“Open” Stem Cell Chromosomes Reveal New Possibilities for Diabetes

Researchers map chromosomal changes that must take place before stem cells can be used to produce pancreatic and liver cells 

Thursday, 02 April 2015

These are pancreatic cells derived from

embryonic stem cells. Credit: UC San Diego

School of Medicine. 

Stem cells hold great promise for treating a number of diseases, in part because they have the unique ability to differentiate, specializing into any one of the hundreds of cell types that comprise the human body. Harnessing this potential, though, is difficult. In some cases, it takes up to seven carefully orchestrated steps of adding certain growth factors at specific times to coax stem cells into the desired cell type. Even then, cells of the intestine, liver and pancreas are notoriously difficult to produce from stem cells. Writing in Cell Stem Cell April 2, researchers at University of California, San Diego School of Medicine have discovered why.

It turns out that the chromosomes in laboratory stem cells open slowly over time, in the same sequence that occurs during embryonic development. It isn’t until certain chromosomal regions have acquired the “open” state that they are able to respond to added growth factors and become liver or pancreatic cells. This new understanding, say researchers, will help spur advancements in stem cell research and the development of new cell therapies for diseases of the liver and pancreas, such as type 1 diabetes.

“Our ability to generate liver and pancreatic cells from stem cells has fallen behind the advances we’ve made for other cell types,” said Maike Sander, MD, professor of paediatrics and cellular and molecular medicine and director of the Pediatric Diabetes Research Center at UC San Diego.

“So we haven’t yet been able to do things like test new drugs on stem cell-derived liver and pancreatic cells. What we have learned is that if we want to make specific cells from stem cells, we need ways to predict how those cells and their chromosomes will respond to the growth factors.”

Sander led the study, together with co-senior author Bing Ren, PhD, professor of cellular and molecular medicine at UC San Diego and Ludwig Cancer Research member.

Chromosomes are the structures formed by tightly wound and packed DNA. Humans have 46 chromosomes – 23 inherited from each parent. Sander, Ren and their teams first made maps of chromosomal modifications over time, as embryonic stem cells differentiated through several different developmental intermediates on their way to becoming pancreatic and liver cells. Then, in analysing these maps, they discovered links between the accessibility (openness) of certain regions of the chromosome and what they call developmental competence – the ability of the cell to respond to triggers like added growth factors.

“We’re also finding that these chromosomal regions that need to open before a stem cell can fully differentiate are linked to regions where there are variations in certain disease states,” Sander says.

In other words, if a person were to inherit a genetic variation in one of these chromosomal regions and his or her chromosome didn’t open up at exactly the right time, he or she could hypothetically be more susceptible to a disease affecting that cell type. Sander’s team is now working to further investigate what role, if any, these chromosomal regions and their variations play in diabetes.

Source: UCSD

Contact: Heather Buschman

Reference:

Epigenetic Priming of Enhancers Predicts Developmental Competence of hESC-Derived Endodermal Lineage Intermediates

Allen Wang, Feng Yue, Yan Li, Ruiyu Xie, Thomas Harper, Nisha A. Patel, Kayla Muth, Jeffrey Palmer, Yunjiang Qiu, Jinzhao Wang, Dieter K. Lam, Jeffrey C. Raum, Doris A. Stoffers, Bing Ren, Maike Sander

Cell Stem Cell, Volume 16, Issue 4, p386–399, 2 April 2015, DOI: http://dx.doi.org/10.1016/j.stem.2015.02.013

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Stem Cells Make Similar Decisions to Humans

Stem Cells Make Similar Decisions to Humans

Wednesday, 25 March 2015

Pancreas explant. 

Scientists at the University of Copenhagen have captured thousands of progenitor cells of the pancreas on video. They have filmed the cells making decisions to both divide and expand the organ or to specialize into the endocrine cells that regulate our blood sugar levels.

The new study reveals that stem cells behave as people in a society, making individual choices but with enough interactions to bring them to their end-goal. The results could eventually lead to a better control over the production of insulin-producing endocrine cells for diabetes therapy.

The research is published in the scientific journal PLOS Biology.

Why one cell matters

In a joint collaboration between the University of Copenhagen and University of Cambridge, Professor Anne Grapin- Botton and a team of researchers including Assistant Professor Yung Hae Kim from DanStem focused on marking the progenitor of the embryonic pancreas, commonly referred to as ‘mothers’, and their ‘daughters’ in different fluorescent colours and then captured them on video to analyse how they make decisions.

Prior to this work, there were methods to predict how specific types of pancreas cells would evolve as the embryo develops. However, by looking at individual cells, the scientists found that even within one group of cells presumed to be of the same type, some will divide many times to make the organ bigger while others will become specialized and will stop dividing.

