPα and C/EBPβ.

In normal skin (left), the stem cells at the base, shown in green, differentiate into skin cells, shown in red. In mice whose skin has neither C/EBP-α nor C/EBP-β (middle), this differentiation is blocked: green-labelled stem cells appear in upper layers of skin, and there are no differentiated skin cells (no red staining). This also happens at the initial stages of basal cell carcinomas. In skin where C/EBP-β is present but has lost its capacity to interact with E2F, a molecule that regulates the cell cycle (right), skin cells start differentiating abnormally, before they have properly exited the stem cell "program" (yellow/orange). This is similar to what is observed in the initial stages of squamous cell carcinomas, a more aggressive and invasive skin tumour. Credit: Claus Nerlov/EMBL.

The EMBL researchers used genetic engineering techniques to delete the genes that encode C/EBPα and β specifically in the skin of mouse embryos, and found that without these proteins the skin of the mice did not form properly.

"Mice with neither C/EBPα nor β had taut and shiny skin that couldn't keep the water inside their bodies, they lacked many of the proteins that make skin mechanically strong and water tight, and they died of de-hydration shortly after birth," Nerlov explains.

However, a single working copy of either the gene for C/EBPα or the gene for C/EBPβ was enough to ensure that skin developed properly. This means that the two proteins normally do the same job in the skin's stem cells - an unexpected redundancy, which may have arisen because there are so many stem cells in skin that a tight control on proliferation is needed to avoid problems like cancer. Or it may simply be a by-product of the fact that these two proteins have different functions in other situations, such as wound healing or repair of sunlight-induced skin damage.

One of the hallmarks of epithelial cancers - which include skin, breast, and oral cancers - is that they have genes turned on which would normally only be expressed in embryonic stem cells, and which may help cancer cells divide indefinitely. Such genes become re-expressed in the skin in the absence of C/EBPs. Therefore, by understanding how C/EBPα and β turn off such 'stem cell' programmes, researchers hope to come a step closer to finding ways to fight such cancers.

When Nerlov and colleagues looked at how C/EBPα and -β work in the skin, they found that these proteins also regulate a number of other molecules that control skin development. Several important pathways known to control skin and hair formation were improperly activated in the mice lacking C/EBPα and -β.

"This is a very important discovery", says Nerlov.

"It opens up a lot of new areas, because we can see how these proteins control virtually every other molecule known to regulate skin cell differentiation. It seems to be a key piece in the puzzle of how our skin is formed and maintained throughout life."

Reference:
C/EBPα and β couple interfollicular keratinocyte proliferation arrest to commitment and terminal differentiation
Rodolphe G. Lopez, Susana Garcia-Silva, Susan J. Moore, Oksana Bereshchenko, Ana B. Martinez-Cruz, Olga Ermakova, Elke Kurz, Jesus M. Paramio & Claus Nerlov
Nature Cell Biology Published online: 13 September 2009, doi:10.1038/ncb1960
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MicroRNAs Are Themselves Regulated

2009-09-12T16:05:00.004+02:00

RNAs taking centre stage
Saturday, 12 September 2009

In an article published in Nature, scientists at the Friedrich Miescher Institute for Biomedical Research (part of the Novartis Research Foundation) have shown that short strands of ribonucleic acid (RNA) which regulate protein production are themselves also regulated. This additional layer of regulation opens up new perspectives for therapeutic approaches.

RNAs, serving as a mere intermediary between DNA and proteins, were long regarded as a poor relation by researchers, attracting little interest. However, following the discovery of small RNAs known as microRNAs, they have increasingly been moving into the limelight. MicroRNAs bind to messenger RNA (mRNA), thereby regulating the translation of genes into proteins.

Recently, various studies have shown that the production of microRNAs is tightly controlled, but their subsequent fate was not clear. It was assumed that mature microRNAs remained stable in the cell for days, and that their possible functions were therefore restricted: a microRNA persisting for a relatively long period cannot be involved in any processes in the cell requiring rapid adaptation.

Regulated regulators
The study carried out by Helge Grosshans, a Research Group Leader at the Friedrich Miescher Institute, has now finally shifted attention away from DNA, spotlighting the key role played by microRNAs in the theatre of cellular processes. As Grosshans and his team report in the current issue of the renowned journal Nature, they discovered a mechanism for active degradation of microRNAs and showed that this mechanism is itself regulated.
Explaining his findings, Grosshans says:

"What was formerly conceived of as a direct, straightforward pathway is gradually turning out to be a dense network of regulatory mechanisms: genes are not simply translated into proteins via mRNA. MicroRNAs control the translation of mRNAs into proteins, and proteins in turn regulate the microRNAs at various levels."

