Rejuvenating stem cells in the aging brain of mice improve the regeneration of injured or diseased areas in the brain

via University of Luxembourg

Scientists from the Luxembourg Centre for Systems Biomedicine (LCSB) of the University of Luxembourg and from the German Cancer Research Center (DKFZ) have been able to rejuvenate stem cells in the brain of aging mice.

The revitalised stem cells improve the regeneration of injured or diseased areas in the brain of old mice. The researchers expect that their approach will provide fresh impetus in regenerative medicine and facilitate the development of stem cell therapies. Their results were published today in the journal “Cell”.

All cells making up our organs originate from stem cells. They divide and the resulting cells develop into specific tissue cells, forming the brain, lungs or bone marrow. With age, however, the stem cells of living organisms lose their ability to proliferate. Many of them lapse into a permanent state of quiescence.

In order to create as accurate as possible computational models of stem cell behaviour, the LCSB’s Computational Biology Group led by Prof. Antonio del Sol applied a novel approach. “Stem cells live in a niche where they constantly interact with other cells and extra-cellular components. It is extremely difficult to model such a plethora of complex molecular interactions on the computer. So we shifted perspective. We stopped thinking about what external factors were affecting the stem cells, and started thinking about what the internal state of a stem cell would be like in its precisely defined niche.”

The novel approach led to in a new computational model developed by Dr. Srikanth Ravichandran of the Computational Biology Group: “Our model can determine which proteins are responsible for the functional state of a given stem cell in its niche – meaning whether it will divide or remain in a state of quiescence. Our model relies on the information about which genes are being transcribed. Modern cell biology technologies enable profiling of gene expression at single cell resolution.”

It was previously unknown why most of the stem cells in the brain of old mice remain in a state of quiescence. From their computational model, the researchers at the LCSB identified a molecule called sFRP5 that keeps the neuronal stem cells inactive in old mice, and prevents proliferation by blocking the Wnt pathway crucial for cell differentiation.

A rejuvenation for stem cells

Then the long-standing expertise in neural stem cells of the collaborators at the German Cancer Research Center (DKFZ) came in: Studying stem cells first in a dish and then later directly in mice, they could experimentally validate the computational prediction. When neutralising the action of sFRP5, the quiescent stem cells did indeed start proliferating more actively. Thus, they were available again to be recruited for the regeneration processes in the aging brain. “With the deactivation of sFRP5, the cells undergo a kind of rejuvenation,” del Sol says: “As a result, the ratio of active to dormant stem cells in the brain of old mice becomes almost as favourable as in young animals.”

“Our results constitute an important step towards the implementation of stem cell-based therapies, for instance for neurodegenerative diseases,” Antonio del Sol says. “We were able to show that, with computational models, it is possible to identify the essential features that are characteristic of a specific state of stem cells.” This approach is not limited to studying the brain. It can also be used to model stem cells of other organs in the body. “The hope is that this will open avenues for regenerative medicine,” says del Sol.

Learn more: Scientists rejuvenate stem cells in the aging brain of mice

 

 

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Age-related loss of stem cell function can be reversed by a 24-hour fast

Intestinal stem cells from mice that fasted for 24 hours, at right, produced much more substantial intestinal organoids than stem cells from mice that did not fast, at left.
Image: Maria Mihaylova and Chia-Wei Cheng

A drug treatment that mimics fasting can also provide the same benefit, study finds

As people age, their intestinal stem cells begin to lose their ability to regenerate. These stem cells are the source for all new intestinal cells, so this decline can make it more difficult to recover from gastrointestinal infections or other conditions that affect the intestine.

This age-related loss of stem cell function can be reversed by a 24-hour fast, according to a new study from MIT biologists. The researchers found that fasting dramatically improves stem cells’ ability to regenerate, in both aged and young mice.

In fasting mice, cells begin breaking down fatty acids instead of glucose, a change that stimulates the stem cells to become more regenerative. The researchers found that they could also boost regeneration with a molecule that activates the same metabolic switch. Such an intervention could potentially help older people recovering from GI infections or cancer patients undergoing chemotherapy, the researchers say.