The scientists witnessed interesting occurrences where the ‘mother’ of two ‘daughters’ made a decision and passed it on to the two ‘daughters’ who then acquired their specialization in synchrony. By observing enough cells, they were able to extract logic rules of decision-making, and with the help of Dr Pau Rué, a mathematician from the University of Cambridge, they developed a mathematical model to make long-term predictions over multiple generations of cells.

Stem cell movies

"It is the first time we have made movies of a quality that is high enough to follow thousands of individual cells in this organ, for periods of time that are long enough for us to follow the slow decision process. The task seemed daunting and technically challenging, but fascinating,” says Professor Grapin-Botton. 

"With these movies we can see and quantify the dynamics of decisions in each cell in the context of the organ, in a way that will inspire the study of many other organs," says Assistant Professor Yung Hae Kim.

"To complement the movies, which are done on isolated pancreas, we developed a method to visualize the family tree of cells in the untouched organ. We initially focused on one generation but now we are also observing their descendants over multiple generations," Kim elaborates.

Next steps in diabetes therapy

The project has been focused on basic research and is highly theoretical, but it now provides tools to control whether a cell should expand or specialize into an endocrine cell on its way to producing insulin.

“It is a worldwide quest to produce such insulin-producing cells from stem cells, for their transplantation in diabetic patients. In the future, this could be done by increasing the probability of specialization or by pushing ‘mother’ cells to pass on the decision to specialize to their two daughters”, Grapin-Botton concludes.

Source: University of Copenhagen

Contact: Yung Hae Kim

Reference:

Cell Cycle–Dependent Differentiation Dynamics Balances Growth and Endocrine Differentiation in the Pancreas

Yung Hae Kim , Hjalte List Larsen, Pau Rué, Laurence A. Lemaire, Jorge Ferrer and Anne Grapin-Botton 

PLOS Biology, March 18, 2015, DOI: 10.1371/journal.pbio.1002111

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New Cells May Help Treat Diabetes

U. Iowa group creates insulin-producing cells that normalize blood-sugar levels in diabetic mice

Thursday, 29 January 2015

University of Iowa researchers reprogrammed

human skin cells to create induced pluripotent

stem cells, which were then differentiated in a

stepwise fashion to create insulin-producing cells.

When these cells were transplanted into diabetic

mice, the cells secreted insulin and reduced the

blood sugar levels of the mice to normal or near-

normal levels. The image shows the insulin-

producing cells (right) and precursor cells (left).

Credit: University of Iowa.

Starting from human skin cells, researchers at the University of Iowa have created human insulin-producing cells that respond to glucose and correct blood-sugar levels in diabetic mice. The findings may represent a first step toward developing patient-specific cell replacement therapy for type 1 diabetes.

In the new study, published Jan. 28 in the journal PLOS ONE, the UI team led by Nicholas Zavazava, MD, PhD, UI professor of internal medicine, reprogrammed human skin cells to create induced pluripotent stem (iPS) cells, which were then coaxed into forming insulin-producing cells. When these cells were transplanted into diabetic mice, the cells secreted insulin and reduced the blood sugar levels of the mice to normal or near-normal levels.

Although the cells were not as effective as pancreatic cells in controlling blood sugar levels, Zavazava says that the results are an "encouraging first step" toward the goal of generating effective insulin-producing cells that can be used to potentially cure type 1 diabetes.

"This raises the possibility that we could treat patients with diabetes with their own cells," says Zavazava, who also is a member of the Fraternal Order of Eagles Diabetes Research Center at the UI.

"That would be a major advance, which will accelerate treatment of diabetes."

In type 1 diabetes, a person's immune system attacks and destroys the pancreatic beta cells that produce insulin. Although it is possible to treat type 1 diabetes with pancreas transplants from deceased donors, the demand for transplants far exceeds the availability of donated organs.

Zavazava's team is among several groups aiming to create an alternative source of insulin-producing pancreatic cells that can be transplanted into patients with type 1 diabetes. However, the UI study is the first to use human iPS cells to create the insulin-producing cells. Creating these cells from a patient's own cells would not only eliminate the need to wait for a donor pancreas, but would also mean patients could receive transplants without needing to take immunosuppressive drugs. Using iPS cells rather than embryonic stem cells as a starting point also avoids the ethical concerns some people have with using embryonic stem cells.

In the mouse study, the insulin-producing cells were placed under the kidney capsule - a thin membrane layer that surrounds the kidney - where they developed into an organ-like structure with its own blood supply. This new "organ" secreted insulin and gradually corrected the blood sugar levels in the diabetic mice over a period of several months. In addition, after the mice became normoglycemic, the glucose levels stayed steady.

By developing the cells in a stepwise fashion, the UI team was able to collect and use only those cells that would develop into pancreatic cells. This meant they were able to remove very immature (undifferentiated) cells that could form tumours. None of the mice developed tumours from the transplanted cells.