In addition, the FMI researchers showed in the nematode Caenorhabditis elegans that, via regulation of degradation, it is possible to influence microRNA activity. This means that microRNAs may, after all, be involved in the regulation of rapidly occurring processes.

The meteoric rise of microRNAs
MicroRNAs are short, single-stranded RNA molecules which interact with mRNAs in a sequence-dependent manner. They thus inhibit translation of mRNAs into proteins. MicroRNAs were first described in 1993 in the nematode Caenorhabditis elegans. They were subsequently also shown to play an important role in regulating development processes and in pathogenesis in higher organisms. The findings of recent years and now also Helge Grosshans's study have shifted attention away from DNA toward RNAs, which are taking centre stage. The term "microRNA" was only introduced in 2001.

Targeted degradation of disease-causing RNAs
But the findings are also relevant in another respect. As microRNAs have been implicated in the development of diseases, efforts to date have focused on replacing disease-causing microRNAs with other microRNAs, or inactivating them with the aid of complementary RNA strands. Unfortunately, it is extremely difficult to deliver RNAs to target cells for therapeutic purposes. Accordingly, the prospects of success for these novel treatment approaches have been uncertain.

In his study, however, Grosshans identified a protein that specifically degrades microRNAs. If it now proves possible to specifically activate or inhibit this protein and its partners that could provide an approach, which is closer to classical and well-established forms of therapy.

"We now assume that a large number of human genes are regulated by microRNAs, so the regulatory mechanism we've discovered has a great potential to significantly influence numerous processes in human cells," Grosshans comments.

About the FMI
The Friedrich Miescher Institute for Biomedical Research (FMI), based in Basel, Switzerland, is a world-class centre for basic research in life sciences. It was founded in 1970 as a joint effort of two Basel-based pharmaceutical companies and is now part of the Novartis Research Foundation. The FMI is devoted to the pursuit of fundamental biomedical research. Areas of expertise are neurobiology, growth control, which includes signalling pathways, and the epigenetics of stem cell development and cell differentiation. The institute counts 320 collaborators. The FMI also offers training in biomedical research to PhD students and postdoctoral fellows from around the world. In addition, the FMI is affiliated with the University of Basel. The Director of the FMI since 2004 is Prof. Susan Gasser.

Reference:
Active turnover modulates mature microRNA activity in C. elegans
Chatterjee S & H. Grosshans
Nature, 24 August 2009, doi:10.1038/nature08349
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See CIRMTV's Channel on YouTube

2009-09-11T14:05:00.002+02:00

California's stem cell research funding agency is using YouTube to tell its story.
Friday, 11 September 2009

The California Institute for Regenerative Medicine's CIRMTV tells the story of stem cell research through short videos about the work of researchers and doctors and — most compellingly — patients.

The 22 videos on the site range from Stanford University stem cell research director Irv Weissman talking about the differences between adult and embryonic stem cells to, most recently, an update about the progress and promise in Parkinson's disease treatments.

See CIRMTV's Channel:
YouTube - CIRMTV's Channel
.........

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'Liposuction Leftovers' Easily Converted to iPS Cells

2009-09-08T07:56:00.001+02:00

'Liposuction Leftovers' Easily Converted to iPS Cells
Tuesday, 08 September 2009

Globs of human fat removed during liposuction conceal versatile cells that are more quickly and easily coaxed to become induced pluripotent stem cells, or iPS cells, than are the skin cells most often used by researchers, according to a new study from Stanford's School of Medicine.

"We've identified a great natural resource," said Stanford surgery professor and co-author of the research, Michael Longaker, MD, who has called the readily available liposuction leftovers "liquid gold." Reprogramming adult cells to function like embryonic stem cells is one way researchers hope to create patient-specific cell lines to regenerate tissue or to study specific diseases in the laboratory.

"Thirty to 40 percent of adults in this country are obese," agreed cardiologist Joseph Wu, MD, PhD, the paper's senior author.

"Not only can we start with a lot of cells, we can reprogram them much more efficiently. Fibroblasts, or skin cells, must be grown in the lab for three weeks or more before they can be reprogrammed. But these stem cells from fat are ready to go right away."

