Researchers Engineer Miniature Human Livers in the Lab

Researchers Engineer Miniature Human Livers in the Lab
Sunday, 31 October 2010

Researchers at the Institute for Regenerative Medicine at Wake Forest University Baptist Medical Center have reached an early, but important, milestone in the quest to grow replacement livers in the lab. They are the first to use human liver cells to successfully engineer miniature livers that function – at least in a laboratory setting – like human livers. The next step is to see if the livers will continue to function after transplantation in an animal model.

The ultimate goal of the research, which will be presented Sunday at the annual meeting of the American Association for the Study of Liver Diseases in Boston, is to provide a solution to the shortage of donor livers available for patients who need transplants. Laboratory-engineered livers could also be used to test the safety of new drugs.

"We are excited about the possibilities this research represents, but must stress that we're at an early stage and many technical hurdles must be overcome before it could benefit patients," said Shay Soker, Ph.D., professor of regenerative medicine and project director.

"Not only must we learn how to grow billions of liver cells at one time in order to engineer livers large enough for patients, but we must determine whether these organs are safe to use in patients."

Pedro Baptista, PharmD, Ph.D., lead author on the study, said the project is the first time that human liver cells have been used to engineer livers in the lab.

"Our hope is that once these organs are transplanted, they will maintain and gain function as they continue to develop," he said.

To engineer the organs, the scientists used animal livers that were treated with a mild detergent to remove all cells (a process called de-cellularization), leaving only the collagen "skeleton" or support structure. They then replaced the original cells with two types of human cells: immature liver cells known as progenitors, and endothelial cells that line blood vessels.

The cells were introduced into the liver skeleton through a large vessel that feeds a system of smaller vessels in the liver. This network of vessels remains intact after the de-cellularization process. The liver was next placed in a bioreactor, special equipment that provides a constant flow of nutrients and oxygen throughout the organ.

After a week in the bioreactor system, the scientists documented the progressive formation of human liver tissue, as well as liver-associated function. They observed widespread cell growth inside the bioengineered organ.

The ability to engineer a liver with animal cells had been demonstrated previously. However, the possibility of generating a functional human liver was still in question.

The researchers said the current study suggests a new approach to whole-organ bioengineering that might prove to be critical not only for treating liver disease, but for growing organs such as the kidney and pancreas. Scientists at the Wake Forest Institute for Regenerative Medicine are working on these projects, as well as many other tissues and organs, and also working to develop cell therapies to restore organ function.

Bioengineered livers could also be useful for evaluating the safety of new drugs.

"This would more closely mimic drug metabolism in the human liver, something that can be difficult to reproduce in animal models," said Baptista.

Source: Wake Forest University Baptist Medical Center
Contact: Karen Richardson
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

Is the Shape of a Genome as Important as Its Content?

Study shows structure exposes genes to regulation and chromosomal crosstalk
Friday, 29 October 2010

The laboratory of Ken-ichi Noma, Ph.D., an assistant

professor at the Wistar Institute, has produced the

first detailed structure of the fission yeast genome.

The researchers demonstrate how the physical

structure of the genome itself helps cells regulate

and control gene expression. Credit: Ken-ichi Noma,

Ph.D./The Wistar Institute.

If there is one thing that recent advances in genomics have revealed, it is that our genes are interrelated, "chattering" to each other across separate chromosomes and vast stretches of DNA. According to researchers at The Wistar Institute, many of these complex associations may be explained in part by the three-dimensional structure of the entire genome. A given cell's DNA spends most of its active lifetime in a tangled clump of chromosomes, which positions groups of related genes near to each other and exposes them to the cell's gene-controlling machinery. This structure, the researchers say, is not merely the shape of the genome, but also a key to how it works.

Their study, published online as a featured article in the journal Nucleic Acids Research, is the first to combine microscopy with advanced genomic sequencing techniques, enabling researchers to literally see gene interactions. It is also the first to determine the three-dimensional structure of the fission yeast genome, S. pombe. Applying this technique to the human genome may provide both scientists and physicians a completely new framework from which to better understand genes and disease, the researchers say.

"People are familiar with the X-shapes our chromosomes form during cell division, but what they may not realize is that DNA only spends a relatively small amount of time in that conformation," said Ken-ichi Noma, Ph.D., an assistant professor in Wistar's Gene Expression and Regulation program and senior author of the study.

"Chromosomes spend the majority of their time clumped together in these large, non-random structures, and I believe these shapes reflect various nuclear processes such as transcription."

To map both individual genes and the overall structure of the genome, Noma and his colleagues combined next generation DNA sequencing with a technique called chromosome conformation capture (3C). They then used fluorescent probes to pinpoint the exact location of specific genes through a microscope. With these data, the researchers were able to create detailed three-dimensional computer models of the yeast genome.

Using this novel approach, the researchers can view genes as they interact with each other. Noma and his colleagues can view where highly active genes are located, or see if genes that are turned on and off together also reside near each other in the three-dimensional structure of the genome. In total, the Wistar researchers also studied 465 so-called gene ontology groups – groups of genes that share a related purpose in the cell, such as structure or metabolism.

"When the chromosomes come together, they fold into positions that bring genes from different chromosomes near each other," Noma said.

"This positioning allows the processes that dictate how and when genes are read to operate efficiently on multiple genes at once."

This structure is not merely an accident of chemical attractions within and among the chromosomes – although that is certainly a part of the larger whole – but an arrangement guided by other molecules in the cell to create a mega-structure that dictates genetic function, Noma says. He envisions a scenario where accessory molecules, such as gene-promoting transcription factors, bind to DNA and contribute to the ultimate structure of the genome as the chromosomes fold together.

"I believe we are looking at a new way to visualize both the genome itself and the movements of all the various molecules that act on the genome," Noma said.

According to the Wistar scientists, their techniques are scalable to the human genome, even though fission yeast only has three chromosomes. In fact, the researchers found signs of "transcription factories" – clusters of related genes that are read, or "transcribed," at discrete sites – which have been proposed to exist in mammals.

Source: The Wistar Institute
Contact: Greg Lester

Reference:
Mapping of long-range associations throughout the fission yeast genome reveals global genome organization linked to transcriptional regulation
Hideki Tanizawa, Osamu Iwasaki, Atsunari Tanaka, Joseph R. Capizzi, Priyankara Wickramasinghe, Mihee Lee, Zhiyan Fu, and Ken-ichi Noma
Nucl. Acids Res., published online October 28, 2010, doi:10.1093/nar/gkq955
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

Researchers Generate iPS Cells to Further Treatments for Lung Diseases

Researchers Generate iPS Cells to Further Treatments for Lung Diseases
Friday, 29 October 2010

A team of researchers from Boston University's Center for Regenerative Medicine and the Pulmonary Center have generated 100 new lines of human induced pluripotent stem cells (iPSC) from individuals with lung diseases, including cystic fibrosis and emphysema. The new stem cell lines could possibly lead to new treatments for these debilitating diseases. The findings, which appear in the current issue of Stem Cells, demonstrate the first time lung disease-specific iPSC have been created in a lab.

iPSCs are derived by reprogramming adult cells into a primitive stem cell state. This process results in the creation of cells that are similar to embryonic stem cells in terms of their capability to differentiate into different types of cells, including endoderm cells that can give rise to liver and lung tissue.

"iPSCs solve many major hurdles currently impacting embryonic stem cell research," said Darrell Kotton, the study's lead author and associate professor of medicine and pathology and laboratory medicine at Boston University's School of Medicine (BUSM).

iPSCs do not require embryos, and the process used to cultivate iPSCs is easier than the techniques used to obtain embryonic stem cells. iPSCs are genetically identical to the patient's cells and potentially can be transplanted back without rejection.