Source: University of Iowa

Contact: Jennifer Brown

Reference:

Human iPS Cell-Derived Insulin Producing Cells Form Vascularized Organoids under the Kidney Capsules of Diabetic Mice

Raikwar SP, Kim E-M, Sivitz WI, Allamargot C, Thedens DR and Nicholas Zavazava

PLoS ONE 10(1): e0116582 (2015), doi:10.1371/journal.pone.0116582

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Reprogramming Cells, Long Term

Positive finding could have major implications for diabetes treatment

Tuesday, 18 November 2014

Harvard Stem Cell Institute (HSCI) researchers, representing five Harvard departments and affiliated institutions as well as the Massachusetts Institute of Technology (MIT), have demonstrated that adult cells, reprogrammed into another cell type in a living animal, can remain functional over a long period.

Joe Zhou is a Harvard Stem Cell Institute

Principal Faculty Member and an Associate

Professor in Harvard's Department of Stem Cell

and Regenerative Biology. Zhou and colleagues

have demonstrated that cells reprogrammed in

vivo can survive, thrive, and be therapeutically

useful. Credit: B. D. Colen/Harvard University.

The work by Joe Zhou, an associate professor in Harvard's Department of Stem Cell and Regenerative Biology, and his collaborators is an important advance in the effort to develop cell-based therapies for tissue repair, and specifically in the effort to develop improved treatment for diabetes.

The researchers used a combination of genes to change pancreatic exocrine cells – one of the main forms of cells in the pancreas – in adult mice that have diabetes into insulin-producing beta cells that appeared to cure about a third of the mice of the metabolic disease, and improved insulin production in most of the other mice. A report on the work was published today in the journal Nature Biotechnology.

The new findings are a major advance in work by HSCI co-director Doug Melton and Zhou, who in 2008 reported having converted exocrine cells into functional beta cells in mice. At that time, however, it was not known how long, and how well, the repurposed cells would function.

"The efficiency of reprogramming has always been an issue," Zhou said.

"Until now, the new cells have either dropped dramatically in number or disappeared completely," he said, noting that since his work with Melton in 2008 there have been reports published in other programing systems that question whether the reprogrammed cells could be stable enough ultimately to be useful.

"What we have demonstrated is that yes, the reprogrammed cells can be useful, and for that to happen you have to create a niche environment in which the cells can survive," Zhou continued.

"We have improved the reprogramming efficiency to a point where one can create a large enough number of the new cells that the new cells create their own niche environment."

Zhou said that the researchers studied the mice for up to about 13 months, approximately half their normal life span, and found that "the cells are still there, and fairly robust. These are diabetic animals, and we were able to, I wouldn't use the word 'cure' because that's a very freighted word for me to use, but they became highly glycaemic animals – though not every animal became normal. That may be because to completely control the glucose level of the animal, you not only need beta cells, you need about a quarter of a million functional beta cells. If you are short of this number, even if the beta cells are perfectly normal," they can't completely control blood sugar levels, Zhou said.

When discussing the implications of the study for the field of cellular reprogramming, Zhou cautioned that the pancreas has a particularly simple cellular organization and structure, and thus findings in the pancreas might not necessarily apply to other organs.

Diabetes is a metabolic disease that is seen in two basic forms.

Type 1 diabetes is an autoimmune disease affecting about 3 million Americans, in which the patient's immune system ultimate destroys all the insulin-producing beta cells in the pancreas, and the patient has to inject insulin in order to regulate blood glucose levels.

Type 2 diabetes, which is now at epidemic prevalence rates in the United States and around the world, is usually caused by being overweight, lack of proper exercise, and improper diet, and can make a patient insulin-resistant, so the insulin the body produces is not sufficient to control blood glucose levels.

If the kind of treatment approach suggested by the new study were to succeed in humans – and that is a question to be answered with further animal, and eventually human, studies – it could be useful in treating both forms of diabetes.

One drawback to the current form of the new approach is that the cellular reprogramming is done with genes, and there might ultimately be unwanted effects on the cells. Zhou said the goal would be to replace the genes with chemicals or, perhaps, RNAs.

"I've talked to many clinicians about whether our approach could be used in humans," Zhou said.

"And the opinion is divided. Some say this could be developed into a human treatment, and some say it should be improved. But there seems to be general agreement that it could potentially be useful."

Source: Harvard University

Contact: B. D. Colen

Reference:

Long-term persistence and development of induced pancreatic beta cells generated by lineage conversion of acinar cells

Weida Li, Claudia Cavelti-Weder, Yinying Zhang, Kendell Clement, Scott Donovan, Gabriel Gonzalez, Jiang Zhu, Marianne Stemann, Ke Xu, Tatsu Hashimoto, Takatsugu Yamada, Mio Nakanishi, Yuemei Zhang, Samuel Zeng, David Gifford, Alexander Meissner, Gordon Weir & Qiao Zhou

Nature Biotechnology (2014), doi:10.1038/nbt.3082

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Stem Cell Success Raises Hopes of Type 1 Diabetes Cure

From human embryonic stem cells to billions of human insulin producing cells

Giant step toward new diabetes treatment

Friday, 10 October 2014

This is an image of human stem cell-derived beta

cells that have formed islet like clusters in a

mouse. Cells were transplanted to the kidney

capsule and photo was taken two weeks later by

which time the beta cells are making insulin and

have cured the diabetes in the mouse. Credit:

Douglas Melton. 