The fact that the cells can also be converted without the need for mouse-derived "feeder cells" may make them an ideal starting material for human therapies. Feeder cells are often used when growing human skin cells outside the body, but physicians worry that cross-species contamination could make them unsuitable for human use.

The findings will be published online Sept. 7 in the Proceedings of the National Academy of Sciences. Longaker is the deputy director of Stanford’s Stem Cell Biology and Regenerative Medicine Institute and director of children’s surgical research at Lucile Packard Children’s Hospital. Wu is an assistant professor of cardiology and radiology, and a member of Stanford’s Cardiovascular Institute.

Even those of us who are not obese would probably be happy to part with a couple of pounds (or more) of flab. Nestled within this unwanted latticework of fat cells and collagen are multipotent cells called adipose, or fat, stem cells. Unlike highly specialized skin-cell fibroblasts, these cells in the fat have a relatively wide portfolio of differentiation options — becoming fat, bone or muscle as needed. It's this pre-existing flexibility, the researchers believe, that gives these cell an edge over the skin cells.

"These cells are not as far along on the differentiation pathway, so they're easier to back up to an earlier state," said first author and postdoctoral scholar Ning Sun, PhD, who conducted the research in both Longaker's and Wu's laboratories.

"They are more embryonic-like than fibroblasts, which take more effort to reprogram."

These reprogrammed iPS cells are usually created by expressing four genes, called Yamanaka factors, normally unexpressed (or expressed at very low levels) in adult cells.

Sun found that the fat stem cells actually express higher starting levels of two of the four reprogramming genes than do adult skin cells — suggesting that these cells are already primed for change. When he added all four genes, about 0.01 percent of the skin-cell fibroblasts eventually became iPS cells but about 0.2 percent of the fat stem cells did so — a 20-fold improvement in efficiency.

The new iPS cells passed the standard tests for pluripotency: They formed tumours called teratomas when injected into immunocompromised mice, and they could differentiate into cells from the three main tissue types in the body, including neurons, muscle and gut epithelium. The researchers are now investigating whether the gene expression profiles of the fat stem cells could be used to identify a subpopulation that could be reprogrammed even more efficiently.

"The idea of reprogramming a cell from your body to become anything your body needs is very exciting," said Longaker, who emphasized that the work involved not just a collaboration between his lab and Wu's, but also between the two Stanford institutes.

"The field now needs to move forward in ways that the Food and Drug Administration would approve — with cells that can be efficiently reprogrammed without the risk of cross-species contamination — and Stanford is an ideal place for that to happen."

"Imagine if we could isolate fat cells from a patient with some type of congenital cardiac disease," said Wu.

"We could then differentiate them into cardiac cells, study how they respond to different drugs or stimuli and see how they compare to normal cells. This would be a great advance."
.........

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Transplanted Human Stem Cells Prolong Survival in Mouse Model of Batten Disease

2009-09-04T03:04:00.001+02:00

Transplanted Human Stem Cells Prolong Survival in Mouse Model of Batten Disease
Friday, 04 September 2009

A new study finds substantial improvement in a mouse model of a rare, hereditary neurodegenerative disease after transplantation of normal human neural stem cells. The research findings, published by Cell Press in the September 4th issue of the journal Cell Stem Cell, show that the transplanted cells provided a critical enzyme that was missing in the brains of the experimental mice and represent an important step toward what may be a successful therapeutic approach for a currently untreatable and devastating disease.

Infantile neuronal ceroid lipofuscinosis (INCL), commonly known as Batten disease, is a fatal neurodegenerative disease in children. It is caused by a mutation in the gene that makes a crucial enzyme called palmitoyl protein thioesterase-1 (PPT1). A deficiency of PPT1 in the brain causes the abnormal accumulation of a cellular lipid storage material called lipofuscin, which leads to neuron death, a decline in cognitive and motor skills, visual impairment, seizures and premature death. Unfortunately, intravenous enzyme replacement therapy is not a viable treatment approach as it is nearly impossible to get the PPT1 enzyme into the brain.

Although there is currently no effective treatment for INCL, it has been hypothesized that transplanted donor cells might be able to secrete the needed enzyme directly into the host brain. A mouse model of INCL that mimics many aspects of the human disease has been developed and provides an excellent experimental model for testing whether a human neural stem cell transplant may be a beneficial disease treatment. Dr. Nobuko Uchida from StemCells, Inc., in Palo Alto, California led a study that tested this hypothesis with banked human neural stem cells that had been purified, expanded, and preserved.