"In a laboratory dish, these cells have the ability to multiply indefinitely so that researchers have more time to investigate the diseased cell and correct its genes," said Kotton.

The study involved patients with different forms of lung disease – cystic fibrosis, alpha-1 antitrypsin deficiency-related emphysema, scleroderma (SSc) and sickle cell disease. The patients underwent skin biopsies and donated tissue samples, which the research team used to cultivate adult stem cells. Using a Boston University-patented vector in the form of a virus, named the Stem Cell Cassette (STEMCCA), the researchers were able to reprogram the skin cells into the primitive pluripotent stem cells known as iPSCs.

"The STEMCCA vector is proving invaluable for reprogramming cells from a variety of species, and this is the first report of the 'humanized' version of our vector for use in reprogramming human cells," said Gustavo Mostoslavsky, a co-author of the study and assistant professor of medicine at BUSM. Together Kotton and Mostoslavsky co-direct the new Boston University Center for Regenerative Medicine (CReM).

To test the differentiation power of the iPSCs, the team showed that the stem cells multiplied and could be differentiated into endoderm tissue, the natural precursor cells of the lung, the primary organ destroyed by the diseases cystic fibrosis and emphysema.

"We hope to build a bank of stem cells that could be used to help treat the two most common forms of inherited lung disease, cystic fibrosis and alpha-1 antitrypsin deficiency," said Kotton.

The next step, he said, is to correct the genetic mutations responsible for causing cystic fibrosis, emphysema and other lung diseases.

Source: Boston University Medical Center
Contact: Gina DiGravio

Reference:
Generation of Transgene-Free Lung Disease-Specific Human Induced Pluripotent Stem Cells Using a Single Excisable Lentiviral Stem Cell Cassette
Aba Somers, Jyh-Chang Jean, Cesar A. Sommer, Amel Omari, Christopher C. Ford, Jason A. Mills, Lei Ying, Andreia Gianotti Sommer, Jenny M. Jean, Brenden W. Smith, Robert Lafyatis, Marie-France Demierre, Daniel J. Weiss, Deborah L. French, Paul Gadue, George J. Murphy, Gustavo Mostoslavsky and Darrell N. Kotton
Stem Cells Vol. 28, Issue 10, pp. 1728–1740, October 2010, DOI: 10.1002/stem.495
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

New Methods Detect Subtleties in Human Genomes Repetitive Landscapes

These techniques spot minute variations linked to evolution, diversity and brain development
Thursday, 28 October 2010

Scientists have invented methods to scout the human genome's repetitive landscapes, where DNA sequences are highly identical and heavily duplicated. These advances, as reported today in Science, can identify subtle but important differences among people in the number and content of repeated DNA segments.

These copy number variations partly account for the normal diversity among people. Copy number variations might also be why some people, and not others, have certain disorders or disease susceptibilities, and might also determine how severely they are affected.

Until about a year ago, locating and counting the number of duplicated copies of DNA sequences was almost impossible. The more copies of a duplicated gene that are present, the harder they are to assess accurately.

"These difficulties resulted in a lack of understanding of the true extent of human copy number variation, " said Dr. Evan E. Eichler, University of Washington (UW) professor of genome sciences and senior author of the Science paper.

"The most dynamic and variable genes are frequently excluded from genome-wide studies."

These hard-to-study genes are also among the most interesting because of their suspected contributions to human evolution, brain development, metabolism and disease immunity.

Researchers in Eichler's lab have developed several analytical and computational techniques to overcome obstacles in looking at multi-copy genes. The lead authors of the study are Peter H. Sudmant and Jacob O. Kitzman, both graduate students in the UW Department of Genome Sciences.

Working with colleagues in the 1000 Genomes Project and at Agilent Technologies, the UW group used the new techniques to deeply probe and compare the genomes of 159 individuals. In assessing the entire genomes of these individuals, the researchers were able to accurately assay previously intractable duplicated genes and gene families.

The researchers demonstrated that the methods could estimate correctly the absolute number of copies of segments as small as 1,900 DNA base pairs, and could count numbers of copies ranging from zero to 48. A human genome is made up of about 3 billion DNA base pair. Each pair consists of two bonded molecules called nucleotides, the basic structural unit of DNA.

"We identified 4.1 million singly unique nucleotide positions informative in distinguishing specific copies," the authors reported. The researchers took this information to genotype the number of copies and the content of genes that had been duplicated to or more different positions on the genome thereby became free to function on their own. These duplicated genes reveal changes that occurred during evolution.

The data allowed the researchers to identify duplicated genes specific to humans, in comparison to apes like gorilla, orang-utans, and chimps. The researchers observed that these duplications occurred in genes associated with brain development. These include genes implicated in the growth and branching of brain cell connections, in abnormally large or small head size, in a particular dopamine (reward signal in the brain) receptor, in visual-spatial and social deficits, in reducing the severity of spinal muscular atrophy, and in intellectual disability and epilepsy.

Copy number variations occur in only about 7 percent to 9 percent of human genes, the researchers found. Most of our genes come standard: two copies. Even among copy number variable genes, the researchers learned that 80 percent of them vary between zero and five copies.

"Extreme gene variation," the researchers noted, "is limited to only a few gene families." In this study, they identified 56 of the most variable gene families. These ranged in median copy number from five to approximately 368.

"These genes were dramatically enriched for segmental duplication," the researchers noted. Segmental duplications are regions that were originally identified in the Human Genome Project as long, repeated blocks of the genome.

The researchers report discovering about 44 "hidden" members of duplicated gene families never before identified in the reference model of the human genome.

"The missing members of these gene families," the researchers suggested, "should be targeted for sequence finishing in order to more accurately capture the architecture and diversity of the human genome."

While duplications of segments of the genome appear to have led to many of the qualities that distinguish human beings from other primate species, areas of the genome in which duplications promote recurrent rearrangements have also been associated with debilitating diseases like intellectual disability, schizophrenia and autism.

"Extreme variation resulting from duplications may contribute to genomic instability associated with disease," the researchers therefore hypothesize.

Overall, the results of the study shows scientists can now leverage newly developed techniques to explore some of the most complex genetic regions of the human genome. Still, a portion of the genome remains impenetrable. About 28 large regions of the human genome have such extraordinary complexity that as yet it is impossible to interpret the underlying pattern of genetic diversity, the authors said.

Despite this limitation, the approaches tested in the study hold promise for improving the understanding of how copy number variation contributes to human health and illness.

"Our approach makes many of the highly duplicated regions of the human genome – and the more than 1,000 previously inaccessible human genes that lie therein –accessible to genetic studies of disease association," the researchers concluded.

Source: University of Washington
Contact: Leila Gray

Reference:
Diversity of Human Copy Number Variation and Multicopy Genes
Peter H. Sudmant, Jacob O. Kitzman, Francesca Antonacci, Can Alkan, Maika Malig, Anya Tsalenko, Nick Sampas, Laurakay Bruhn, Jay Shendure, 1000 Genomes Project, Evan E. Eichler
Science 29 October 2010, Vol 330, 641-646.
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

1000 Genomes Project Publishes Analysis of Completed Pilot Phase

Produces tool for research into genetic contributors to human disease
Thursday, 28 October 2010



Human chromosomes.

 Small genetic differences between individuals help explain why some people have a higher risk than others for developing illnesses such as diabetes or cancer. Today in the journal Nature, the 1000 Genomes Project, an international public-private consortium, published the most comprehensive map of these genetic differences, called variations, estimated to contain approximately 95 percent of the genetic variation of any person on Earth.