Harvard stem cell researchers today announced that they have made a giant leap forward in the quest to find a truly effective treatment for type 1 diabetes, a condition that affects an estimated three million Americans at a cost of about $15 billion annually: With human embryonic stem cells as a starting point, the scientists are for the first time able to produce, in the kind of massive quantities needed for cell transplantation and pharmaceutical purposes, human insulin-producing beta cells equivalent in most every way to normally functioning beta cells.

Doug Melton, who led the work and who twenty-three years ago, when his then infant son Sam was diagnosed with type 1 diabetes, dedicated his career to finding a cure for the disease, said he hopes to have human transplantation trials using the cells to be underway within a few years.

"We are now just one pre-clinical step away from the finish line," said Melton, whose daughter Emma also has type 1 diabetes.

A report on the new work has today been published by the journal Cell.

Felicia W. Pagliuca, Jeff Millman, and Mads Gurtler of Melton's lab are co-first authors on the Cell paper. The research group and paper authors include a Harvard undergraduate.

"You never know for sure that something like this is going to work until you've tested it numerous ways," said Melton, Harvard's Xander University Professor and a Howard Hughes Medical Institute Investigator.

"We've given these cells three separate challenges with glucose in mice and they've responded appropriately; that was really exciting.”

"It was gratifying to know that we could do something that we always thought was possible," he continued, "but many people felt it wouldn't work. If we had shown this was not possible, then I would have had to give up on this whole approach. Now I'm really energized."

The stem cell-derived beta cells are presently undergoing trials in animal models, including non-human primates, Melton said.

Elaine Fuchs, the Rebecca C. Lancefield Professor at Rockefeller University, and a Howard Hughes Medical Institute Investigator who is not involved in the work, hailed it as "one of the most important advances to date in the stem cell field, and I join the many people throughout the world in applauding my colleague for this remarkable achievement.”

"For decades, researchers have tried to generate human pancreatic beta cells that could be cultured and passaged long term under conditions where they produce insulin. Melton and his colleagues have now overcome this hurdle and opened the door for drug discovery and transplantation therapy in diabetes," Fuchs said.

And Jose Oberholtzer, M.D., Associate Professor of Surgery, Endocrinology and Diabetes, and Bioengineering at the University of Illinois at Chicago, and its Director of the Islet and Pancreas Transplant Program and the Chief of the Division of Transplantation, said work described in today's Cell "will leave a dent in the history of diabetes. Doug Melton has put in a life-time of hard work in finding a way of generating human islet cells in vitro. He made it. This is a phenomenal accomplishment."

Doug Melton, who when his then infant son was

diagnosed 23 years ago with type 1 diabetes

dedicated his career to finding a cure for the

disease, has successfully produced insulin

producing, glucose sensitive, human beta cells

from human embryonic stem cells. Credit: B. D.

Colen/Harvard University.

Melton, co-scientific director of the Harvard Stem Cell Institute, and the University's Department of Stem Cell and Regenerative Biology – both of which were created more than a decade after he began his quest – said that when he told his son and daughter they were surprisingly calm.

"I think like all kids, they always assumed that if I said I'd do this, I'd do it," he said with a self-deprecating grin.

Type 1 diabetes is an autoimmune metabolic condition in which the body kills off all the pancreatic beta cells that produce the insulin needed for glucose regulation in the body. Thus the final pre-clinical step in the development of a treatment involves protecting from immune system attack the approximately 150 million cells that would have to be transplanted into each patient being treated. Melton is collaborating on the development of an implantation device to protect the cells with Daniel G. Anderson, the Samuel A. Goldblith Professor of Applied Biology, Associate Professor in the Department of Chemical Engineering, the Institute of Medical Engineering and Science, and the Koch Institute at MIT.

Melton said that the device Anderson and his colleagues at MIT are currently testing has thus far protected beta cells implanted in mice from immune attack for many months.

"They are still producing insulin," Melton said.

Cell transplantation as a treatment for diabetes is still essentially experimental, uses cells from cadavers, requires the use of powerful immunosuppressive drugs, and has been available to only a very small number of patients.

MIT's Anderson said the new work by Melton's lab is "an incredibly important advance for diabetes. There is no question that ability to generate glucose-responsive, human beta cells through controlled differentiation of stem cells will accelerate the development of new therapeutics. In particular, this advance opens doors to an essentially limitless supply of tissue for diabetic patients awaiting cell therapy."