"We took a novel approach and transplanted normal, non-tumourigenic, and non-genetically modified human neural stem cells to deliver the deficient enzyme in the mouse model of INCL," explains Dr. Uchida.

"We transplanted self-renewing human neural stem cells because, theoretically, these transplants can provide life-long production of the missing enzyme."

Dr. Uchida and colleagues found that the purified human neural stem cells engrafted to the brain of INCL mice, migrated extensively, and produced enough PPT1 in the host mice to elicit significant improvement. Specifically, the INCL mice exhibited reduced lipofuscin, widespread neuroprotection, and a delayed loss of motor coordination.

"Early intervention with neural stem cell transplants into the brains of INCL patients may supply a continuous and long-lasting source of the missing PPT1 and provide some therapeutic benefit through protection of endogenous neurons," concludes Dr. Uchida.

"These data support our rationale for continued development in humans and the potential for a medical breakthrough in this deadly disease."

Notably, StemCells, Inc., recently reported positive results from the first Phase 1 clinical trials assessing the safety of these human neural stem cells as a potential treatment for Batten disease.

Reference:
Neuroprotection of Host Cells by Human Central Nervous System Stem Cells in a Mouse Model of Infantile Neuronal Ceroid Lipofuscinosis
Stanley J. Tamaki, Yakop Jacobs, Monika Dohse, Alexandra Capela, Jonathan D. Cooper, Michael Reitsma, Dongping He, Robert Tushinski, Pavel V. Belichenko, Ahmad Salehi, William Mobley, Fred H. Gage, Stephen Huhn, Ann S. Tsukamoto, Irving L. Weissman and Nobuko Uchida
Cell Stem Cell, Volume 5, Issue 3, 310-319, 4 September 2009, doi:10.1016/j.stem.2009.05.022
.........

ZenMaster

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What Makes Us Uniquely Human?

2009-09-02T04:00:00.001+02:00

Discovery of novel genes could unlock the mystery
Wednesday, 02 September 2009

Humans and chimpanzees are genetically very similar, yet it is not difficult to identify the many ways in which we are clearly distinct from chimps. In a study published online in Genome Research, scientists have made a crucial discovery of genes that have evolved in humans after branching off from other primates, opening new possibilities for understanding what makes us uniquely human.

The prevailing wisdom in the field of molecular evolution was that new genes could only evolve from duplicated or rearranged versions of pre-existing genes. It seemed highly unlikely that evolutionary processes could produce a functional protein-coding gene from what was once inactive DNA.

However, recent evidence suggests that this phenomenon does in fact occur. Researchers have found genes that arose from non-coding DNA in flies, yeast, and primates. No such genes had been found to be unique to humans until now, and the discovery raises fascinating questions about how these genes might make us different from other primates.

In this work, David Knowles and Aoife McLysaght of the Smurfit Institute of Genetics at Trinity College Dublin undertook the painstaking task of finding protein-coding genes in the human genome that are absent from the chimp genome. Once they had performed a rigorous search and systematically ruled out false results, their list of candidate genes was trimmed down to just three. Then came the next challenge.

"We needed to demonstrate that the DNA in human is really active as a gene," said McLysaght.

The authors gathered evidence from other studies that these three genes are actively transcribed and translated into proteins, but furthermore, they needed to show that the corresponding DNA sequences in other primates are inactive. They found that these DNA sequences in several species of apes and monkeys contained differences that would likely disable a protein-coding gene, suggesting that these genes were inactive in the ancestral primate.

The authors also note that because of the strict set of filters employed, only about 20% of human genes were amenable to analysis. Therefore, they estimate there may be approximately 18 human-specific genes that have arisen from non-coding DNA during human evolution.

This discovery of novel protein-coding genes in humans is a significant finding, but raises a bigger question: What are the proteins encoded by these genes doing?

"They are unlike any other human genes and have the potential to have a profound impact," McLysaght noted. While these genes have not been characterized yet and their functions remain unknown, McLysaght added that it is tempting to speculate that human-specific genes are important for human-specific traits.