Researchers produced the map using next-generation DNA sequencing technologies to systematically characterize human genetic variation in 180 people in three pilot studies. Moreover, the full scale-up from the pilots is already under way, with data already collected from more than 1,000 people.

“The pilot studies of the 1000 Genomes Project laid a critical foundation for studying human genetic variation,” said Richard Durbin, Ph.D., of the Wellcome Trust Sanger Institute and co-chair of the consortium.

“These proof-of-principle studies are enabling consortium scientists to create a comprehensive, publicly available map of genetic variation that will ultimately collect sequence from 2,500 people from multiple populations worldwide and underpin future genetics research.”

Genetic variation between people refers to differences in the order of the chemical units — called bases — that make up DNA in the human genome. These differences can be as small as a single base being replaced by a different one — which is called a single nucleotide polymorphism (abbreviated SNP) — or is as large as whole sections of a chromosome being duplicated or relocated to another place in the genome. Some of these variations are common in the population and some are rare. By comparing many individuals to one another and by comparing one population to other populations, researchers can create a map of all types of genetic variation.

The 1000 Genomes Project’s aim is to provide a comprehensive public resource that supports researchers aiming to study all types of genetic variation that might cause human disease. The project’s approach goes beyond previous efforts in capturing and integrating data on all types of variation, and by studying samples from numerous human populations with informed consent allowing free data release without restriction on use. Already, these data have been used in studies of the genetic basis for disease.

“By making data from the project freely available to the research community, it is already impacting research for both rare and common diseases,” said David Altshuler, M.D., Ph.D., Deputy Director of the Broad Institute of Harvard and MIT, and a co-chair of the project.

“Biotech companies have developed genotyping products to test common variants from the project for a role in disease. Every published study using next-generation sequencing to find rare disease mutations, and those in cancer, used project data to filter out variants that might obscure their results.”

The project has studied populations with European, West African and East Asian ancestry. Using the newest technologies for sequencing DNA, the project’s nine centres sequenced the whole genome of 179 people and the protein-coding genes of 697 people. Each region was sequenced several times, so that more than 4.5 terabases (4.5 million million base letters) of DNA sequence were collected. A consortium involving academic centres on multiple continents and technology companies that developed and sell the sequencing equipment carried out the work.

To process these data required many technical and computational innovations, including standardized ways to organize, store, analyze and share DNA sequencing data. Launched in 2008, the 1000 Genomes Project started with three pilot projects to develop, evaluate and compare strategies for producing a catalogue of genetic variations. Funded through numerous mechanisms by foundations and national governments, the 1000 Genome Project will cost some $120 million over five years, ending in 2012.

When the work began, sequencing was very expensive, so the project began with two approaches aimed at increasing efficiency: One strategy, called “low-pass”, combines partial data from many people; the second, only focused on the part of the genome that encodes protein-coding genes. By comparing these strategies to “gold standard” data produced at great completeness and accuracy, the project was able to show that both the alternative approaches work well and have complementary strengths. Researchers will use both strategies in the full-scale project because, although sequencing costs have decreased, it is still relatively expensive.

“We have shown for the first time that a new approach to sequencing — low coverage of many samples — works efficiently and well,” said Gil McVean, Ph.D., Professor of Statistical Genetics at the University of Oxford.

“This proof of principle is now being applied not only in the 1000 Genomes Project, but in disease research, as well.”

The resulting map of human genetic variation includes about 15 million SNPs, 1 million short insertion/deletion changes, and more than 20,000 structural variations. Many of the genetic variants had previously been identified, but more than half were new. The project’s database contains more than 95 percent of the currently measurable variants found in any individual, and continuing work will eventually identify more than 99 percent of human variants.

Richard Gibbs, Ph.D., director of the Human Genome Sequencing Center at the Baylor College of Medicine (one of the project’s sequencing centres) said:

“What really excites me about this project is the focus on identifying variants in the protein-coding genes that have functional consequences. These will be extremely useful for studies of disease and evolution.”

The improved map produced some surprises. For example, the researchers discovered that on average, each person carries between 250 and 300 genetic changes that would cause a gene to stop working normally, and that each person also carried between 50 and 100 genetic variations that had previously been associated with an inherited disease. No human carries a perfect set of genes. Fortunately, because each person carries at least two copies of every gene, individuals likely remain healthy, even while carrying these defective genes, if the second copy works normally.

In addition to looking at variants that are shared between many people, the researchers also investigated in detail the genomes of six people: two mother-father-daughter nuclear families. By finding new variants present in the daughter but not the parents, the team was able to observe the precise rate of mutations in humans, showing that each person has approximately 60 new mutations that are not in either parent.

With the completion of the pilot phase, the 1000 Genomes Project has moved into full-scale studies in which 2,500 samples from 27 populations will be studied over the next two years. Data from the pilot studies and the full-scale project are freely available on the project web site, http://www.1000genomes.org/.

Researchers studying specific illnesses, such as heart disease or cancer, use maps of genetic variation to help them identify genetic changes that may contribute to the illnesses. Over the last five years, the first generation of such studies (called genome-wide association studies or GWAS) have been based on an earlier map of genetic variation called the HapMap. Built using older technology, HapMap lacks the completeness and detail of the 1000 Genomes Project.

“The 1000 Genomes Project map fills in the gaps between the HapMap landmarks, helping researchers identify all candidate genes in a region associated with a disease,” said Lisa Brooks, Ph.D., program director for genetic variation at the National Human Genome Research Institute, a part of the National Institutes of Health.

“Once a disease-associated region of the genome is identified, experimental studies must be done to identify which variants, genes, and regulatory elements cause the increased disease risk. With the new map, researchers can just look up all the candidate genes and almost all of the variants in the database, saving them many steps in finding the causes.”

Additional information about the project, including a list of all participants and organizations, can be found at http://www.1000genomes.org/.

About the National Institutes of Health:
The National Institutes of Health - "The Nation's Medical Research Agency" - is a component of the U.S. Department of Health and Human Services. It is the primary federal agency for conducting and supporting basic, clinical and translational medical research, and it investigates the causes, treatments and cures for both common and rare diseases.


About the National Human Genome Research Institute:
The National Human Genome Research Institute is one of 27 institutes and centers at National Institutes of Health, an agency of the Department of Health and Human Services. NHGRI's Division of Extramural Research supports grants for research and for training and career development.


About the Wellcome Trust:
The Wellcome Trust is a global charitable foundation dedicated to achieving extraordinary improvements in human and animal health. It is independent of both political and commercial interests.


About the Wellcome Trust Sanger Institute:
The Wellcome Trust Sanger Institute, which receives the majority of its funding from the Wellcome Trust, was founded in 1992. In October 2006, new funding was awarded by the Wellcome Trust to exploit the wealth of genome data now available to answer important questions about health and disease.


About the European Molecular Biology Laboratory:
The European Molecular Biology Laboratory is a basic research institute funded by public research monies from 20 member countries and supports research by approximately 85 independent groups covering the spectrum of molecular biology.

About the European Bioinformatics Institute:
European Bioinformatics Institute (EBI) is part of the European Molecular Biology Laboratory (EMBL) and is located on the Wellcome Trust Genome Campus in Hinxton near Cambridge (UK).


About the Broad Institute of MIT and Harvard:
The Eli and Edythe L. Broad Institute of MIT and Harvard, founded in 2003 by MIT, Harvard and its affiliated hospitals, and Los Angeles philanthropists Eli and Edythe L. Broad, includes faculty, professional staff and students from throughout the MIT and Harvard biomedical research communities and beyond, with collaborations spanning over a hundred private and public institutions in more than 40 countries worldwide.