Richard A. Insel, M.D., chief scientific officer of the Juvenile Diabetes Research Foundation, a founder of Melton's work, said the "JDRF is thrilled with this advancement toward large scale production of mature, functional human beta cells by Dr. Melton and his team. This significant accomplishment has the potential to serve as a cell source for islet replacement in people with type 1 diabetes and may provide a resource for discovery of beta cell therapies that promote survival or regeneration of beta cells and development of screening biomarkers to monitor beta cell health and survival to guide therapeutic strategies for all stages of the disease."

Melton expressed gratitude to both the Juvenile Diabetes Research Foundation and the Helmsley Trust, saying "their support has been, and continues to be essential."

While diabetics can keep their glucose metabolism under general control by injecting insulin multiple times a day, that does not provide the kind of exquisite fine tuning necessary to properly control metabolism, and that lack of control leads to devastating complications from blindness to loss of limbs.

About 10 percent of the more than 26 million Americans living with type 2 diabetes are also dependent upon insulin injections, and would presumably be candidates for beta cell transplants, Melton said.

"There have been previous reports of other labs deriving beta cell types from stem cells, no other group has produced mature beta cells as suitable for use in patients," he said.

"The biggest hurdle has been to get to glucose sensing, insulin secreting beta cells, and that's what our group has done."

Source: Harvard University

Contact: B. D. Colen

Reference:

Generation of Functional Human Pancreatic β Cells In Vitro

Felicia W. Pagliuca, Jeffrey R. Millman, Mads Gürtler, Michael Segel, Alana Van Dervort, Jennifer Hyoje Ryu, Quinn P. Peterson, Dale Greiner, Douglas A. Melton

Cell, 9 October 2014, Volume 159, Issue 2, p428–439

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First Disease-specific Human Embryonic Stem Cell Line by Nuclear Transfer

Major step toward cell-based therapies for life-threatening diseases

Tuesday, 29 April 2014

Using somatic cell nuclear transfer, a team of scientists led by Dr. Dieter Egli at the New York Stem Cell Foundation (NYSCF) Research Institute and Dr. Mark Sauer at Columbia University Medical Center has created the first disease-specific embryonic stem cell line with two sets of chromosomes.

Embryonic stem cells from an adult with type 1

diabetes were created by transferring the nucleus

from a skin cell of the patient into a donor oocyte.

Credit: Bjarki Johannesson, NYSCF. 

As reported today in Nature, the scientists derived embryonic stem cells by adding the nuclei of adult skin cells to unfertilized donor oocytes using a process called somatic cell nuclear transfer (SCNT). Embryonic stem cells were created from one adult donor with type 1 diabetes and a healthy control. In 2011, the team reported creating the first embryonic cell line from human skin using nuclear transfer when they made stem cells and insulin-producing beta cells from patients with type 1 diabetes. However, those stem cells were triploid, meaning they had three sets of chromosomes, and therefore could not be used for new therapies.

The investigators overcame the final hurdle in making personalized stem cells that can be used to develop personalized cell therapies. They demonstrated the ability to make a patient-specific embryonic stem cell line that has two sets of chromosomes (a diploid state), the normal number in human cells. Reports from 2013 showed the ability to reprogram foetal fibroblasts using SCNT; however, this latest work demonstrates the first successful derivation by SCNT of diploid pluripotent stem cells from adult and neonatal somatic cells.

"From the start, the goal of this work has been to make patient-specific stem cells from an adult human subject with type 1 diabetes that can give rise to the cells lost in the disease," said Dr. Egli, the NYSCF scientist who led the research and conducted many of the experiments.

"By reprograming cells to a pluripotent state and making beta cells, we are now one step closer to being able to treat diabetic patients with their own insulin-producing cells."

"I am thrilled to say we have accomplished our goal of creating patient-specific stem cells from diabetic patients using somatic cell nuclear transfer," said Susan L. Solomon, CEO and co-founder of NYSCF.

"I became involved with medical research when my son was diagnosed with type 1 diabetes, and seeing today's results give me hope that we will one day have a cure for this debilitating disease. The NYSCF laboratory is one of the few places in the world that pursues all types of stem cell research. Even though many people questioned the necessity of continuing our SCNT work, we felt it was critical to advance all types of stem-cell research in pursuit of cures. We don't have a favourite cell type, and we don't yet know what kind of cell is going to be best for putting back into patients to treat their disease."

The research is the culmination of an effort begun in 2006 to make patient-specific embryonic stem cell lines from patients with type 1 diabetes. Ms. Solomon opened NYSCF's privately funded laboratory on March 1, 2006, to facilitate the creation of type 1 diabetes patient-specific embryonic stem cells using SCNT. Initially, the stem cell experiments were done at Harvard and the skin biopsies from type 1 diabetic patients at Columbia; however, isolation of the cell nuclei from these skin biopsies could not be conducted in the federally funded laboratories at Columbia, necessitating a safe-haven laboratory to complete the research. NYSCF initially established its lab, now the largest independent stem cell laboratory in the nation, to serve as the site for this research.

