Fat-Based Stem Cell Treatment Suggests a New Way to Slow Scarring in Scleroderma Patients


Scleroderma is an autoimmune disease that causes chronic scarring of the skin and internal organs. The deposition of massive quantities of collagen decrease the pliability and elasticity of the skin, lungs, and blood vessels. As you might guess, the prognosis of scleroderma patients is quite poor and this disease causes a good deal of suffering and morbidity.

Treatments options usually include steroids, and other drugs that suppress the immune system, all of which have severe side effects.

New research from scientists at the Hospital for Special Surgery in New York City and other collaborating institutions, led by Dr. Teresa T. Lu, may have identified a new mechanism in operation during the onset and maintenance of scleroderma. This work was published in the Journal of Clinical Investigation.

In this study, scleroderma patients were shown to possess diminished numbers of “adipose-derived stromal cells” (ADSCs) in the layer of fat that underlies the upper layers of the skin. These fatty tissues are referred to as “dermal white adipose tissue.” The loss of these dermal white adipose tissue ADSCs tightly correlates with the onset of scarring in two different mouse model systems that recapitulate scleroderma in laboratory mice. These observations may show that ADSC loss contributes to scarring of the skin.

Why do these ADSCs die? Lu and her coworkers discovered that ADSC survival depends on the presence of particular molecules secreted by immune cells called “dendritic cells.” Skin-based dendritic cells secrete a molecule called lymphotoxin B. Although this molecule is called a toxin, it is required for ADSC survival. In laboratory mice that suffered from a scleroderma-like disease, artificial stimulation of the lymphotoxin B receptor in ADSCs amplified and eventually restored the numbers of ADSCs in the skin. Could stimulating ADSCs in this manner help treat scleroderma patients?

According the Dr. Lu, the administrating author of this publication, injecting “ADSCs is being tried in scleroderma; the possibility of stimulating the lymphotoxin B pathway to increase the survival of these stem cells is very exciting.” Dr. Lu continued, “By uncovering these mechanisms and targeting them with treatments, perhaps one day we can better treat the disease.”

Lu also thinks that a similar strategy that targets stem cells from other tissues might provide a treatment for other rheumatological conditions – such as systemic lupus erythematosis and rheumatoid arthritis. Additionally, bone and cartilage repair might also benefit from such a treatment strategy.

In the coming years, Dr. Lu and her colleagues hope to test the applicability of this work in human cells. If such a strategy works in human cells, then the next stop would be trial in human scleroderma patients. The success of such a treatment strategy would be a welcome addition to the treatment options for scleroderma patients, but only if this treatment is shown to be proven safe and effective.

“Improving ADSC therapy would be a major benefit to the field of rheumatology and to patients suffering from scleroderma,” said Lu.

USC Researchers Isolate Human Nephon Progenitor Cells – Future Possibilities for Kidney Regeneration


Researchers at the Saban Research Institute of Children’s Hospital of Los Angeles and the University of Southern California (USC) have reported the isolation of human nephron progenitor (NP) cells. These results, which were published in the journal Stem Cell Translational Medicine, might very well elucidate how progenitor cells differentiate into become renal cells and then develop into kidneys. Such insights could, possibly provide new strategies to promote renal regeneration after chronic kidney failure or acute kidney injury.

Kidneys are composed of about a million tiny filtration units known as “nephrons.” These diminutive structures filter waste and concentrate those wastes into urine, which is leaked into the bladder. In humans, approximately 500,000 to 1,000,000 nephrons are generated before week 34 – 36 of fetal gestation. However, at this point in development, the NP cells are exhausted and kidney development (known as “nephrogenesis”) effectively ceases. If the kidney loses a large enough quantity of nephrons after this time period, such losses may lead to irreversible kidney failure, since no further cell repair or regeneration is possible.

kidney_nephron_civyrose

In past studies, NPs were made from induced pluripotent stem cells, or by utilizing animal models. Scientists at USC and Children’s Hospital of Los Angeles (CHLA), chose a different tactic; they designed an efficient protocol by which they could directly isolate human NPs. To accomplish this, Dr. Laura Perin and her colleagues used RNA-labeling probes to obtain cells that expressed the SIX2 and CITED1 genes. Cells expressing both of these genes are almost certainly NPs, since SIX2 and CITED1 are master regulatory genes that promote renal development.

Dr. Perin, co-director of CHLA’s GOFARR Laboratory for Organ Regenerative Research and Cell Therapeutics in Urology, added, “In addition to defining the genetic profile of human NP, this system will facilitate studies of human kidney development, providing a novel tool for renal regeneration and bioengineering purposes.”