Organizations that committed major support to the project include: 454 Life Sciences, a Roche company, Branford, Conn.; Life Technologies Corporation, Carlsbad, Calif.; BGI-Shenzhen, Shenzhen, China; Illumina Inc., San Diego; the Max Planck Institute for Molecular Genetics, Berlin, Germany; the Wellcome Trust Sanger Institute, Hinxton, Cambridge, UK; and the National Human Genome Research Institute, which supports the work being done by Baylor College of Medicine, Houston, Texas; the Broad Institute, Cambridge, Mass.; and Washington University, St. Louis, Missouri. Researchers at many other institutions are also participating in the project including groups in Barbados, Canada, China, Colombia, Finland, the Gambia, India, Malawi, Pakistan, Peru, Puerto Rico, Spain, the UK, the US, and Vietnam.


Source: Broad Institute of MIT and Harvard
Contact: Nicole Davis

Reference:
A map of human genome variation from population scale sequencing
The 1000 Genomes Project Consortium
Nature 467, 1061–1073, (28 October 2010), DOI:10.1038/nature09534
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

Genome-wide Study of Human Stem Cells

Landmark identification of key human stem cell genes paves the way for future stem cell research
Tuesday, 26 October 2010

A team of scientists from Singapore led by the Genome Institute of Singapore (GIS) and the Institute of Molecular and Cell Biology (IMCB), two biomedical research institutes of Singapore's Agency of Science, Technology and Research (A*STAR), have discovered the most important genes in human embryonic stem cells (hESCs), a crucial breakthrough in discovering how human stem cells work.

Their research, published in the journal Nature, is the first ever genome-wide study of human stem cells on such a massive scale, and its results are crucial in understanding how stem cells may one day be used to treat debilitating conditions such as Parkinson's disease and traumatic spinal injury.

GIS Senior Group Leader for Stem Cell and Development Biology and Associate Director for Biology Dr Ng Huck Hui, and IMCB Principal Investigator Dr Frederic Bard combined the strengths of their teams. They investigated the 21,000 genes in the entire human genome to find those, which regulate the two characteristic properties of hESCs – the capacity to turn into any type of cell in the human body (pluripotency), and the ability to retain that capacity indefinitely. Out of the several key genes they identified, a particular gene known as PRDM14 was discovered to make it easier to turn a type of human cell (fibroblasts) into pluripotent stem cells. The discoveries contribute to a fundamental understanding of the nature of stem cells and helps efforts to improve techniques to turn mature adult cells into hESCs.

In addition, the scientists found that PRDM14 played a key role in hESCs, but not in mouse ESCs. This significant new finding highlights the fundamental differences between stem cells from different species, and highlights the greater need to use human cells in stem cell research.

"Very little is known about the molecular machines that drive stem cell states or the transcriptional profiles of hESCs. Our study helps to build a better understanding of hESCs and this will help in the development of technologies to further the utilities of these cells such as their potential to be used for clinical and therapeutic applications," said Dr Ng.

"Dr Bard's scientific expertise was invaluable in helping us crack another piece of the stem cell puzzle. I definitely look forward to collaborating with him on more projects that aim to peel away the mysteries surrounding stem cells," he added.

Dr Alan Colman, Executive Director of the Singapore Stem Cell Consortium, said:

"Huck Hui Ng and his colleagues continue to keep Singapore at the top table of countries plundering the secrets of human embryonic stem cell regulation. This time they have deployed the first genome-wide functional screen to identify factors that maintain 'stemness' in these cells and yet again reveal major differences between mouse and human embryonic stem cells in the control of this important property."

Senior Scientist at the Developmental & Stem Cell Biology Program, The Hospital for Sick Children, Dr Janet Rossant added:

"The unprecedented scale of this screen has added considerable new information to our understanding of pluripotency and will help efforts to improve reprogramming of adult cells."

Professor Lee Eng Hin, Executive Director of the Biomedical Research Council, A*STAR, applauded the discovery and said:

"This is an example of a great cross institutional collaboration. The combined strength of stem cell and genomics experts has led to a great piece of world-class work. I hope to see more of such valuable partnerships in the future."

Source: Agency for Science, Technology and Research (A*STAR), Singapore
Contact: Joshua Tan
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

How Genes are Selectively Silenced

How Genes are Selectively Silenced
Monday, 25 October 2010

Our genetic material is often compared to a book. However, it is not so much like a novel to be read in one piece, but rather like a cookbook. The cell reads only those recipes which are to be cooked at the moment. The recipes are the genes; 'reading' in the book of the cell means creating RNA copies of individual genes, which will then be translated into proteins.

The cell uses highly complex, sophisticated regulatory mechanisms to make sure that not all genes are read at the same time. Particular gene switches need to be activated and, in addition, there are particular chemical labels in the DNA determining which genes are transcribed into RNA and which others will be inaccessible, i.e. where the book literally remains closed. The biological term for this is epigenetic gene regulation.

Among the epigenetic mechanisms, which are well studied, is the silencing of genes by methyl groups. This is done by specialized enzymes called methyltransferases, which attach methyl labels to particular 'letters' of a gene whereby access to the whole gene is blocked.

"One of the great mysteries of modern molecular biology is: How do methyltransferases know where to attach their labels in order to selectively inactivate an individual gene?" says Professor Ingrid Grummt of the German Cancer Research Center (DKFZ).

Grummt has now come much closer towards unravelling this mystery. She has focused on studying those text passages in the genetic material, which do not contain any recipes. Nevertheless, these texts are transcribed into RNA molecules in a controlled manner.

"These so-called noncoding RNAs do not contain recipes for proteins. They are important regulators in the cell which we are just beginning to understand," says Ingrid Grummt.

In her most recent work, Grummt and her co-workers have shown for the first time that epigenetic regulation and regulation by noncoding RNAs interact. The scientists artificially introduced a noncoding RNA molecule called pRNA into cells. As a result, methyl labels are attached to a particular gene switch so that the genes behind it are not read. The trick is that pRNA exactly matches (is complementary to) the DNA sequence of this gene switch. The investigators found out that pRNA forms a kind of plait, or triple helix, with the two DNA strands in the area of this gene switch. Methyltransferases, in turn, are able to specifically dock to this 'plait' and are thus directed exactly to the place where a gene is to be blocked.

More than half of our genetic material is transcribed into noncoding RNA. This prompts Ingrid Grummt to speculate:

"It is very well possible that there are exactly matching noncoding RNA molecules for all genes that are temporarily silenced. This would explain how such a large number of genes can be selectively turned on and off."

Source: Helmholtz Association of German Research Centres
Contact: Dr. Sibylle Kohlstädt

Reference:
Interaction of noncoding RNA with the rDNA promoter mediates recruitment of DNMT3b and silencing of rRNA genes
Kerstin-Maike Schmitz, Christine Mayer, Anna Postepska and Ingrid Grummt
Genes & Development 2010, DOI: 10.1101/gad.590910
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

Gene Identified that Prevents Stem Cells from Turning Cancerous

Gene Identified that Prevents Stem Cells from Turning Cancerous
Friday, 15 October 2010

Stem cells, the prodigious precursors of all the tissues in our body, can make almost anything, given the right circumstances. Including, unfortunately, cancer. Now research from Rockefeller University shows that having too many stem cells, or stem cells that live for too long, can increase the odds of developing cancer. By identifying a mechanism that regulates programmed cell death in precursor cells for blood, or hematopoietic stem cells, the work is the first to connect the death of such cells to a later susceptibility to tumours in mice. It also provides evidence of the potentially carcinogenic downside to stem cell treatments, and suggests that nature has sought to balance stem cells' regenerative power against their potentially lethal potency.