“Fasting has many effects in the intestine, which include boosting regeneration as well as potential uses in any type of ailment that impinges on the intestine, such as infections or cancers,” says Omer Yilmaz, an MIT assistant professor of biology, a member of the Koch Institute for Integrative Cancer Research, and one of the senior authors of the study. “Understanding how fasting improves overall health, including the role of adult stem cells in intestinal regeneration, in repair, and in aging, is a fundamental interest of my laboratory.”

David Sabatini, an MIT professor of biology and member of the Whitehead Institute for Biomedical Research and the Koch Institute, is also a senior author of the paper, which appears in the May 3 issue of Cell Stem Cell.

“This study provided evidence that fasting induces a metabolic switch in the intestinal stem cells, from utilizing carbohydrates to burning fat,” Sabatini says. “Interestingly, switching these cells to fatty acid oxidation enhanced their function significantly. Pharmacological targeting of this pathway may provide a therapeutic opportunity to improve tissue homeostasis in age-associated pathologies.”

The paper’s lead authors are Whitehead Institute postdoc Maria Mihaylova and Koch Institute postdoc Chia-Wei Cheng.

Boosting regeneration

For many decades, scientists have known that low caloric intake is linked with enhanced longevity in humans and other organisms. Yilmaz and his colleagues were interested in exploring how fasting exerts its effects at the molecular level, specifically in the intestine.

Intestinal stem cells are responsible for maintaining the lining of the intestine, which typically renews itself every five days. When an injury or infection occurs, stem cells are key to repairing any damage. As people age, the regenerative abilities of these intestinal stem cells decline, so it takes longer for the intestine to recover.

“Intestinal stem cells are the workhorses of the intestine that give rise to more stem cells and to all of the various differentiated cell types of the intestine. Notably, during aging, intestinal stem function declines, which impairs the ability of the intestine to repair itself after damage,” Yilmaz says. “In this line of investigation, we focused on understanding how a 24-hour fast enhances the function of young and old intestinal stem cells.”

After mice fasted for 24 hours, the researchers removed intestinal stem cells and grew them in a culture dish, allowing them to determine whether the cells can give rise to “mini-intestines” known as organoids.

The researchers found that stem cells from the fasting mice doubled their regenerative capacity.

“It was very obvious that fasting had this really immense effect on the ability of intestinal crypts to form more organoids, which is stem-cell-driven,” Mihaylova says. “This was something that we saw in both the young mice and the aged mice, and we really wanted to understand the molecular mechanisms driving this.”

Metabolic switch

Further studies, including sequencing the messenger RNA of stem cells from the mice that fasted, revealed that fasting induces cells to switch from their usual metabolism, which burns carbohydrates such as sugars, to metabolizing fatty acids. This switch occurs through the activation of transcription factors called PPARs, which turn on many genes that are involved in metabolizing fatty acids.

The researchers found that if they turned off this pathway, fasting could no longer boost regeneration. They now plan to study how this metabolic switch provokes stem cells to enhance their regenerative abilities.

They also found that they could reproduce the beneficial effects of fasting by treating mice with a molecule that mimics the effects of PPARs. “That was also very surprising,” Cheng says. “Just activating one metabolic pathway is sufficient to reverse certain age phenotypes.”

Jared Rutter, a professor of biochemistry at the University of Utah School of Medicine, described the findings as “interesting and important.”

“This paper shows that fasting causes a metabolic change in the stem cells that reside in this organ and thereby changes their behavior to promote more cell division. In a beautiful set of experiments, the authors subvert the system by causing those metabolic changes without fasting and see similar effects,” says Rutter, who was not involved in the research. “This work fits into a rapidly growing field that is demonstrating that nutrition and metabolism has profound effects on the behavior of cells and this can predispose for human disease.”

The findings suggest that drug treatment could stimulate regeneration without requiring patients to fast, which is difficult for most people. One group that could benefit from such treatment is cancer patients who are receiving chemotherapy, which often harms intestinal cells. It could also benefit older people who experience intestinal infections or other gastrointestinal disorders that can damage the lining of the intestine.

The researchers plan to explore the potential effectiveness of such treatments, and they also hope to study whether fasting affects regenerative abilities in stem cells in other types of tissue.

