Major Step Forward In Cell Reprogramming, Researchers Report

From ScienceDaily.com

A team of Harvard Stem Cell Institute (HSCI) researchers has made a major advance toward producing induced pluripotent stem cells, or iPS cells, that are safe enough to use in treating diseases in patients.

“This demonstrates that we’re halfway home, and remarkably we got halfway home with just one chemical,” said Kevin Eggan, an HSCI principal faculty member who is the senior author of the paper being published online today by the journal Cell Stem Cell.

“There are four genes that do this, and with just one chemical we replaced half the genes,” said Eggan, who is also an assistant professor in Harvard’s Department of Stem Cell and Regenerative Biology. “The one chemical replaces those two genes in different ways at different times in the experiment. The experiments we performed not only led to discovery of the chemical, but they also explained how it works,” he said.

The chemical that the team used is a small molecule that members named RepSox in honor of another Boston team. It replaces Sox2 and cMyc, two of the four genes currently being used to reprogram adult skin cells into an embryonic-like state. Because cMyc is a tumor promoter and iPS cells created using it could never be used to treat patients, researchers have been looking for ways to turn back the cellular clock without the use of genes.

Lee Rubin, director of translational medicine at HSCI and the other senior author on the research team, said that “our goals were to try to as discretely and specifically as possible guide the cells through the deprogramming process” from the adult state to the embryonic-like state.

Finding a way to produce safe iPS cells that are the biological equivalent of embryonic stem cells is especially important because the cells can then be created from the cells of individual patients for transplantation into those patients. Thus, a patient with Parkinson’s disease might be treated with neurons created from his own cells, theoretically eliminating the need for immunosuppressive drugs, or the possibility of rejection of the transplanted cells. Similarly, patient-specific iPS cells could be used to create muscle for damaged hearts, or other individualized treatments.

Additionally, iPS cells derived from the skin cells of patients with specific diseases can be used as a source of differentiated cells to study those disease processes in a laboratory dish, and manipulated to find better drug targets and more effective therapeutics.

“This discovery is exciting because it demonstrates the feasibility of using chemicals to make safer patient-specific stem cells for transplantation medicine,” said Justin K. Ichida, a postdoctoral fellow in Eggan’s lab and the first author on the study. “One of the most important things we learned from this study is that, with respect to molecular pathways, there may be several ways to convert one type of cell into another. By using a nonbiased chemical screening approach, we uncovered a previously unknown way to make stem cells. The big challenge over the next decade will be to figure out how to make the right cells for disease treatment. This approach will be important for achieving that goal.”

Other co-first authors on the study are Joel Blanchard, Kelvin Lam, and Esther Y. Son. Additional contributors include Julia E. Chung, Dieter Egli, Kyle M. Loh, Ava C. Carter, Francesco P. Di Gorgio, Kathryn Koszak, Danwei Huangfu, Hidenori Akutsu, and David R. Liu.

The study was funded in part by the Harvard Stem Cell Institute, the Stowers Medical Institute, the Howard Hughes Medical Institute, and the New York Stem Cell Foundation.

‘Liposuction Leftovers’ Easily Converted To Induced Pluripotent Stem Cells

From ScienceDaily.com

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.

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Researchers May Have Found Equivalent of Embryonic Stem Cells

By Rob Stein, WashingtonPost.com

Chinese scientists have bred mice from cells that might offer an alternative to human embryonic stem cells, producing the most definitive evidence yet that the technique could help sidestep many of the explosive ethical issues engulfing the controversial field but raising alarm that the advance could lead to human cloning and designer babies.

In papers published online Thursday by two scientific journals, separate teams of researchers from Beijing and Shanghai reported that they had for the first time created virtual genetic duplicates of mice using skin cells from adult animals that had been coaxed into the equivalent of embryonic stem cells.

The findings were welcomed by supporters and opponents of human embryonic stem cell research as a long-sought vital step in proving that the cells could be as useful as embryonic cells for studying and curing many illnesses.

The results come just as the Obama administration has eased federal restrictions on government funding for embryonic stem cell research, and they could influence how to prioritize millions of dollars in new spending in the field.

But because of concerns that the techniques might make cloning and genetic engineering of embryos easier, the work could reignite calls for a ban on attempts to clone people and for restrictions on genetic manipulation of embryos.

“The implications of this are both enormously important and troublesome,” said Robert Lanza, a stem cell researcher at Advanced Cell Technology in Worcester, Mass. “It revives many of the issues raised by reproductive cloning.”

Many scientists believe human embryonic stem cell research could revolutionize medicine by enabling doctors to use genetically matched tissue to treat many diseases. But the field has been mired in controversy because embryos are destroyed to obtain the cells.