Research associate Maria Garcia-Fernandez, Hermann Steller, head of the Strang Laboratory of Apoptosis and Cancer Biology, and their colleagues explored the activity of a gene called Sept4, which encodes a protein, ARTS, that increases programmed cell death, or apoptosis, by antagonizing other proteins that prevent cell death. ARTS was originally discovered by Sarit Larisch, a visiting professor at Rockefeller, and is found to be lacking in human leukaemia and other cancers, suggesting it suppresses tumours. To study the role of ARTS, the experimenters bred a line of mice genetically engineered to lack the Sept4 gene.

For several years, Garcia-Fernandez studied cells that lacked ARTS, looking for signs of trouble relating to cell death. In mature B and T cells, she could not find any, however, so she began to look at cells earlier and earlier in development, until finally she was comparing hematopoietic progenitor and stem cells. Here she found crucial differences, to be published Friday in Genes and Development.

Newborn ARTS-deprived mice had about twice as many hematopoietic stem cells as their normal, ARTS-endowed peers, and those stem cells were extraordinary in their ability to survive experimentally induced mutations.

"The increase in the number of hematopoietic progenitor and stem cells in Sept4-deficient mice brings with it the possibility of accelerating the accumulation of mutations in stem cells," says Garcia-Fernandez.

"They have more stem cells with enhanced resistance to apoptosis. In the end, that leads to more cells accumulating mutations that cannot be eliminated."

Indeed, the ARTS-deprived mice developed spontaneous tumours at about twice the rate of their controls.

"We make a connection between apoptosis, stem cells and cancer that has not been made in this way before: this pathway is critically important in stem cell death and in reducing tumour risk," Steller says.

"The work supports the idea that the stem cell is the seed of the tumour and that the transition from a normal stem cell to a cancer stem cell involves increased resistance to apoptosis."

ARTS interferes with molecules called inhibitor of apoptosis proteins (IAPs), which prevent cells from killing themselves. By inhibiting these inhibitors, under the right circumstances ARTS helps to take the brakes off the process of apoptosis, permitting the cell to die on schedule. Pharmaceutical companies are working to develop small molecule IAP antagonists, but this research is the first to show that inactivating a natural IAP antagonist actually causes tumours to grow, Steller says. It also suggests that the premature silencing of the Sept4/ARTS pathway at the stem cell level may herald cancer to come.

"This work not only defines the role of the ARTS gene in the underlying mechanism of mammalian tumour cell resistance to programmed cell death, but also links this gene to another hallmark of cancer, stem and progenitor cell proliferation," said Marion Zatz, who oversees cell death grants, including Steller's, at the NIH's National Institute of General Medical Sciences.

"The identification of the ARTS gene and its role in cancer cell death provides a potential target for new therapeutic approaches."

Source: Rockefeller University
Contact: Brett Norman

Reference:
Sept4/ARTS is required for stem cell apoptosis and tumor suppression
Maria Garcia-Fernandez, Holger Kissel, Samara Brown, Travis Gorenc, Andrew J. Schile, Shahin Rafii, Sarit Larisch, and Hermann Steller
Genes and Development, Oct 15, 2010; 24 (20)
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

Control of Foetal Heart-muscle-cell Proliferation

3-way control of foetal heart-cell proliferation could help regenerate cardiac cells
Saturday, 09 October 2010

Heart muscle cells do not normally replicate in adult tissue, but multiply with abandoned during development. This is why the loss of heart muscle after a heart attack is so dire — you can't grow enough new heart muscle to make up for the loss.

A team of researchers at the University of Pennsylvania School of Medicine describe the interconnections between three-molecules that control foetal, heart-muscle-cell proliferation in a mouse model that will help cardiologists better understand the natural repair process after heart attacks and help scientists learn how to expand cardiac stem cells for regenerative therapies.

The research team, led by Jonathan Epstein, MD, chair of the Department of Cell and Developmental Biology, and Chinmay Trivedi, MD, PhD, an Instructor in the same department, report their findings in the cover article of the most recent issue of Developmental Cell.

This is a thickened heart wall due to

loss of Hdac2-Hopx function.

Credit: Jon Epstein, MD, University

of Pennsylvania School of Medicine.

The Penn team showed that an enzyme called Hdac2 directly modifies a protein called Gata4, and a third protein called Hopx, which appears to have adopted a new function. Hopx is a member of a family of ancient, evolutionally conserved proteins that normally bind DNA. In this case, however, rather than binding to DNA, it works to bring two other proteins, Hdac2 and Gata4, together. By performing this unexpected matchmaker function, Hopx helps to control the rate at which heart muscle cells divide.

"Although the degree to which hearts can repair themselves after injury is controversial, if there is a natural regeneration process, even if normally insufficient and modest, then approaches leveraging this insight this could be useful for boosting new growth so that it has a clinically significant effect," says Epstein.

"We are eager to see if drugs like Hdac inhibitors will have this effect."

The scientists found an unexpected function for Hdac2 as well. This enzyme normally acts as a switch that regulates how DNA is packaged inside the cell, and therefore how large groups of genes are turned on and off. Epstein said that his team was surprised discover that in the developing heart this packaging role was not the critical function.

"Rather, Hdac2 seems to be working directly on other proteins, and not on DNA structure, to control replication of heart muscle cells," he says.

Hdac inhibitors are already in trials for cancer and one, valproic acid, has been used for decades to treat seizures. These inhibitors are a new class of agents that inhibit the proliferation of tumour cells in culture. Hdac inhibitors that are used to fight T cell lymphoma could possibly be used to enhance cardiac cell proliferation, say after a heart attack, when growing new heart muscle to replace damaged tissue would be is most needed.

"This could help to explain why Hdac inhibitors improve outcomes after heart attacks in animal models," says Trivedi.

Source: University of Pennsylvania School of Medicine
Contact: Karen Kreeger

Reference:
Hopx and Hdac2 Interact to Modulate Gata4 Acetylation and Embryonic Cardiac Myocyte Proliferation
Chinmay M. Trivedi, Wenting Zhu, Qiaohong Wang, Cheng Jia, Hae Jin Kee, Li Li, Sridhar Hannenhalli, Jonathan A. Epstein
Developmental Cell, Volume 19, Issue 3, 450-459, 14 September 2010, 10.1016/j.devcel.2010.08.012
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

Stem Cells Repair Damaged Spinal Cord Tissue

Stem Cells Repair Damaged Spinal Cord Tissue
Friday, 08 October 2010


Karolinska Institutet, Stockholm.

Researchers at the Swedish medical university Karolinska Institute have shown how stem cells, together with other cells, repair damaged tissue in the mouse spinal cord. The results are of potential significance to the development of therapies for spinal cord injury.

There is hope that damage to the spinal cord and brain will one day be treatable using stem cells (i.e. immature cells that can develop into different cell types). Stem cell-like cells have been found in most parts of the adult human nervous system, although it is still unclear how much they contribute to the formation of new, functioning cells in adult individuals.


Professor Jonas Frisén.

Credit: Camilla Svensk.

A joint study by Professor Jonas Frisén's research group at Karolinska Institute and their colleagues from France and Japan, and published in Cell Stem Cell, shows how stem cells and several other cell types contribute to the formation of new spinal cord cells in mice and how this changes dramatically after trauma. The research group has identified a type of stem cell, called an ependymal cell, in the spinal cord. They show that these cells are inactive in the healthy spinal cord, and that the cell formation that takes place does so mainly through the division of more mature cells. When the spinal cord is injured, however, these stem cells are activated to become the dominant source of new cells.