Learn more: Fasting boosts stem cells’ regenerative capacity

 

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Success in generating cartilage tissue by printing stem cells using a 3D-bioprinter

via University of Gothenburg

A team of researchers at Sahlgrenska Academy has managed to generate cartilage tissue by printing stem cells using a 3D-bioprinter. The fact that the stem cells survived being printed in this manner is a success in itself. In addition, the research team was able to influence the cells to multiply and differentiate to form chondrocytes (cartilage cells) in the printed structure.

The findings have been published in Nature’s Scientific Reports magazine. The research project is being conducted in collaboration with a team of researchers at the Chalmers University of Technology who are experts in the 3D printing of biological materials. Orthopedic researchers from Kungsbacka are also involved in the research collaboration.

The team used cartilage cells harvested from patients who underwent knee surgery, and these cells were then manipulated in a laboratory, causing them to rejuvenate and revert into “pluripotent” stem cells, i.e. stem cells that have the potential to develop into many different types of cells. The stem cells were then expanded and encapsulated in a composition of nanofibrillated cellulose and printed into a structure using a 3D bioprinter. Following printing, the stem cells were treated with growth factors that caused them to differentiate correctly, so that they formed cartilage tissue.

Tricked into thinking that they aren’t alone

The publication in Scientific Reports is the result of three years of hard work.

“In nature, the differentiation of stem cells into cartilage is a simple process, but it’s much more complicated to accomplish in a test tube. We’re the first to succeed with it, and we did so without any animal testing whatsoever,” says Stina Simonsson, Associate Professor of Cell Biology, who lead the research team’s efforts.

Most of the team’s efforts had to do with finding a procedure so that the cells survive printing, multiply and a protocol that works that causes the cells to differentiate to form cartilage.

“We investigated various methods and combined different growth factors. Each individual stem cell is encased in nanocellulose, which allows it to survive the process of being printed into a 3D structure. We also harvested mediums from other cells that contain the signals that stem cells use to communicate with each other so called conditioned medium. In layman’s terms, our theory is that we managed to trick the cells into thinking that they aren’t alone,” clarifies Stina Simonsson. Therefore the cells multiplied before we differentiated them.
A key insight gained from the team’s study is that it is necessary to use large amounts of live stem cells to form tissue in this manner.

The cartilage formed by the stem cells in the 3D bioprinted structure is extremely similar to human cartilage. Experienced surgeons who examined the artificial cartilage saw no difference when they compared the bioprinted tissue to real cartilage, and have stated that the material has properties similar to their patients’ natural cartilage. Just like normal cartilage, the lab-grown material contains Type II collagen , and under the microscope the cells appear to be perfectly formed, with structures similar to those observed in samples of human-harvested cartilage.

Potential for use in osteoarthritis therapies

The study represents a giant step forward in the ability to generate new, endogenous cartilage tissue. In the not too distant future, it should be possible to use 3D bioprinting to generate cartilage based on a patient’s own, “backed-up” stem cells. This bioprinted tissue can be used to repair cartilage damage, or to treat osteoarthritis, in which joint cartilage degenerates and breaks down. The condition is very common – one in four Swedes over the age of 45 suffer from some degree of osteoarthritis.

In theory, this research has created the opportunity to generate large amounts of cartilage, but one major issue must be resolved before the findings can be used in practice to benefit patients.

“The structure of the cellulose we used might not be optimal for use in the human body. Before we begin to explore the possibility of incorporating the use of 3D bioprinted cartilage into the surgical treatment of patients, we need to find another material that can be broken down and absorbed by the body so that only the endogenous cartilage remains, the most important thing for use in a clinical setting is safety” explains Stina Simonsson.

Learn more: Success in the 3D Bioprinting of Cartilage

 

 

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Edited stem cells fight inflammation caused by arthritis and other chronic conditions

ELLA MARUSHCHENKO
Using CRISPR technology, a team of researchers led by Farshid Guilak, PhD, at Washington University School of Medicine in St. Louis, rewired stem cells’ genetic circuits to produce an anti-inflammatory arthritis drug when the cells encounter inflammation. The technique eventually could act as a vaccine for arthritis and other chronic conditions.

Goal is vaccine that targets inflammation in joints

Using new gene-editing technology, researchers have rewired mouse stem cells to fight inflammation caused by arthritis and other chronic conditions. Such stem cells, known as SMART cells (Stem cells Modified for Autonomous Regenerative Therapy), develop into cartilage cells that produce a biologic anti-inflammatory drug that, ideally, will replace arthritic cartilage and simultaneously protect joints and other tissues from damage that occurs with chronic inflammation.