In 2008, all of the research was moved to the NYSCF laboratory when the Harvard scientists determined they could no longer move forward, as restrictions in Massachusetts prevented their obtaining oocytes. Dr. Egli left Harvard University and joined NYSCF; at the same time, NYSCF forged a collaboration with Dr. Sauer who designed a unique egg-donor program that allowed the scientists to obtain oocytes for the research.

"This project is a great example of how enormous strides can be achieved when investigators in basic science and clinical medicine collaborate. I feel fortunate to have been able to participate in this important project," said Dr. Sauer. Dr. Sauer is vice chair of the Department of Obstetrics and Gynecology, professor of obstetrics and gynaecology, and chief of reproductive endocrinology at Columbia University Medical Center and program director of assisted reproduction at the Center for Women's Reproductive Care.

Patients with type 1 diabetes lack insulin-producing beta cells, resulting in insulin deficiency and high blood-sugar levels. Therefore, producing beta cells from stem cells for transplantation holds promise as a treatment and potential cure for type 1 diabetes. Because the stem cells are made using a patient's own skin cells, the beta cells for replacement therapy would be autologous, or from the patient, matching the patient's DNA.

Generating autologous beta cells using SCNT is only the first step in developing a complete cell replacement therapy for type 1 diabetes. In type 1 diabetes, the body's immune system attacks its own beta cells; therefore, further work is underway at NYSCF, Columbia, and other institutions to develop strategies to protect existing and therapeutic beta cells from attack by the immune system, as well as to prevent such attack.

The technique described in the report published today can also be translated for use in the development of personalized autologous cell therapies for many other diseases and conditions including Parkinson's disease, macular degeneration, multiple sclerosis, and liver diseases and for replacing or repairing damaged bones.

As part of the work, the scientists systematically analysed the factors that affect stem-cell derivation after SCNT. The reprogramming of skin cells from a type 1 diabetes patient by SCNT has long been sought, but has been challenging to achieve because of logistical difficulties in obtaining human oocytes for research, as well as an incomplete understanding of the biology of human oocytes.

The scientists found that the addition of specific chemicals, called histone deacetylase inhibitors, and an efficient protocol for human oocyte activation were critical to achieving development to the stage at which embryonic stem cells are derived. These findings are consistent with the 2013 report by Tachibana and colleagues that used foetal cells. Though the authors of the 2013 paper also performed studies with cells of an infant with Leigh syndrome, they did not demonstrate that diploid pluripotent stem cells could be derived from these cells. Because foetal cells are less mature than the cells after birth, it was critical to determine if diploid pluripotent stem cells could be derived from the cells of both infants and adults.

As an additional optimization of the SCNT protocol, the scientists found that it was important to maintain the integrity of the plasma membrane during manipulation, and that to do so, the agent used in the manipulations had to be at a low dose. The scientists applied this optimized protocol to skin cells of a male new-born and the cells of the adult patient with type 1 diabetes. From these two cell lines, the scientists produced a total of four SCNT-derived embryonic stem cell lines. All cell lines were diploid and could give rise to neurons, pancreatic cells, and cartilage, as well as various other cell types, demonstrating their pluripotency. Importantly, the cells of the type 1 diabetes patient also gave rise to insulin-producing beta cells.

Therefore, this is the first report of the derivation of diploid pluripotent stem cells from a patient. And together with a paper published this month in Cell Stem Cell by Chung et al., it is also the first report of diploid embryonic stem cell lines derived from a human after birth.

Dr. Nissim Benvenisty and his laboratory at Hebrew University of Jerusalem collaborated on this report by demonstrating that the cells produced were, in fact, embryonic stem cells by using microarrays to perform gene expression analysis of the cells.

Dr. Rudolph Leibel, a co-author and co-director with Dr. Robin Goland of the Naomi Berrie Diabetes Center, where aspects of these studies were conducted, said:

"This accomplishment is the product of an ongoing inter-institutional collaboration across scientific and clinical disciplines, supported by thoughtful philanthropy. The resulting technical and scientific insights bring closer the promise of cell replacement for a wide range of human disease."

NYSCF continues pursuing SCNT research despite many scientific obstacles and in light of the advent of induced pluripotent stem (iPS) cells, as it is not yet clear which type of stem cells will prove best for personalized treatments. Many thought that iPS cells, first created from human cells in 2007, would replace the need for patient-specific embryonic stem cells because they allow patient- and disease-specific stem cell lines to be generated by genetically reprogramming adult cells into becoming pluripotent cells. However, it is not clear how similar iPS cells are to naturally occurring embryonic stem cells, which remain the gold standard, and what will be the preferred cell type for therapies.