On a rather sanguine note, Perin noted that these experiments, which constitute proof-of-concept work, may create new applications to researchers who might be able to use her laboratory’s techniques to isolated progenitor cells for other organs, the pancreas, heart, or lung. “This technique provides a ‘how to’ of human tissue during development,” said Perin.

“It is an important tool that will allow scientists to study cell renewal and differentiation in human cells, perhaps offering clues to how to regulate such development,” added first author of this paper, Stefano Da Sacco.

Functional, Though Not Completely Structurally Normal Tissue-Engineered Livers Made from Adult Liver Cells


Tracy C. Grikscheit and her research team from the Saban Research Institute at the Children’s Hospital Los Angeles have produced functional, tissue-engineered human and mouse liver from adult stem and progenitor cells.

The largest organ in our bodies, the liver executes many vital functions. It is located in the upper right portion of the abdomen protected by the rib cage. The liver has two main lobes that are divided into many tiny lobules.

Liver cells are supplied by two different sources of blood. The hepatic artery provides oxygen-rich blood from the heart and the portal vein supplies nutrients from the intestine and the spleen. Normally, veins return blood from the body to the heart, but the portal vein allows chemicals from the digestive tract to enter the liver for “detoxification” and filtering prior to entering the general circulation. The portal vein also delivers the precursors liver cells need to produce the proteins, cholesterol, and glycogen required for normal body activities.

The liver also makes bile. Bile is a mixture of water, bile acids (made from stored cholesterol in the liver), and other sundry chemicals. Bile made by the liver is then stored in the gallbladder. When food enters the duodenum (the uppermost part of the small intestine), the gallbladder contracts and secretes bile is secreted into the duodenum, to aid in the digestion of fats in food.

The liver also stores extra sugar in the form of glycogen, which is converted back into glucose when the body needs it for energy. It also produces blood clotting factors, processes and stores iron for red blood cell production, converts toxic nitrogenous wastes (usually in the form of ammonium) into urea, which is excreted in urine. Finally, the liver also metabolizes foreign substances, like drugs into substances that can effectively excreted by the kidneys.

Both adults and children are affected by various types of liver disease. Liver can be caused by infectious hepatitis, which is caused by a variety of viruses, chronic alcoholism, inherited liver abnormalities (e.g., Wilson’s disease, hemochromatosis, Gilbert’s disease) or various types of liver cancer. One in ten people in the United States suffer from liver cancer and need a liver transplant. Liver transplantation is the only effective treatment for end-stage liver disease, but the scarcity of liver donors and the necessity of life-long immunosuppressive therapy limit treatment options. In some cases (such as inborn errors of metabolism or acute bouts of liver insufficiency), patients may be effectively treated by transplanting small quantities of functional liver tissue.

Alternate approaches that have been investigated, but these protocols have significant limitations. For example, “hepatocyte transplantation” involves the infusion of liver cells from a donated liver. This protocol, however, wastes many cells that do not integrate into the existing liver and such a treatment is usually little more than a stop-gap solution, since most patients require a liver transplant within a year of this treatment.

Human-induced pluripotent stem (iPS) cells are another possibility but, so far, iPS cells differentiate into immature rather than mature, functional, proliferative hepatocytes.

A need remains for a robust treatment that can eliminate the need for immunosuppressive theory. “We hypothesized that by modifying the protocol used to generate intestine, we would be able to develop liver organoid units that could generate functional tissue-engineered liver when transplanted,” said Dr. Grikscheit.

Grikscheit and her co-workers extracted hardy, multicellular clusters of liver cells known as liver organoid units (LOUs) from resected human and mouse livers. These LOUs were loaded onto scaffolds made from nonwoven polyglycolic acid fibers. These scaffolds are completely biodegradable and they provide a structure upon which the LOUs can grow, fuse, and form a structure that resembles a liver.

After transplantation of the LOU/scaffold combinations, they generated tissue-engineered livers or TELis. Tissue-engineered livers developed from the human and mouse LOUs and possessed a variety of key liver-specific cell types that are required for normal hepatic function. However, the cellular organization of these TELis did differ from native liver tissue.

The tissue-engineered livers (TELis) made by Grikscheit’s laboratory contained normal liver components such as hepatocytes that properly expressed the liver-specific protein albumin, CK19-expressing bile ducts, vascular structures surrounded by smooth muscles that expressed smooth muscle-specific actin, desmin-expressing stellate cells, and CD31-expressing endothelial cells. The production of albumin by the TELi hepatocytes indicated that these cells were executing their normal secretory function. In a mouse model of liver failure, their tissue-engineered liver provided some hepatic function. In addition, the hepatocytes proliferated in the tissue-engineered liver.