The cells were developed at Washington University School of Medicine in St. Louis and Shriners Hospitals for Children-St. Louis, in collaboration with investigators at Duke University and Cytex Therapeutics Inc., both in Durham, N.C. The researchers initially worked with skin cells taken from the tails of mice and converted those cells into stem cells. Then, using the gene-editing tool CRISPR in cells grown in culture, they removed a key gene in the inflammatory process and replaced it with a gene that releases a biologic drug that combats inflammation.

The research is available online April 27 in the journal Stem Cell Reports.

“Our goal is to package the rewired stem cells as a vaccine for arthritis, which would deliver an anti-inflammatory drug to an arthritic joint but only when it is needed,” said Farshid Guilak, PhD, the paper’s senior author and a professor of orthopedic surgery at Washington University School of Medicine. “To do this, we needed to create a ‘smart’ cell.”

Many current drugs used to treat arthritis — including Enbrel, Humira and Remicade — attack an inflammation-promoting molecule called tumor necrosis factor-alpha (TNF-alpha). But the problem with these drugs is that they are given systemically rather than targeted to joints. As a result, they interfere with the immune system throughout the body and can make patients susceptible to side effects such as infections.

“We want to use our gene-editing technology as a way to deliver targeted therapy in response to localized inflammation in a joint, as opposed to current drug therapies that can interfere with the inflammatory response through the entire body,” said Guilak, also a professor of developmental biology and of biomedical engineering and co-director of Washington University’s Center of Regenerative Medicine. “If this strategy proves to be successful, the engineered cells only would block inflammation when inflammatory signals are released, such as during an arthritic flare in that joint.”

As part of the study, Guilak and his colleagues grew mouse stem cells in a test tube and then used CRISPR technology to replace a critical mediator of inflammation with a TNF-alpha inhibitor.

“Exploiting tools from synthetic biology, we found we could re-code the program that stem cells use to orchestrate their response to inflammation,” said Jonathan Brunger, PhD, the paper’s first author and a postdoctoral fellow in cellular and molecular pharmacology at the University of California, San Francisco.

Over the course of a few days, the team directed the modified stem cells to grow into cartilage cells and produce cartilage tissue. Further experiments by the team showed that the engineered cartilage was protected from inflammation.

“We hijacked an inflammatory pathway to create cells that produced a protective drug,” Brunger said.

The researchers also encoded the stem/cartilage cells with genes that made the cells light up when responding to inflammation, so the scientists easily could determine when the cells were responding. Recently, Guilak’s team has begun testing the engineered stem cells in mouse models of rheumatoid arthritis and other inflammatory diseases.

If the work can be replicated in animals and then developed into a clinical therapy, the engineered cells or cartilage grown from stem cells would respond to inflammation by releasing a biologic drug — the TNF-alpha inhibitor — that would protect the synthetic cartilage cells that Guilak’s team created and the natural cartilage cells in specific joints.

“When these cells see TNF-alpha, they rapidly activate a therapy that reduces inflammation,” Guilak explained. “We believe this strategy also may work for other systems that depend on a feedback loop. In diabetes, for example, it’s possible we could make stem cells that would sense glucose and turn on insulin in response. We are using pluripotent stem cells, so we can make them into any cell type, and with CRISPR, we can remove or insert genes that have the potential to treat many types of disorders.”

With an eye toward further applications of this approach, Brunger added, “The ability to build living tissues from ‘smart’ stem cells that precisely respond to their environment opens up exciting possibilities for investigation in regenerative medicine.”

Learn more: Stem cells edited to fight arthritis

 

 

 

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New technology could lead to a better way to differentiate stem cells for nerve regrowth

Iowa State University researchers, left to right, Metin Uz, Suprem Das, Surya Mallapragada and Jonathan Claussen are developing technologies to promote nerve regrowth. The monitor shows mesenchymal stem cells (the white) aligned along graphene circuits (the black). Larger photo. Photo by Christopher Gannon

Researchers looking for ways to regenerate nerves can have a hard time obtaining key tools of their trade.

Schwann cells are an example. They form sheaths around axons, the tail-like parts of nerve cells that carry electrical impulses. They promote regeneration of those axons. And they secrete substances that promote the health of nerve cells.