The stem cells then give rise to cells that form scar tissue and to a type of support cell that is an important component of spinal cord functionality. The scientists also show that a certain family of mature cells known as astrocytes produce large numbers of scar-forming cells after injury.

"The stem cells have a certain positive effect following injury, but not enough for spinal cord functionality to be restored," says Jonas Frisén.

"One interesting question now is whether pharmaceutical compounds can be identified to stimulate the cells to form more support cells in order to improve functional recovery after a spinal trauma."

Source: Karolinska Institutet
Contact: Katarina Sternudd

Reference:
Origin of new glial cells in the intact and injured adult spinal cord
Fanie Barnabé-Heider, Christian Göritz, Hanna Sabelström, Hirohide Takebayashi, Frank W. Pfrieger, Konstantinos Meletis & Jonas Frisén,
Cell Stem Cell, Volume 7, Issue 4, 470-482, 8 October 2010, 10.1016/j.stem.2010.07.014
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

Researcher Discovers Mechanism for Changing Adult Cells into Stem-like Cells

Scientist says 'It is fascinating to understand why the magic happens'
Friday, 08 October 2010

In 2006, Dr. Shinya Yamanaka of Kyoto University in Japan set the stem cell and regenerative medicine research world on fire when he successfully transformed differentiated mouse skin cells into cells that looked and behave like embryonic stem cells. Embryonic stem cells, the subject of much controversy when used in research, have the ability to differentiate into any type of tissue.

Yamanaka's creation of induced pluripotent stem cells [iPSCs] meant that in the future, research to improve human disease might be able to use iPSCs in lieu of embryonic stem cells. Since then, researchers around the world have been able to replicate his process. However, no one has been able to unlock the mechanism that allows cells to be regressed from differentiated to undifferentiated cells — until now.

University of Colorado Cancer Center researcher Chuan-Yuan Li, PhD, and his group have discovered that so-called "grim-reaper" caspase genes are the gatekeepers that can open the door to allow differentiated adult cells to regress to undifferentiated iPSCs.

"By doing experiments in which we added caspase inhibitor genes to the Yamanaka protocol, we discovered that when caspases are turned off, you cannot make iPSCs," says Li, professor of radiation oncology at the University of Colorado School of Medicine.

"We were able to shut down the process almost completely."

The discovery is the cover article in the Oct. 8, 2010 issue of Cell Stem Cell.

"For practical reasons, the discovery is important because even though the transformation to iPSCs is a straightforward process on surface, it is not very efficient, and this information can help increase efficiency," Li says.

"It can also help with the problem of cells that don't complete the transformation process acting like cancer cells. And from a purely scientific perspective, it is fascinating to understand why the magic happens."

Li's group had been working on the roles of caspases in wound healing when Yamanaka published his initial iPSC work in mice. That got Li thinking about potential roles of caspases in iPSC generation.

"I thought maybe caspases could also induce iPS cells instead of the four transcriptional factors that Yamanaka used," he says.

"If that was true, it would be very exciting."

For six months, his group tried different experiments using various caspase genes to coax human skin cells into iPS cells, but they had no success. Although caspases were not sufficient to make iPS cells, Li kept going with the idea that caspases were somehow involved.

They made their discovery when they introduced the caspase inhibitors into skin cells, which almost completely shut down the induction of iPS cells.

Caspases, Li says, appear to loosen up the built-in controls that make a cell differentiated or undifferentiated, just like a clutch allow a driver to switch gears while driving. Undifferentiated stem-like cells and differentiated cells from one person have the exact same genes. The difference between them is which genes are turned on or off.

In other words, he says, caspases could be the key to a kind of cellular reincarnation — taking a cell that, during human development, became a skin cell back to its original state to become any kind of cell.

"About twenty years ago, a scientist who was among the first to clone the caspase 3 gene named the gene Yama, the Hindu Lord of Death who was responsible for both killing a being and setting him on his way into his reincarnated life," Li said.

"It is now becoming clear that caspases don't just kill, but they can change the cell's fate. They could be a mediator of epigenetic changes in multi-cellular organisms."

Source: University of Colorado, Denver
Contact: David Kelly

Reference:
Apoptotic Caspases Regulate Induction of iPSCs from Human Fibroblasts
Fang Li, Zhimin He, Jingping Shen, Qian Huang, Wenrong Li, Xinjian Liu, Yujun He, Frank Wolf, Chuan-Yuan Li
Cell Stem Cell, Volume 7, Issue 4, 508-520, 8 October 2010, 10.1016/j.stem.2010.09.003
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

Stem Cells Shape Up to Their Surroundings

Stem Cells Shape Up to Their Surroundings
Friday, 08 October 2010

Many scientists aspire to take control over the stem cell differentiation process, so that we can grow organs and implants perfectly matched to each patient in the future. Now research in the Journal of Tissue Engineering, published by SAGE-Hindawi, explains how engineering the topography on which stem cells grow, and the mechanical forces working on them, can be as powerful an agent for change as their chemical environment.

Stem cells respond to the stiffness, chemistry and topography of the environments they find themselves in – and scientists building their understanding of the complex signalling controlling these responses hope to harness this knowledge to take stem cell research further. As well as increasing the potential to guide stem cells to create desired materials for research and clinical applications, using nanoscale topographies could eliminate (or alternatively enhance) steps including those involving feeder layers and synthetic induction supplements currently used in stem cell culture. In addition, tomorrow's increasingly sophisticated prosthetics for regenerative medicine could feature surfaces with varied tissue zones for different purposes, thanks to this improved understanding.

In their article, Laura McNamara of the University of Glasgow, UK, Centre for Cell Engineering, together with colleagues from Columbia University, New York, Nanotechnology Centre for Mechanics in Regenerative Medicine and the Bone and Joint Research Group at the University of Southampton, UK, review the latest developments in the use of nano-topography to direct stem cell differentiation. In particular, they look at skeletal (mesenchymal) stem cells.

Evidence is mounting that researcher’s can both maintain stem cells in the undifferentiated state, and determine the direction of their fate, by precise control of the surface features beneath them. Stem cells have an uncanny ability to detect and respond to nanoscale grooves, pits and ridges, and are particularly sensitive to the spacing and regularity of these features.

Nano-topographical responsiveness has been observed in diverse cell types including fibroblasts, osteoblasts, osteoclasts, endothelial, smooth muscle, epithelial, and epitenon cells.

"This is intriguing from a biomaterials perspective," says McNamara, "as it demonstrates that surface features of just a few nanometres can influence how cells will respond to, and form tissue on, materials."

Stem cells detect surface features with a variety of mechano-sensors, including integrin-linked focal adhesions. These respond to the mechanical constraints of the surface by inducing signalling cascades, such as the ERK-MAPK pathway. When the cell's rearranging cytoskeleton physically pulls on components of the cell's nucleus, this force works together with chemical signalling. Together these indirect (biochemical signal-mediated) and direct (force-mediated) factors can modulate nuclear components, altering gene expression to direct stem cell responses.

One interesting finding has been that topography can in some cases have the same effect as biochemical differentiation factors. The potential to eliminate the need for the latter opens the door to development of improved clinical prostheses with topographies that can directly modulate stem cell fate. In particular, the authors envisage applications involving engineered topography components for stem cells in regenerative medicine, for instance, in orthopaedics and dental implants. A combination of different topographies could be used to differentially functionalise implants for distinct applications, or demarcate particular "zones" within a single device.