A cellular therapy for liver disease that utilizes technologies like this would completely change the treatment options for many patients. In particular, children with metabolic disorders and require a new liver to survive might see particular benefits if such a treatment can come to the clinic. By generating functional hepatocytes comparable to those in native liver, establishing that these cells are functional and proliferative, Grikscheit and her colleagues have moved one step closer to that goal.

To access this paper, please see: Nirmala Mavila et al., “Functional Human and Murine Tissue-Engineered Liver Is Generated From Adult Stem/Progenitor Cells,” Stem Cells Translational Medicine, August 2016 DOI: 10.5966/sctm.2016-0205.

AUF1 Gene Important Inducer of Muscle Repair


A new study in the laboratory of Robert J. Schneider at NYU Langone and his collaborators has uncovered a gene that plays integral roles in the repair of injured muscle throughout life. This investigation shows that this previously “overlooked” gene might play a pivotal role in “sarcopenia,” which refers to the loss of muscle tissues with age.

This collaboration between scientists at NYU Langone Medical Center and the University of Colorado at Boulder showed that the levels of a protein called AUF1 determine if stem cell populations retain the ability to regenerate muscle after injury and as mice age.

Changes in the activity of AUF1 have also been linked by past studies to human muscle diseases. More than 30 genetic diseases, known collectively as myopathies, show defective muscle regeneration and these anomalies cause muscles to weaken or waste away.

For example, muscular dystrophy is a disease in which abnormal muscles fail to function properly and undergo normal repair. Although the signs and symptoms of Duchenne Muscular Dystrophy vary, in some cases wildly, this disease develops in infants and affects and weakens the torso and limb muscles beginning in young adulthood. Sarcopenia, in healthy individuals occurs in older patients.

Skeletal muscles have a stem cell population set aside for muscle repair known as satellite cells. These cells divide and differentiate into skeletal muscle when skeletal muscle is damaged, and as we age, the capacity of muscle satellite cells to repair muscle decreases.

AUF1 is a protein that regulates muscle stem cell function by inducing the degradation of specific, targeted messenger RNAs (mRNAs). According to Robert Schneider, “This work places the origin of certain muscle diseases squarely within muscle stem cells, and shows that AUF1 is a vital controller of adult muscle stem cell fate.” He continued: “The stem cell supply is remarkably depleted when the AUF1 signal is defective, leaving muscles to deteriorate a little more each time repair fails after injury.”

The experiments in this study demonstrated that mice that lack AUF1 display accelerated skeletal muscle wasting as they age. These AUF1-depleted mice also showed impaired skeletal muscle repair following injury. When the molecular characteristics of these AUF1-depleted muscle satellite cells were examined, Schneider and his collaborators showed that auf1−/− satellite cells had increased stability and overexpression of so-called “ARE-mRNAs.” ARE mRNAs contain AU-rich elements at their tail-ends. AUF1 proteins bind to these ARE mRNAs and induce their degradation. In the absence of AUF1, muscle satellite cells accumulate ARE mRNAs. One of these ARE mRNAs includes that which encodes matrix metalloprotease, MMP9. Overexpression of MMP9 by aging muscle satellite cells causes degradation of the skeletal muscle matrix, which prevents satellite-cell-mediated regeneration of muscles. Consequently, the muscle satellite cells return to their quiescent state and fail to divide and repair skeletal muscle.

When Schneider and his coworkers and collaborators blocked MMP9 activity in auf1−/− mice, they found that they had restored skeletal muscle repair and maintenance of the satellite cell population.

These experiments suggest that repurposing drugs originally developed for cancer treatment that blocks MMP9 activity might be a way to dial down age-related sarcopenia.

“This provides a potential path to clinical treatments that accelerate muscle regeneration following traumatic injury, or in patients with certain types of adult onset muscular dystrophy,” said Schneider.

This work was published here: Devon M. Chenette et al., “Targeted mRNA Decay by RNA Binding Protein AUF1 Regulates Adult Muscle Stem Cell Fate, Promoting Skeletal Muscle Integrity,” Cell Reports, 2016; DOI: 10.1016/j.celrep.2016.06.095.

Behavior Of Brain Stem Cells Controlled By Cerebrospinal Fluid Signals


The choroid plexus is a network of blood vessels in each ventricle of the brain. It is derived from the pia mater and produces the cerebrospinal fluid.  The choroid plexus, unfortunately, has been ignored to some degree when it comes to brain research.  However, CSF turns to be an important regulator of adult neural stem cells, research indicates.