In other words, they’re very useful to researchers hoping to regenerate nerve cells, specifically peripheral nerve cells, those cells outside the brain and spinal cord.

But Schwann cells are hard to come by in useful numbers.

So researchers have been taking readily available and noncontroversial mesenchymal stem cells (also called bone marrow stromal stem cells that can form bone, cartilage and fat cells) and using a chemical process to turn them, or as researchers say, differentiate them into Schwann cells. But it’s an arduous, step-by-step and expensive process.

Researchers at Iowa State University are exploring what they hope will be a better way to transform those stem cells into Schwann-like cells. They’ve developed a nanotechnology that uses inkjet printers to print multi-layer graphene circuits and also uses lasers to treat and improve the surface structure and conductivity of those circuits.

It turns out mesenchymal stem cells adhere and grow well on the treated circuit’s raised, rough and 3-D nanostructures. Add small doses of electricity – 100 millivolts for 10 minutes per day over 15 days – and the stem cells become Schwann-like cells.

The researchers’ findings are featured on the front cover of the scientific journal Advanced Healthcare Materials. Jonathan Claussen, an Iowa State assistant professor of mechanical engineering and an associate of the U.S. Department of Energy’s Ames Laboratory, is lead author. Suprem Das, a postdoctoral research associate in mechanical engineering and an associate of the Ames Laboratory; and Metin Uz, a postdoctoral research associate in chemical and biological engineering, are first authors.

The project is supported by funds from the Roy J. Carver Charitable Trust, the U.S. Army Medical Research and Materiel Command, Iowa State’s College of Engineering, the department of mechanical engineering and the Carol Vohs Johnson Chair in Chemical and Biological Engineering held by Surya Mallapragada, an Anson Marston Distinguished Professor in Engineering, an associate of the Ames Laboratory and a paper co-author.

“This technology could lead to a better way to differentiate stem cells,” Uz said. “There is huge potential here.”

The electrical stimulation is very effective, differentiating 85 percent of the stem cells into Schwann-like cells compared to 75 percent by the standard chemical process, according to the research paper. The electrically differentiated cells also produced 80 nanograms per milliliter of nerve growth factor compared to 55 nanograms per milliliter for the chemically treated cells.

The researchers report the results could lead to changes in how nerve injuries are treated inside the body.

“These results help pave the way for in vivo peripheral nerve regeneration where the flexible graphene electrodes could conform to the injury site and provide intimate electrical stimulation for nerve cell regrowth,” the researchers wrote in a summary of their findings.

The paper reports several advantages to using electrical stimulation to differentiate stem cells into Schwann-like cells:

  • doing away with the arduous steps of chemical processing
  • reducing costs by eliminating the need for expensive nerve growth factors
  • potentially increasing control of stem cell differentiation with precise electrical stimulation
  • and creating a low maintenance, artificial framework for neural damage repairs.

A key to making it all work is a graphene inkjet printing process developed in Claussen’s research lab. The process takes advantages of graphene’s wonder-material properties – it’s a great conductor of electricity and heat, it’s strong, stable and biocompatible – to produce low-cost, flexible and even wearable electronics.

But there was a problem: once graphene electronic circuits were printed, they had to be treated to improve electrical conductivity. That usually meant high temperatures or chemicals. Either could damage flexible printing surfaces including plastic films or paper.

Claussen and his research group solved the problem by developing computer-controlled laser technology that selectively irradiates inkjet-printed graphene oxide. The treatment removes ink binders and reduces graphene oxide to graphene – physically stitching together millions of tiny graphene flakes. The process makes electrical conductivity more than a thousand times better.

The collaboration of Claussen’s group of nanoengineers developing printed graphene technologies and Mallapragada’s group of chemical engineers working on nerve regeneration began with some informal conversations on campus.

That led to experimental attempts to grow stem cells on printed graphene and then to electrical stimulation experiments.

“We knew this would be a really good platform for electrical stimulation,” Das said. “But we didn’t know it would differentiate these cells.”

But now that it has, the researchers say there are new possibilities to think about. The technology, for example, could one day be used to create dissolvable or absorbable nerve regeneration materials that could be surgically placed in a person’s body and wouldn’t require a second surgery to remove.

 

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