In 2006, scientists discovered that they could induce adult cells to regress to a stage that appeared identical to embryonic stem cells, called induced pluripotent stem (iPS) cells. Although scientists have become increasingly adept at creating and manipulating such cells, questions have lingered about whether they are truly equivalent. The new experiments were designed to put the cells to what has long been considered the most rigorous test.

In the studies, published in the journals Nature and Cell Stem Cell, the researchers used viruses to flip genetic switches in the DNA of skin cells from adult mice to turn them into iPS cells in the laboratory. The researchers then injected some of the iPS cells into very early embryos that are capable of forming a placenta but not of fully developing on their own. The resulting embryos were then transferred into the wombs of surrogate mice.

One team of scientists led by Qi Zhou of the Chinese Academy of Sciences created 37 iPS cell lines, three of which produced 27 live offspring, the first of which they named Tiny. One of the offspring, a 7-week-old male, went on to impregnate a female and produce young of its own. Altogether, the researchers bred at least 100 first-generation mice and hundreds of second-generation mice that were nearly identical genetically to the mice from which the iPS cells were derived.

“This gives us hope for future therapeutic interventions using patients’ own reprogrammed cells,” Fanyi Zeng of Shanghai Jiao Tong University, who worked with Zhou, said during a telephone briefing for reporters.

The second group of researchers, led by Shaorong Gao of the National Institute of Biological Sciences in Beijing, created five iPS cell lines, one of which was able to produce embryos that survived until birth. Although four animals were born, only one lived to adulthood. Nevertheless, the work is “proof that iPS cells are functionally equivalent to embryonic stem cells,” Gao said in a telephone interview.

Other researchers agreed, praising the work as a long-awaited confirmation of the cells’ equivalence.

“This clearly says for the first time that iPS cells pass the most stringent test,” said Konrad Hochedlinger, a stem cell researcher at Harvard University.

Opponents of human embryonic stem cell research said the findings provide the latest in a growing body of evidence for why such research is no longer necessary.

“Nobody has been able to find anything that embryonic stem cells can do that these cells can’t do,” said Richard M. Doerflinger of the U.S. Conference of Catholic Bishops. “This was the last remaining barrier.”

The Chinese scientists and others, however, said continued research on embryonic stem cells remains crucial to validate iPS cells and because it remains unclear which cells will turn out to be most useful for different purposes.

But the cells’ ability to produce almost genetically identical offspring raised the fear that rogue scientists might misuse the technique to attempt to clone humans.

“The culture wars are not over,” said Jonathan D. Moreno, a University of Pennsylvania bioethicist. “There was a lot of celebration about the end of the ethical issues with induced pluripotent stem cells. But this is the paradigm case that shows that the old debates are rapidly being transformed into something even more complicated.”

Lanza also raised the prospect that the techniques could one day be used essentially to steal someone’s DNA to make a baby. “With just a little piece of your skin, or some blood from the hospital, anyone could have your child — even an ex-girlfriend or neighbor,” he wrote in an e-mail. “This isn’t rocket science — with a little practice, any IVF clinic in the world could probably figure out how to get it to work.”

In addition, researchers could genetically engineer traits into the cells before using them to create embryos for designer babies.

“For instance, the technology already exists to genetically increase the muscle mass in animals by knocking out a gene known as mystatin, and could be used by a couple who wants a great child athlete,” Lanza wrote.

Others dismissed such concerns, saying many scientific, ethical and regulatory hurdles remain. They said that just because the process works in mice does not necessarily mean it would work in humans, that many states outlaw human cloning and that federal regulators could step in to prevent it.

Scientists Find Differences in Embryonic Stem Cells and Reprogrammed Skin Cells

From Newswise.com

UCLA researchers have found that embryonic stem cells and skin cells reprogrammed into embryonic-like cells have inherent molecular differences, demonstrating for the first time that the two cell types are clearly distinguishable from one another.

The data from the study suggest that embryonic stem cells and the reprogrammed cells, known as induced pluripotent stem (iPS) cells, have overlapping but still distinct gene expression signatures. The differing signatures were evident regardless of where the cell lines were generated, the methods by which they were derived or the species from which they were isolated, said Bill Lowry, a researcher with the Broad Stem Cell Research Center and a study author.

“We need to keep in mind that iPS cells are not perfectly similar to embryonic stem cells,” said Lowry, an assistant professor of molecular, cell and developmental biology. “We’re not sure what this means with regard to the biology of pluripotent stem cells. At this point our analyses comprise just an observation. It could be biologically irrelevant, or it could be manifested as an advantage or a disadvantage.”

The study appears in the July 2, 2009 issue of the journal Cell Stem Cell.