A new study led by Prof. Fiona Doetsch at the Biozentrum of the University of Basel, Switzerland has shown that signals secreted by the choroid plexus dynamically change during aging, and these different signals affect the behavior of aged stem cells.

In the adult brain, neural stem cell populations in various places throughout the central nervous system divide to give rise to neurons and glial cells throughout our lives. These stem cells reside in unique micro-environments (known as niches) that provide key signals that regulate stem cell self-renewal and differentiation. Stem cells in the adult brain contact the ventricles, which are cavities in the brain filled with CSF. CSF bathes and protects the brain and is produced by the cells of the choroid plexus.

Ventricular System of the Brain

Doetsch and her coworkers have shown that the choroid plexus is a key component of the stem cell niche, and that the properties of this stem cell niche change throughout life and affect stem cell behavior.

Doetsch’s group discovered that the choroid plexus secretes a cocktail of important signaling factors into the CSF. These CSF-secreted growth factors are important in stem cell regulation throughout life. As we age, the levels of stem cell division and formation of new neurons decrease. They also showed that although stem cells are still present in the aged brain, and have the capacity to divide, their ability to do so have significantly decreased.

Graphical abstract

“One reason is that signals in the old choroid plexus are different. As a consequence, stem cells receive different messages and are less capable to form new neurons during aging. In other words, compromising the fitness of stem cells in this brain region,” said Violeta Silva Vargas, first author of the paper that appeared in the journal Cell Stem Cell. “But what is really amazing is that when you cultivate old stem cells with signals from young fluid, they can still be stimulated to divide, behaving like the young stem cells.”

In the future, Doetsch and her group plans to tease out the composition of the signaling factors secreted by the choroid plexus.  They would also like to know how the composition of this growth factor cocktail changes as a result of changes in brain states and how these changes affect neural stem cells. This could provide new ways to understand brain function in health and disease.

“We can imagine the choroid plexus as a watering can that provides signals to the stem cells. Our investigations also open a new route for understanding how different physiological states of the body influence stem cells in the brain during health and disease, and opens new ways for thinking about therapy,” said Doetsch.

This work was published here: Violeta Silva-Vargas et al., “Age-Dependent Niche Signals from the Choroid Plexus Regulate Adult Neural Stem Cells,” Cell Stem Cell, 2016; DOI: 10.1016/j.stem.2016.06.013.

Computer Simulations of MSC-Heart Muscle Interactions Identify A Family of MSCs that Produce Few Side Effects


A research team at the Icahn School of Medicine at Mount Sinai has utilized a mathematical modeling to simulate the delivery of human mesenchymal stem cells to a damaged heart. In doing so, they found that a particular subset of harvested MSCs minimizes the risks associated with this therapy. This study represents a development that could lead to novel strategies to repair and regenerate heart muscle and might even improve stem cell treatments for heart attack patients.

In the United States alone, one person suffers a myocardial infarction or heart attack every 43 seconds (on the average). The urgency of this situation has motivated stem cells scientists and cardiologists to develop novel therapies to repair and regenerate heart muscle. One of these therapies includes the implantation of human mesenchymal stem cells (hMSCs). However, in clinical trials the benefits of hMSC implantation have often been modest and even transient. This might reflect our understanding of the mechanism by which hMSCs influence cardiac function.

Kevin D. Costa and his colleagues at the Icahn School of Medicine have used mathematical modeling to simulate the electrical interactions between implanted hMSCs and endogenous heart cells. They hoped to eventually understand the possible adverse effects of hMSC transplantation and new methods for reducing some potential risks of this therapy.

Implanted hMSCs can disrupt the electrical connections between heart muscle cells and can even cause the heart to beat irregularly; a condition called “arrhythmias.” One particular type of hMSCs, however, did not express an ion channel called EAG1 (which stands for “ether-a-go-go”). The EAG1-less hMSCs did not cause arrhythmias at nearly the rate as the EAG1-containing hMSC, in computer simulations run by Costa’s group.

These EAG1-less hMSCs, also known as “Type C” MSCs, minimized electrochemical disturbances in cardiac single-cell and tissue-level electrical activity. The benefits of these EAG1-less hMSCs may enhance the safety of hMSC treatments in heart attack patients who receive stem cell therapy. This advance could therefore lead to new clinical trials and future improvements in treatment of patients with heart failure.

Costa’s study might provide a template for future computational studies on mesenchymal stem cells. It also provides novel insights into hMSC-heart cell interactions that can guide future experimental studies to understand the mechanisms that underlie hMSC therapy for the heart.