Reference:
Recent de novo origin of human protein-coding genes
Knowles DG, McLysaght A.
Genome Res doi:10.1101/gr.095026.109
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ZenMaster

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We Are All Human Mutants

2009-08-27T23:38:00.003+02:00

Measurement of mutation rate in humans by direct sequencing
Thursday, 27 August 2009

An international team of 16 scientists today reports the first direct measurement of the general rate of genetic mutation at individual DNA letters in humans. The team sequenced the same piece of DNA - 10,000,000 or so letters or 'nucleotides' from the Y chromosome - from two men separated by 13 generations, and counted the number of differences. Among all these nucleotides, they found only four mutations.

In 1935 one of the founders of modern genetics, J. B. S. Haldane, studied men in London with the blood disease haemophilia and estimated that there would be one in 50,000 incidence of mutations causing haemophilia in the gene affected - the equivalent of a mutation rate of perhaps one in 25 million nucleotides across the genome. Others have measured rates at a few further specific genes or compared DNA from humans and chimpanzees to produce general estimates of the mutation rate expressed more directly in nucleotides of DNA.

Remarkably, the new research, published today in Current Biology, shows that these early estimates were spot on - in total, we all carry 100-200 new mutations in our DNA. This is equivalent to one mutation in each 15 to 30 million nucleotides. Fortunately, most of these are harmless and have no apparent effect on our health or appearance.

"The amount of data we generated would have been unimaginable just a few years ago," says Dr Yali Xue from the Wellcome Trust Sanger Institute and one of the project's leaders.

"But finding this tiny number of mutations was more difficult than finding an ant's egg in the emperor's rice store."

Team member Qiuju Wang recruited a family from China who had lived in the same village for centuries. The team studied two distant male-line relatives - separated by thirteen generations - whose common ancestor lived two hundred years ago.

To establish the rate of mutation, the team examined an area of the Y chromosome. The Y chromosome is unique in that, apart from rare mutations, it is passed unchanged from father to son; so mutations accumulate slowly over the generations.

Despite many generations of separation, researchers found only 12 differences among all the DNA letters examined. The two Y chromosomes were still identical at 10,149,073 of the 10,149,085 letters examined. Of the 12 differences, eight had arisen in the cell lines used for the work. Only four were true mutations that had occurred naturally through the generations.

We have known for a long time that mutations occur occasionally in each of us, but have had to guess exactly how often. Now, thanks to advances in the technology for reading DNA, this new research has been possible.

Understanding mutation rates is key to many aspects of human evolution and medical research: mutation is the ultimate source of all our genetic variation and provides a molecular clock for measuring evolutionary timescales. Mutations can also lead directly to diseases like cancer. With better measurements of mutation rates, we could improve the calibration of the evolutionary clock, or test ways to reduce mutations, for example.

Even with the latest DNA sequencing technology, the researchers had to design a special strategy to search for the vanishingly rare mutations. They used next-generation sequencing to establish the order of letters on the two Y chromosomes and then compared these to the Y chromosome reference sequence.

Having identified 23 candidate SNPs - or single letter changes in the DNA - they amplified the regions containing these candidates and checked the sequences using the standard Sanger method. A total of four naturally occurring mutations were confirmed. Knowing this number of mutations, the length of the area that they had searched and the number of generations separating the individuals, the team were able to calculate the rate of mutation.

"These four mutations gave us the exact mutation rate - one in 30 million nucleotides each generation - that we had expected," says the study's coordinator, Chris Tyler-Smith, also from The Wellcome Trust Sanger Institute.

"This was reassuring because the methods we used - harnessing next-generation sequencing technology - had not previously been tested for this kind of research. New mutations are responsible for an array of genetic diseases. The ability to reliably measure rates of DNA mutation means we can begin to ask how mutation rates vary between different regions of the genome and perhaps also between different individuals."

Participating centres were The Wellcome Trust Sanger Institute, Hinxton, Cambridgeshire, UK; Department of Otorhinolaryngology-Head and Neck Surgery, and Institute of Otolaryngology, Chinese People's Liberation Army General Hospital, Beijing, China; and Beijing Genomics Institute at Shenzhen, Shenzhen, China.