Though it is now possible to derive stem cell lines with a patient's genotype using iPS technology, the generation of stem cells using oocytes may have an advantage for use in cell replacement for diseases such as type 1 diabetes. The generation of pluripotent stem cell lines by SCNT uses human oocytes, while iPS cells use recombinant DNA, RNA, or chemicals, each of which requires its own safety testing and approval for clinical use. Human oocytes are already used routinely around the world to generate clinically relevant cells. The generation of pluripotent stem cell lines using human oocytes may therefore be particularly suitable for the development of cell-replacement therapies. Therefore, this work brings the scientists a significant step closer to this goal.

Drs. Mitsutoshi Yamada and Bjarki Johannesson, postdoctoral fellows at the NYSCF Research Institute, were the co-first authors of the paper.

Source: New York Stem Cell Foundation

Contact: David McKeon

Reference:

Human oocytes reprogram adult somatic nuclei of a type 1 diabetic to diploid pluripotent stem cells

Mitsutoshi Yamada, Bjarki Johannesson, Ido Sagi, Lisa Cole Burnett, Daniel H. Kort, Robert W. Prosser, Daniel Paull, Michael W. Nestor, Matthew Freeby, Ellen Greenberg, Robin S. Goland, Rudolph L. Leibel, Susan L. Solomon, Nissim Benvenisty, Mark V. Sauer& Dieter Egli

Nature 28 April 2014, doi:10.1038/nature13287

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Replacing Insulin Through Stem Cell-derived Pancreatic Cells under the Skin

A newly created method of placing stem cell-derived pancreatic cells in capsules under the skin to replace insulin is tested in diabetic disease models

Tuesday, 25 March 2014

Microscopic view of Islet of Langerhans in the

pancreas. Beta cells in the Islets are responsible

for producing insulin. Credit: Courtesy of Sanford-

Burnham Medical Research Institute. 

Sanford-Burnham Medical Research Institute (Sanford-Burnham) and UC San Diego School of Medicine scientists have shown that by encapsulating immature pancreatic cells derived from human embryonic stem cells (hESC), and implanting them under the skin in animal models of diabetes, sufficient insulin is produced to maintain glucose levels without unwanted potential trade-offs of the technology. The research suggests that encapsulated hESC-derived insulin-producing cells hold great promise as an effective and safe cell-replacement therapy for insulin-dependent diabetes.

"Our study critically evaluates some of the potential pitfalls of using stem cells to treat insulin-dependent diabetes," said Pamela Itkin-Ansari, Ph.D., adjunct assistant professor in the Development, Aging, and Regenerative Program at Sanford-Burnham, with a joint appointment at UC San Diego.

"We have shown that encapsulated hESC-derived pancreatic cells are able to produce insulin in response to elevated glucose without an increase in the mass or their escape from the capsule. These results are important because it means that the encapsulated cells are both fully functional and retrievable," said Itkin-Ansari.

In the study, published online in Stem Cell Research, Itkin-Ansari and her team used bioluminescent imaging to see if encapsulated cells stay in the capsule after implantation.

Previous attempts to replace insulin-producing cells, called beta cells, have met with significant challenges. For example, researchers have tried treating diabetics with mature beta cells, but because mature cells are fragile and scarce, the method is fraught with problems. Moreover, since the cells come from organ donors, they may be recognized as foreign by the recipient's immune system — requiring patients to take immunosuppressive drugs to prevent their immune system from attacking the donor's cells, ultimately leaving patients vulnerable to infections, tumours, and other adverse events.

Encapsulation technology was developed to protect donor cells from exposure to the immune system — and has proven extremely successful in preclinical studies.

Itkin-Ansari and her research team previously made an important contribution to the encapsulation approach by showing that pancreatic islet progenitor cells are an optimal cell type for encapsulation. They found that progenitor cells were more robust than mature beta cells to encapsulate, and while encapsulated, they matured into insulin-producing cells, which secreted insulin only when needed.

"We were thrilled to see that the cells remained fully encapsulated for up to 150 days, the longest period tested,” said Itkin-Ansari.

"Equally important is that we show that the progenitor cells develop glucose responsiveness without a significant change in mass — meaning they don't outgrow their capsule.”

"Next steps for the development of the approach will be to figure out the size of the capsule required to house the number of progenitor beta cells needed to respond to glucose in humans. And of course we want to learn how long a capsule will function once implanted. Given these goals and continued successful results, I expect to see the technology become a treatment option for patients with insulin-dependent diabetes," said Itkin-Ansari.

Source: Sanford-Burnham Medical Research Institute 

Contact: Susan Gammon, Ph.D.