This work was published in Joshua Mayourian et al., “Modeling Electrophysiological Coupling and Fusion between Human Mesenchymal Stem Cells and Cardiomyocytes, PLOS Computational Biology, 2016; 12 (7): e1005014. DOI: 10.1371/journal.pcbi.1005014.

Adjustable Gels Used to Determine Those Molecules That Drive Stem Cell Differentiation


Scientists have been very interested in the details of stem cell differentiation. To that end, several laboratories have designed hydrogels that mimic the stiffness of biological tissue in order to grow stem cells and study their differentiation.

In one enterprising laboratory, led by Rein Ulijn of the City University of New York and the University of Strathclyde, scientists have used a novel culture-based gel system to study mesenchymal stem cell differentiation and identify those metabolites used by stem cells when they select bone and cartilage cell fates. When these molecules are provided to standard stem cell cultures, these molecules can guide stem cells to generate desired cell types. This new study illustrates how new biomaterials can provide an exacting model system that can help scientists precisely determine those identifying factors that drive stem cell differentiation.

Stem-cell scientists have known that the rigidity of a hydrogel surface can instruct stem cells to differentiate. A rigid surface, as it turns out, can result in bone cell formation, whereas soft surfaces induce the differentiation of cells into neuron-like cells. With this information, Ulijn and others developed a protocol that generates gels by combining small building-block molecules that spontaneously form a network of nanosized fibers. Furthermore, by varying the concentration of these building blocks, the stiffness of these gels can be adjusted. By mimicking the stiffness of bone (40 kilopascal) or cartilage (15 kilopascal), the gel stimulates stem cells applied to its surface to differentiate accordingly.

“This paper is a great example of how chemistry can help make step changes in biology,” said Matthew Dalby of the University of Glasgow and co-senior author. “As a biologist, I needed simple yet tunable cell-culture gels that would give me a defined system to study metabolites in the laboratory. Rein had developed the chemistry to allow this to happen.”

The available gels for growing stem cells are typically derived from animal products. Unfortunately, this can affect the reproducibility of results, since different preparations of particular animal products can have rather different properties. Synthetic components usually require coatings or coupling of cell-adhesive ligands. However, the gel developed by Ulijn’s group is composed of two simple synthetic peptide derivatives. One component binds to copies of itself with high directional preference, which results in the spontaneous formation of nanoscale fibers when the molecules are dissolved in water. The second components consists of a surfactant-like molecule that binds to the fiber surface and presents simple, cell-compatible chemical groups to any cells.

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The components are held together by relatively weak and reversible interactions, e.g., hydrogen bonding and aromatic stacking. Interestingly, variants of these gels are commercially available through a spinoff company called Biogelx, Ltd., where Ulijn serves as chief scientific officer.

“We wanted a platform that provides nanofiber morphology and as-simple-as-possible chemistry and tunable stiffness to serve as a blank-slate background so that we could focus on changes in stem cell metabolism,” said Ulijn. “Matt and his team performed metabolomics analysis to find out how the key metabolites within a stem cell are used up during the differentiation process.”

Particular transcription factors are often the ingredients scientists use to induce stem cell fate in the case of induced pluripotent stem cells. However, Dalby and Ulijn think that certain metabolites might drive those pathways that cause the different intracellular concentrations of transcription factors that drive the various differentiation pathways.

One metabolite featured in the study is cholesterol sulfate. Cholesterol sulfate is used up during osteogenesis on a rigid matrix and can also be used to convert stem cells into bone-like cells in cell culture.

In their paper, Ulijn and his coworkers showed how small molecules, like cholesterol sulfate, can put into motion those cell-signaling pathways that culminate in the activation of the transcription factors that drive the transcription of major bone-related genes. The expression of these bone-specific genes drives bone formation, and this demonstrates a connection between the metabolites and the activation of transcription factors.

It must be noted that this gel does not precisely recapitulate the microenvironment inside the body. Therefore, it is unclear if the stem cells grown on it behave differently on the designed gel surfaces than they would in the body.

Although the full list of metabolites derived from the analysis is preliminary, “it could certainly point researchers in the right direction,” Ulijn said. “Our ambition is to simplify drug discovery by using the cell’s own metabolites as drug candidates,” Dalby said.

This paper was published here: Alakpa et al., “Tunable Supramolecular Hydrogels for Selection of Lineage Guiding Metabolites in Stem Cell Cultures,” Chem, 2016 DOI:10.1016/j.chempr.2016.07.001.