Reference:
Human Y Chromosome Base-Substitution Mutation Rate Measured by Direct Sequencing in a Deep-Rooting Pedigree
Yali Xue, Qiuju Wang, Quan Long, Bee Ling Ng, Harold Swerdlow, John Burton, Carl Skuce, Ruth Taylor, Zahra Abdellah, Yali Zhao, Asan, Daniel G. MacArthur, Michael A. Quail, Nigel P. Carter, Huanming Yang andChris Tyler-Smith
Current Biology, 27 August 2009, doi:10.1016/j.cub.2009.07.032

See also:
Team 19: Human Evolution
The Wellcome Trust Sanger Institute
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ZenMaster

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Unique Study Isolates DNA from Linnaeus' Botanical Collections

2009-08-27T23:05:00.002+02:00

Unique Study Isolates DNA from Linnaeus' Botanical Collections
Thursday, 27 August 2009

Researchers at Uppsala University has succeeded in extracting long DNA fragments from dried, pressed plant material collected in the 1700s by Linnaeus' apprentice Adam Afzelius. It is hoped that the study, led by Associate Professor Katarina Andreasen, will shed light on whether plants growing today at Linnaeus' Hammarby estate outside Uppsala reflect the species cultivated by Linnaeus himself.

A large number of plants of uncertain provenance grow at Carl Linnaeus' Hammarby estate, a museum and popular tourist destination. Have they been present since Linnaeus' time? In addition to probing this question, the current study will test the limits of DNA-sequencing methods with regard to old plant material and has already demonstrated that it is possible to sequence plant material more than 200 years old. The study is now published in the scientific journal Taxon.

"This opens up a number of exciting research possibilities in connection with material from herbaria throughout the world", says Associate Professor Katarina Andreasen.

The researchers hope to initiate corresponding DNA investigations of plant material from the garden at Hammarby as soon as possible.

"It would be fun, if we can show that the old material is genetically identical with the plants currently growing at Hammarby, to create a living herbarium for summer visitors to the garden", says Katarina Andreasen.

Linnaeus' significance for the science of systematic biology, as reflected in locations in Sweden (Uppsala and Småland) and collection locations in seven other countries, is the focus of a World Heritage Site nomination. Carl Linnaeus laid the foundations of systematic biology through the aid of an extensive scientific network. If UNESCO approves the nomination, preserved animals and plants will constitute, for the first time, a central aspect of a World Heritage Site.

Links:
Linnaeus' Hammarby
The Linnaeus Garden
The Linnean Society of London
The Swedish Linnaeus Society
The Linnaeus Herbarium
The Linnaean Correspondence
Carl von Linné on Internet
.........

ZenMaster

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Mitochondrial Gene Replacement in Primate Offspring and Embryonic Stem Cells

2009-08-26T22:44:00.001+02:00

Breakthrough could help break the chain of several maternally-based diseases passed from generation to generation
Wednesday, 26 August 2009

Researchers at Oregon Health & Science University's Oregon National Primate Research Center (ONPRC) believe they have developed one of the first forms of genetic therapy – a therapy aimed at preventing serious diseases in unborn children. Specifically, the therapy would combat inherited diseases passed on from mothers to their children through mutated DNA in cell mitochondria. The research is published in the Aug. 26 advance online edition of the journal Nature.

"We believe this discovery in nonhuman primates can rapidly be translated into human therapies aimed at preventing inherited disorders passed from mothers to their children through the mitochondrial DNA, such as certain forms of cancer, diabetes, infertility, myopathies and neurodegenerative diseases," explained Shoukhrat Mitalipov, Ph.D.. Dr. Mitalipov is an associate scientist in the Division of Reproductive Sciences at ONPRC, the Oregon Stem Cell Center and the departments of Obstetrics and Gynecology and Molecular & Medical Genetics of Oregon Health & Science University (OHSU).

"Currently there are 150 known diseases caused by mutations of the mitochondrial DNA, and approximately 1 out of every 200 children is born with mitochondrial mutations."

Mitochondria are structures that are found in all cells that provide energy for cell growth and metabolism, which is why they are often called the cell's "power plant." The structures produce energy to power each individual cell. Mitochondria also carry their own genetic material.

When an egg cell is fertilized by a sperm cell during reproduction, the embryo almost exclusively inherits the maternal mitochondria present in the egg. This means that any disease-causing genetic mutations that a mother carries in her mitochondrial DNA can be passed on to her offspring. The method developed by OHSU researchers transfers the mother's chromosomes to a donated egg that has had its chromosomes removed, but which has healthy mitochondria, thereby preventing the disease from being passed on to one's offspring.

How the OHSU researchers' method works
Scientists collected groups of unfertilized eggs from two female rhesus macaque monkeys (monkeys A and B). They then removed the chromosomes, which contain the genes found in the cell nucleus, from the eggs of monkey B, and then transplanted the nuclear genes from the eggs of monkey A into the eggs of monkey B. Then the eggs from monkey B, which now contained their own mitochondria but monkey A's nuclear genes, were fertilized. The fertilized eggs developed into embryos that were implanted in surrogate monkeys.