Orthopaedic implants designed with specific regions tailored to integrate with bone and improve the chances of implant fixation might be seamlessly joining other areas of the implant programmed to reduce excessive bony in-growth, for example. Some surfaces with clinical potential include nano-structured titanium and diamond. A growing number of precision nanofabrication techniques are becoming available to help carve out the substrates needed for this research.

Skeletal stem cells have even been shown to grow into non-skeletal cells (known as transdifferentiation) on surfaces with the right groves and ridges – in some studies this has produced neural tissue.

"With the emergence of mechanical stimuli as critical modulators of cellular functionality, nano-topography should prove an excellent tool for development of novel biomaterials capable of promoting desirable cellular behaviour, discouraging unwanted cell responses, and preventing or ameliorating pathological changes," the authors suggest.

Source: SAGE Publications UK
Contact: Jayne Fairley

Reference:
Nanotopographical Control of Stem Cell Differentiation
Laura E. McNamara, Rebecca J. McMurray, Manus J. P. Biggs, Fahsai Kantawong, Richard O. C. Oreffo, and Matthew J. Dalby
Journal of Tissue Engineering, Vol. 2010 (2010), doi:10.4061/2010/120623
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

How to Create a Human Embryonic Stem Cell Line

U of Michigan creates the state’s first human ESC line
Sunday, 03 October 2010

University of Michigan researchers have created the state's first human embryonic stem cell line, achieving a long-sought goal that provides the foundation for future efforts to develop innovative disease treatments.

The new cell line, known as UM4-6, is the culmination of years of planning and preparation at U-M and was made possible by Michigan voters' November 2008 approval of a state constitutional amendment permitting scientists here to derive embryonic stem cell lines using surplus embryos from fertility clinics.

"This historic achievement opens the door on a new era for U-M researchers, one that holds enormous promise for the treatment of many seriously debilitating and life-threatening diseases," said U-M President Mary Sue Coleman.

"This accomplishment will enable the University of Michigan to take its place among the world's leaders in every aspect of stem cell research."

Work on UM4-6 began in May, and detailed characterization of the line was completed in late September. The project was conducted without federal funds, using private gifts to U-M's Consortium for Stem Cell Therapies and internal U-M resources. With the derivation of UM4-6, U-M joins a select group of fewer than a dozen U.S. universities that have created human embryonic stem cell lines.

"The real importance of today's announcement is that the ability to derive new embryonic stem cell lines will allow us to take the next step: disease-specific research that could someday lead to new treatments," said Gary Smith, leader of the derivation project at the U-M Consortium for Stem Cell Therapies.

The consortium will distribute UM4-6 samples to stem cell researchers across campus and to their collaborators statewide. In addition, U-M researchers hope — pending the resolution of a federal court case that seeks to bar federal funding for human embryonic stem cell research — to submit UM4-6 to the U.S. National Institutes of Health for inclusion in the national registry of human embryonic stem cell lines that are eligible for federal research funding.

"We've spent a lot of time making sure that our entire process is in compliance with the NIH guidelines for registering embryonic stem cell lines so that other scientists will be able to use these lines to conduct NIH-funded research," said Smith, co-director of the consortium and a professor of obstetrics and gynaecology at the Medical School.



This microscope image (400x magnification) shows

the 5-day-old embryo, known at that stage as a

blastocyst, used to create U-M’s first human

embryonic stem cell line. Credit: Image courtesy

of Gary Smith.

 UM4-6 was derived from a cluster of about 30 cells removed from a donated five-day-old embryo roughly the size of the period at the end of this sentence. That embryo was created for reproductive purposes but was no longer needed for that purpose and was therefore about to be destroyed.

The embryonic stem cells were extracted and placed in a culture dish containing nutrients that nourished them while preventing them from differentiating into specialized cell types. The cells divided and spread over the surface of the dish. When they began to crowd the dish, the cells were gently removed and placed into several fresh culture dishes, a process called re-plating. The re-plating process was repeated every seven to 10 days.

This microscope image (400x magnification)

shows an oval cluster of roughly 1,000

human embryonic stem cells growing together

as a colony. The colony is part of the UM4-6

line. Credit: Image courtesy of Gary Smith.

Once cell colonies have been successfully re-plated many times over several months, a new embryonic stem cell line — a collection of millions of genetically identical cells generated from a single embryo — has been established. Various tests are then performed to confirm that the cells display all the traits of normal embryonic stem cells, including the ability to form the body's specialized cell types. Conducting those tests is called characterizing an embryonic stem cell line.

While the creation of Michigan's first human embryonic stem cell line stands as a research milestone, the many steps that led to the achievement were nearly as important.

"We have addressed all the regulatory issues and have derived this line according to the highest ethical standards. We have a laboratory equipped with cutting-edge equipment and people with the know-how," Smith said.

“All our efforts have finally started to bear fruit, so now the truly exciting and novel work can begin."


Gary Smith, co-director of the U-M Consortium

for Stem Cell Therapies, observing the growth

of the UM4-6 human embryonic stem cell line

at the consortium's derivation laboratory. The

oval object in the centre of the computer screen

is a colony containing roughly 1,000 human

embryonic stem cells. Credit: Photo by Scott

Soderberg/U-M Photo Services.

 In March 2009, four months after voters approved the state constitutional amendment, U-M announced the creation of a consortium to establish new human embryonic stem cell lines that will aid in the search for disease treatments and cures.

Based at the Medical School, the A. Alfred Taubman Medical Research Institute's Consortium for Stem Cell Therapies secured funding commitments of nearly $2 million to start the program. Lab space totalling 1,254 square feet was obtained in the Biomedical Science Research Building, and the labs were outfitted with state-of-the-art equipment. Four new research associates were hired to do the work.

Throughout much of last year, U-M researchers worked to ensure that their proposed embryo-donation and cell-line derivation projects would comply with federal law and the Michigan Constitution, as well as extensive new regulations established last summer by the National Institutes of Health.

The project required approval by U-M's Human Pluripotent Stem Cell Research Oversight Committee and the Medical School's Institutional Review Board. Both committees are composed of physicians, scientists, ethicists, attorneys and community members who evaluated whether the project would be conducted ethically, legally and to the benefit of patients.

In November 2009, the U-M Consortium for Stem Cell Therapies received final approval to begin accepting donated embryos created for reproductive purposes but no longer needed or unsuitable for clinical use. Many Michigan couples, as well as some from outside the state, have contacted the university and expressed the desire to donate their surplus embryos — which would otherwise be discarded — for human embryonic stem cell research.

In March, the first attempts were made to establish an embryonic stem cell line, a process called derivation. After several attempts, the consortium team succeeded with UM4-6, using a 5-day-old embryo known at that stage as a blastocyst.

In addition to deriving new embryonic stem cell lines, consortium researchers spent much of this year refining recently developed techniques to convert adult skin cells into induced pluripotent stem cells, known as iPS cells. These reprogrammed cells display many of the most scientifically valuable properties of embryonic stem cells while enabling researchers to bypass embryos altogether.

Earlier this year, the consortium established its first iPS cells, using skin samples donated by healthy individuals and by patients with diseases including amyotrophic lateral sclerosis (ALS, also known as Lou Gehrig's disease) and several ataxias, said Sue O'Shea, a professor of cell and developmental biology at the Medical School and co-director of the Consortium for Stem Cell Therapies. Consortium workers are now attempting to grow iPS cell lines that will be used to study disease mechanisms.

"There are few university programs in the United States deriving disease-affected embryonic stem cell lines," O'Shea said.

"Our special niche will be creating, studying and understanding normal and abnormal development of disease-affected stem cell lines — both embryonic and iPS cell lines."