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Converting Adult Somatic Cells To Pluripotent Stem Cells Using A Single Virus

From ScienceDaily.com

A Boston University School of Medicine-led research team has discovered a more efficient way to create induced Pluripotent Stem (iPS) cells, derived from mouse fibroblasts, by using a single virus vector instead of multiple viruses in the reprogramming process.

The result is a powerful laboratory tool and a significant step toward the application of embryonic stem cell-like cells for clinical purposes such as the regeneration of organs damaged by inherited or degenerative diseases, including emphysema, diabetes, inflammatory bowel disease, and Alzheimer’s Disease.

Prior research studies have required multiple retroviral vectors for reprogramming — steps that depended on four different viruses to transfer genes into the cells’ DNA – essentially a separate virus for each reprogramming gene (Oct4. Klf4, Sox2 and cMyc). Upon activation these genes convert the cells from their adult, differentiated status to what amounts to an embryonic-like state.

However, the high number of genomic integrations — 15 to 20 — that typically occurs when multiple viruses are used for reprogramming, poses a safety risk in humans, as some of these genes (i.e. cMyc) can cause cancer. In addition, the viruses can integrate in cell locations turning on potential oncogenes.

The major milestone the six-member research team, led by Gustavo Mostoslavsky, Boston University Assistant Professor of Medicine in the Gastroenterology Section, achieved was combining the four vectors into a single “stem cell cassette” containing all four genes. The cassette (named STEMCCA) is comprised of a single multicistronic mRNA encoding the four transcription factors using a combination of 2A peptide technology and an internal ribosomal entry site (IRES).

With the STEMCCA vector, the researchers were able to generate iPS cells more efficiently — 10 times higher than previously reported studies.

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Scientists Create Monkey Stem Cells

(Ivanhoe Newswire) — The successful creation of the first induced pluripotent stem (iPS) cell line from adult monkey skin may have important implications for direct reprogramming capabilities across different species.

Previous studies have shown that induction of four key transcription factors can reprogram adult mouse and human skin cells into iPS cells. Until now, iPS cell creation had not been demonstrated in any other species.

Researchers at Peking University in Beijing, China retrofit viruses to express the four key factors to infect adult monkey skin cells. This technique led to the creation of cells that displayed multiple hallmarks of embryonic stem cells. These cells possessed the ability to develop into multiple types of differentiated cells.

These findings could potentially be useful for the creation of clinically valuable primate models for human disease. The direct reprogramming model may also be a universal strategy for generating iPS cells in other species.

SOURCE: Cell Stem Cell, 2008;3:587-590

Patient-derived induced stem cells retain disease traits

From Genengnews.com

When neurons started dying in Clive Svendsen’s lab dishes, he couldn’t have been more pleased.

The dying cells the same type lost in patients with the devastating neurological disease spinal muscular atrophy confirmed that the University of Wisconsin-Madison stem cell biologist had recreated the hallmarks of a genetic disorder in the lab, using stem cells derived from a patient. By allowing scientists the unparalleled opportunity to watch the course of a disease unfold in a lab dish, the work marks an enormous step forward in being able to study and develop new therapies for genetic diseases.

As reported this week in the journal Nature, Svendsen and colleagues at UW-Madison and the University of Missouri-Columbia created disease-specific stem cells by genetically reprogramming skin cells from a patient with spinal muscular atrophy, or SMA. In this inherited disease, the most common genetic cause of infant mortality, a mutation leads to the death of the nerves that control skeletal muscles, causing muscle weakness, paralysis, and ultimately death, usually by age two.

Genetic reprogramming of skin cells, first reported in late 2007 by UW-Madison stem cell biologists James Thomson and Junying Yu and a Japanese group led by Shinya Yamanaka, turns back the cells’ developmental clock and returns them to an embryonic-like state from which they can become any of the body’s 220 different cell types. The resulting induced pluripotent stem cells, known as iPS cells, harness the blank-slate developmental potential of embryonic stem cells without the embryo and have been heralded as a powerful potential way to study development and disease.

Just one year later, the new work is fulfilling that promise.

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Caption: The nerves that control muscles, known as motor neurons (shown here in red), are lost in the devastating genetic disease called spinal muscular atrophy, causing weakness, paralysis, and early death. A team of UW-Madison stem cell biologists recreated the hallmarks of this disease in the lab using genetically reprogrammed stem cells created from a young SMA patient’s skin. The work gives scientists the opportunity to study the full progression of a disease in the lab and should improve understanding and treatment of genetic disorders. The motor neurons shown here were grown from cells from the patient’s healthy mother.
Photo: provided by Clive Svendsen, cnsvendsen@wisc.edu