The initial implantation of two embryos resulted in the birth of healthy twin monkeys, nicknamed "Mito" and "Tracker" (in reference to the procedure used for imaging of mitochondria). These monkeys are the world's first animals derived by spindle transfer.

Follow-up testing showed that there was little to no trace of cross-animal mitochondrial transfer using this procedure. This demonstrates that the researchers were successful in isolating nuclear genetic material from mitochondrial genetic material during the transfer process.

"In theory, this research has demonstrated that it is possible to use this therapy in mothers carrying mitochondrial DNA diseases so that we can prevent those diseases from being passed on to their offspring," added Mitalipov.

"We believe that with the proper governmental approvals, our work can rapidly be translated into clinical trials for humans, and, eventually, approved therapies."

"This breakthrough is an excellent example of how OHSU's research findings can often be rapidly translated into health therapies that benefit residents of our state and the country as a whole," said Dr. Joe Robertson, M.D., M.B.A., president of OHSU.

"Recent findings suggest that mitochondrial disorders play a role in at least some proportion of many human disorders," said Duane Alexander, M.D, director of the Eunice Kennedy Shriver National Institute of Child Health and Human Development, which provided funding for the study.

"Pending further research, the findings hold the potential of allowing a couple to have a child who is biologically their own, but is free of any conditions associated with defects in maternal mitochondria."

Using the technique, the researchers created fertilized eggs and achieved three successful pregnancies in rhesus monkeys, which have resulted in four healthy newborns. Recent advances in the transfer of hereditary material and in microscopy facilitated the achievement, they wrote.

The researchers said that the technique did not appear to pose any risk of chromosomal damage. Analysis of 5-6-day-old embryos (blastocysts) resulting from the fertilized eggs, and of embryonic stem cell lines established from them, did not uncover any evidence of damage to the chromosomes. Analysis of cells from the infant monkeys born after the procedure failed to detect any mitochondrial DNA from the mother.

Reference:
Mitochondrial gene replacement in primate offspring and embryonic stem cells
Masahito Tachibana, Michelle Sparman, Hathaitip Sritanaudomchai, Hong Ma, Lisa Clepper, Joy Woodward, Ying Li, Cathy Ramsey, Olena Kolotushkina & Shoukhrat Mitalipov
Nature advance online publication 26 August 2009, doi:10.1038/nature08368

See also:
DNA swap could avoid inherited diseases
David Cyranoski
Nature News 26 August 2009, doi:10.1038/news.2009.860
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ZenMaster

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Scientists Grows Retina Cells from Skin-derived Stem Cells

2009-08-24T22:13:00.003+02:00

Scientists Grows Retina Cells from Skin-derived Stem Cells
Monday, 24 August 2009

A team of scientists from the University of Wisconsin-Madison School of Medicine and Public Health has successfully grown multiple types of retina cells from two types of stem cells — suggesting a future in which damaged retinas could be repaired by cells grown from the patient's own skin.

Even sooner, the discovery will lead to laboratory models for studying genetically linked eye conditions, screening new drugs to treat those conditions and understanding the development of the human eye.

A Waisman Center research team led by David Gamm, an assistant professor of ophthalmology and visual sciences, and Jason Meyer, a research scientist, announced their discovery in the Aug. 24 edition of the Proceedings of the National Academy of Sciences.

"This is an important step forward for us, as it not only confirms that multiple retinal cells can be derived from human iPS cells using the Wisconsin approach, but also shows how similar the process is to normal human retinal development," Gamm says.

"That is quite remarkable given that the starting cell is so different from a retinal cell and the whole process takes place in a plastic dish. We continue to be amazed at how deep we can probe into these early events and find that they mimic those found in developing retinas. Perhaps this is the way to close the gap between what we know about building a retina in mice, frogs and flies with that of humans."

Gamm says the work built on the strong tradition of stem cell research at UW-Madison. James Thomson, a School of Medicine and Public Health faculty member and director of regenerative medicine at the Morgridge Institute for Research on the UW-Madison campus, announced that he had made human stem cells from skin, called induced pluripotent stem (iPS cells), in November 2007. Su-Chun Zhang, UW-Madison professor of anatomy and a Waisman researcher, was among the first to create neural cells from embryonic stem cells. Zhang was also part of the Gamm lab's retinal study.