"These efforts represent the culmination of several years of work to bring Michigan laws into line with the laws of other states and expand the University of Michigan's facilities for pluripotent stem cell research, so we can follow the science wherever it leads and get to new treatments sooner rather than later," said Sean Morrison, director of the Center for Stem Cell Biology at U-M's Life Sciences Institute.

In the months and years ahead, consortium researchers will use genetically abnormal embryos to create cell lines that carry genes for diseases such as cystic fibrosis, Huntington's disease, Rett syndrome, spinal muscular atrophy and Tay-Sachs disease.

"We are extremely gratified that we are now able to make new embryonic stem cell and iPS cell lines available to researchers everywhere, who will put them to use in the discovery of effective treatments for a wide variety of human diseases," said Dr. Eva L. Feldman, director of the Taubman Institute.

"It demonstrates the wisdom of the voters of the state of Michigan, who put their faith and confidence in the work of their scientific community. It is also a tribute to Alfred Taubman, who donated his time, his money and his leadership to make this day possible."

Embryonic stem cells are the body's master cells; they can replicate endlessly and form all of the more than 200 cell types in the human body. Scientists hope these remarkably versatile cells — and the iPS cells that mimic them — can someday replace faulty cells or diseased tissues in failing organs. This fledgling field is known as regenerative medicine, and the U-M Consortium for Stem Cell Therapies intends to play a leadership role in this research.

Source: University of Michigan News
Contact: Jim Erickson
.........

ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---

Researchers Engineer Adult Stem Cells That Do Not Age

Bone marrow stem cells injected into skeletal muscle reverse heart failure
Sunday, 03 October 2010

Biomedical researchers at the University at Buffalo have engineered adult stem cells that scientists can grow continuously in culture, a discovery that could speed development of cost-effective treatments for diseases including heart disease, diabetes, immune disorders and neurodegenerative diseases.

UB scientists created the new cell lines – named "MSC Universal" – by genetically altering mesenchymal stem cells, which are found in bone marrow and can differentiate into cell types including bone, cartilage, muscle, fat, and beta-pancreatic islet cells. They identified a growth-factor chain of action that prompts bone marrow stem cells to repair cardiac tissue and reverse heart failure.

Earlier research from this group showed for the first time that injecting mesenchymal (bone marrow) stem cells into skeletal muscle in an animal model increased two-fold the production of myocytes, a type of heart muscle cell.


Biomedical researchers at the University

at Buffalo have engineered adult stem cells

that scientists can grow continuously in

culture. Credit: Douglas Levere,

University at Buffalo, NY.

The current findings provide insight into how the injected stem cells may rejuvenate the host tissue.

"By thoroughly understanding the interplay of stem cells and host tissue, and characterizing stem-cell-derived growth factors, it is possible to assemble a cocktail of these factors and use it for tissue repair, much like the use of insulin for diabetes patients," says Techung Lee, PhD, senior author.

Lee is associate professor of biochemistry and biomedical engineering in the UB School of Medicine and Biomedical Sciences and the School of Engineering and Applied Sciences, respectively.

Bone marrow mesenchymal stem cells [MSCs] possess an impressive ability to produce a plethora of growth factors, most of which remain to be characterized, Lee says.

"These growth factors appear to account for most of the observed therapeutic benefits in preclinical and clinical studies. Using skeletal muscle as a depot for the injected MSCs, we found that the MSC-derived growth factors activate production of host muscle tissue-derived growth factors."

The researchers say the breakthrough overcomes a frustrating barrier to progress in the field of regenerative medicine: the difficulty of growing adult stem cells for clinical applications.

Because mesenchymal stem cells have a limited life span in laboratory cultures, scientists and doctors who use the cells in research and treatments must continuously obtain fresh samples from bone marrow donors, a process both expensive and time-consuming. In addition, mesenchymal stem cells from different donors can vary in performance.

The cells that UB researchers modified show no signs of aging in culture, but otherwise appear to function as regular mesenchymal stem cells do – including by conferring therapeutic benefits in an animal study of heart disease. Despite their propensity to proliferate in the laboratory, MSC-Universal cells did not form tumours in animal testing.

Lee notes that current clinical trials of myocardial stem cell therapy require surgery, injecting the cells directly into the heart or into the heart muscle, invasive methods that can result in harmful scar tissue, arrhythmia, calcification or small vessel blockages. Lee's research group found that only 1-to-2 percent of MSCs infused into the myocardium actually grafted into the heart, and there was no evidence that they differentiated into heart muscle cells.

"For these reasons, and because patients with heart failure are not good surgical risks, it made sense to explore a non-invasive cell delivery approach," Lee notes.

"Our stem cell research is application-driven," says Techung Lee.

"If you want to make stem cell therapies feasible, affordable and reproducible, we know you have to overcome a few hurdles. Part of the problem in our health care industry is that you have a treatment, but it often costs too much. In the case of stem cell treatments, isolating stem cells is very expensive. The cells we have engineered grow continuously in the laboratory, which brings down the price of treatments."

Stem cells help regenerate or repair damaged tissues, primarily by releasing growth factors that encourage existing cells in the human body to function and grow.

Lee's group has shown that the instructive signal that generates the repair of cardiac tissue appears to come from at least a group of MSC-derived factors belonging to the IL-6 type cytokine family. Cytokines are small proteins made by the cells that act on other cells to stimulate or inhibit their function.

"These IL-6 type cytokines typically activate their cell/tissue targets through two specific proteins, known as JAK and STAT3, a cytosolic and a nuclear protein, respectively," explains Lee.

"These cytokines then instruct the host cell to produce another panel of growth factors.”

"The combined effects of the growth factors from injected stem cells and growth factors produced by host tissues cause tissue repair and achieve healing. Being able to use the factors for therapy rather than stem cells will make therapy to repair hearts much easier," he says.

UB has applied for a patent to protect Lee's discovery, and the university's Office of Science, Technology Transfer and Economic Outreach (UB STOR) is discussing potential license agreements with companies interested in commercializing MSC-Universal.

Lee's ongoing work indicates that this feature makes it feasible to repair tissue damage by injecting mesenchymal stem cells into skeletal muscle, a less invasive procedure than injecting the cells directly into an organ requiring repair. In a rodent model of heart failure, Lee and collaborators showed that intramuscular delivery of mesenchymal stem cells improved heart chamber function and reduced scar tissue formation.

UB STOR commercialization manager Michael Fowler believes MSC-Universal could be key to bringing new regenerative therapies to the market. The modified cells could provide health care professionals and pharmaceutical companies with an unlimited supply of stem cells for therapeutic purposes, Fowler says.

Lee says his research team has generated two lines of MSC-Universal cells: a human line and a porcine line. Using the engineering technique he and colleagues developed, scientists can generate an MSC-Universal line from any donor sample of mesenchymal stem cells, he says.

"I imagine that if these cells become routinely used in the future, one can generate a line from each ethnic group for each gender for people to choose from," Lee says.

Source: Adapted from Press releases from University at Buffalo, State University of New York.
Contact: Charlotte Hsu and Lois Baker.

Reference:
Activation of host tissue trophic factors through JAK/STAT3 signaling: A mechanism of mesenchymal stem cell-mediated cardiac repair
Shabbir A, Zisa D, Lin H, Mastri M, Roloff G, Suzuki G, Lee T
Am J Physiol Heart Circ Physiol. 2010 Sep 17, doi:10.1152/ajpheart.00488.2010
.........


ZenMaster

---

For more on stem cells and cloning, go to CellNEWS at
http://cellnews-blog.blogspot.com/

---