Reference:

Human embryonic stem cell derived islet progenitors mature inside an encapsulation device without evidence of increased biomass or cell escape

Kaitlyn Kirk, Ergeng Hao, Reyhaneh Lahmy, Pamela Itkin-Ansari

Stem Cell Research, online 24 March 2014, http://dx.doi.org/10.1016/j.scr.2014.03.003

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ZenMaster

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For more on stem cells and cloning, go to CellNEWS at

http://cellnews-blog.blogspot.com/

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Scientists Reprogram Skin Cells into Insulin-producing Pancreas Cells

Gladstone-led study represents important step towards a cure for type 1 diabetes

Friday, 07 February 2014

Gladstone Postdoctoral Scholar Ke Li, PhD 

(above) and Investigator Sheng Ding, MD, PhD,

have identified a new method of generating

functional, insulin-producing β-cells, representing

a potential therapeutic strategy for combating

type 1 diabetes. Credit: Chris Goodfellow.

A cure for type 1 diabetes has long eluded even the top experts. Not because they do not know what must be done — but because the tools did not exist to do it. But now scientists at the Gladstone Institute’s Investigator Sheng Ding, MD, PhD, harnessing the power of regenerative medicine, have developed a technique in animal models that could replenish the very cells destroyed by the disease. The team's findings, published online today in the journal Cell Stem Cell, are an important step towards freeing an entire generation of patients from the life-long injections that characterize this devastating disease.

Type 1 diabetes, which usually manifests during childhood, is caused by the destruction of b-cells, a type of cell that normally resides in the pancreas and produces a hormone called insulin. Without insulin, the body's organs have difficulty absorbing sugars, such as glucose, from the blood. Once a death sentence, the disease can now be managed with regular glucose monitoring and insulin injections. A more permanent solution, however, would be to replace the missing b-cells. But these cells are hard to come by, so researchers have looked towards stem cell technology as a way to make them.

"The power of regenerative medicine is that it can potentially provide an unlimited source of functional, insulin-producing b-cells that can then be transplanted into the patient," said Dr. Ding, who is also a professor at the University of California, San Francisco (UCSF), with which Gladstone is affiliated.

"But previous attempts to produce large quantities of healthy b-cells — and to develop a workable delivery system — have not been entirely successful. So we took a somewhat different approach."

One of the major challenges to generating large quantities of b-cells is that these cells have limited regenerative ability; once they mature it's difficult to make more. So the team decided to go one step backwards in the life cycle of the cell.

The team first collected skin cells, called fibroblasts, from laboratory mice. Then, by treating the fibroblasts with a unique 'cocktail' of molecules and reprogramming factors, they transformed the cells into endoderm-like cells. Endoderm cells are a type of cell found in the early embryo, and which eventually mature into the body's major organs — including the pancreas.

"Using another chemical cocktail, we then transformed these endoderm-like cells into cells that mimicked early pancreas-like cells, which we called PPLC's," said Gladstone Postdoctoral Scholar Ke Li, PhD, the paper's lead author.

"Our initial goal was to see whether we could coax these PPLC's to mature into cells that, like b-cells, respond to the correct chemical signals and — most importantly — secrete insulin. And our initial experiments, performed in a petri dish, revealed that they did."

The research team then wanted to see whether the same would occur in live animal models. So they transplanted PPLC's into mice modified to have hyperglycaemia (high glucose levels), a key indicator of diabetes.

"Importantly, just one week post-transplant, the animals' glucose levels started to decrease gradually approaching normal levels," continued Dr. Li.

"And when we removed the transplanted cells, we saw an immediate glucose spike, revealing a direct link between the transplantation of the PPLC's and reduced hyperglycaemia."

But it was when the team tested the mice eight weeks post-transplant that they saw more dramatic changes: the PPLC's had given rise to fully functional, insulin-secreting b-cells.

"These results not only highlight the power of small molecules in cellular reprogramming, they are proof-of-principle that could one day be used as a personalized therapeutic approach in patients," explained Dr. Ding.

"I am particularly excited about the prospect of translating these findings to the human system," said Matthias Hebrok, PhD, one of the study's authors and director of the UCSF Diabetes Center.

"Most immediately, this technology in human cells could significantly advance our understanding of how inherent defects in b-cells result in diabetes, bringing us notably closer to a much-needed cure."

Source: Gladstone Institutes 

Contact: Anne Holden

Reference:

Small Molecules Facilitate the Reprogramming of Mouse Fibroblasts into Pancreatic Lineages

Ke Li, Saiyong Zhu, Holger A. Russ, Shaohua Xu, Tao Xu, Yu Zhang, Tianhua Ma, Matthias Hebrok, Sheng Ding

Cell Stem Cell, Volume 14, Issue 2, 228-236, doi: 10.1016/j.stem.2014.01.006

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ZenMaster

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For more on stem cells and cloning, go to CellNEWS at

http://cellnews-blog.blogspot.com/

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