A team of scientists from the University of Wisconsin-Madison's School of Medicine and Public Health, led by David Gamm, assistant professor of ophthalmology and visual sciences, and Jason Meyer, research scientist, has successfully grown multiple types of retina cells from two types of stem cells — suggesting a future in which damaged retinas could be repaired by cells grown from the patient's own skin. Pictured here in a microscopic photograph are early retinal cells (green) and early brain cells (blue). Credit: courtesy David Gamm.

Meyer says the retina project began by using embryonic stem cells, but incorporated the iPS cells as they became available. Ultimately, the group was able to grow multiple types of retina cells beginning with either type of stem cell, starting with a highly enriched population of very primitive cells with the potential to become retina. This is critical, as it reduces contamination from unwanted cells early in the process. In normal human development, embryonic stem cells begin to differentiate into more specialized cell types about five days after fertilization. The retina develops from a group of cells that arise during the earliest stages of the developing nervous system. The Wisconsin team took cells from skin, turned them back into cells resembling embryonic stem cells, then triggered the development of retinal cell types.

"This is one of the most comprehensive demonstrations of a cell-based system for studying all of the key events that lead to the generation of specialized neural cells,'' Meyer says.

"It could serve as a foundation for unlocking the mechanisms that produce human retinal cells."

Because the group was successful using the iPS cells, they expect this advance to lead to studying retinal development in detail and treating conditions that are genetically linked. For example, skin from a patient with retinitis pigmentosa could be reprogrammed into iPS cells, then retina cells, which would allow researchers to screen large numbers of potential drugs for treating or curing the condition.

Likewise, someday ophthalmologists may be able to repair damage to the retina by growing rescue or repair cells from the patient's skin. Earlier this year, scientists from the University of Washington showed that human ES cells had the potential to replace retinal cells lost during disease in mice.

"We're able to produce significant numbers of photoreceptor cells and other retinal cell types using our system, which are lost in many disorders," Meyer says. Photoreceptors are light-sensitive cells that absorb light and transmit the image as an electrical signal to the brain.

The team had similar success in creating the multiple specialized types of retina cells from embryonic stem cells, underscoring the similarities between ES and iPS cells. However, Gamm emphasizes that there are differences between these cell types as well. More work is needed to understand their potential and their limitations.
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ZenMaster

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'Glow-in-the-dark' Red Blood Cells Made from Human Embryonic Stem Cells

2009-08-24T19:22:00.001+02:00

'Glow-in-the-dark' Red Blood Cells Made from Human Embryonic Stem Cells
Monday, 24 August 2009

Victorian stem cell scientists from Monash University have modified a human embryonic stem cell (hESC) line to glow red when the stem cells become red blood cells.

The modified hESC line, ErythRED, represents a major step forward to the eventual aim of generating mature, fully functional red blood cells from human embryonic stem cells.

The research, conducted by a team led by Professors Andrew Elefanty and Ed Stanley at the Monash Immunology and Stem Cell Laboratories that included scientists at the Murdoch Children's Research Institute, was published in today's issue of the prestigious journal, Nature Methods.

The work, funded by the Australian Stem Cell Centre (ASCC), will help scientists to track the differentiation of embryonic stem cells into red blood cells.

Whilst hESCs have the potential to turn into any cell type in the body, it remains a scientific challenge to reliably turn these stem cells into specific cell types such as red blood cells. The development of the ErythRED embryonic stem cell line, which fluoresces red when haemoglobin genes are switched on, is an important development that will help researchers to optimise the conditions that generate these cells.

"The elegant work of the Elefanty-Stanley group unlocks the entrance to the long sought and elusive differentiation pathway that leads to expression of adult haemoglobin genes," said Professor Joe Sambrook, Scientific Director of the ASCC.

"Not only will the ErythRED cell line lead to more efficient creation of red blood cells from human embryonic stem cells, but these cells are a crucial tool for monitoring the behaviour of the cells when transplanted into animal models" said Professor Andrew Elefanty.

Reference:
ErythRED, a hESC line enabling identification of erythroid cells
Tanya Hatzistavrou, Suzanne J Micallef, Elizabeth S Ng, Jim Vadolas, Edouard G Stanley & Andrew G Elefanty
Nature Methods, Published online: 23 August 2009, doi:10.1038/nmeth.1364
.........

ZenMaster

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