Activation of the Proteasome Enhances Stem Cell Function and Lifespan


As we age, the capacity of our stem cells to heal and replace damaged cells and tissues decline. This age-associated decrease in adult stem cell function seems to be a major contributor to the physiological decline during aging. A new paper, by Efstathios Gonos and his colleagues at the National Hellenic Research Foundation in Athens, Greece gives one possible technique that might improve the function of stem cells in an aging body.

Cells contain a multiprotein complex called the “proteasome” that degrades unneeded or defective proteins. The proteasome controls protein half-lives, function, and the protein composition of the cell. Functional failure of the proteasome has been linked to various biological phenomena including senescence and aging. The role of the proteasome in stem cells aging, however has received little attention to date.

Proteasome figure

Gonos and his coworkers used mesenchymal stem cells from umbilical cord Wharton’s Jelly and human fat. Because they were able to compare the proteasome activity in very young and aged stem cells, Gonos and others discovered a significant age-related decline in proteasome content and activity between these two types of stem cells. The proteasome from Warton’s Jelly mesenchymal stem cells were consistently more active and displayed more normal function and activity than those from human fat.  In fact, not only were the protease activities of the proteasomes from the aging stem cells decreased, but they also displayed structural alterations.

These differences in proteasomal activity were not only reproducible, but when the proteasome of young stem cells were compromised, the “stemness,” or capacity of the stem cells to act as undifferentiated cells, was negatively affected.

Even more surprisingly, once after mesenchymal stem cells from human donors lost their ability to proliferate and act as stem cells (their stemness, that is) their decline could be counteracted by artificially activating their proteasomes. Activating the proteasome seems to help the cell “clean house,” get rid of junk proteins, and rejuvenate themselves.

proteasomes-and-stem-cells

Gonos and his team found that the stem cell-specific protein, Oct4, binds to the promoter region of the genes that encode the β2 and β5 proteasome subunits. Oct4 might very well regulate the expression of these proteasome-specific genes.

From this paper, it seems that a better understanding the mechanisms regulating protein turnover in stem cells might bring forth a way to stem cell-based interventions that can improve health during old age and lifespan.

This paper was published in Free Radical Biology and Medicine, Volume 103, February 2017, Pages 226–235.

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.

Nanog Gene Reverses Aging in Adult Stem Cells


Professor Stelios Andreadis from SUNY Buffalo and his colleagues have, in a series of elegant experiments, shown that the gene Nanog can stimulate dormant cellular processes that seem to be vital for preventing weak bones, clogged arteries and other telltale signs of aging. The findings might help counteract premature aging disorders such as Hutchinson-Gilford progeria syndrome.

“Our research into Nanog is helping us to better understand the process of aging and ultimately how to reverse it,” said Andreadis.

In order to delay or even reverse the ravages of aging, the human body holds a reservoir of nonspecialized progenitor cells that can regenerate organs. These cells are collectively called “adult stem cells,” and they are in every tissue of the body. Adult stem cells can rapidly respond to tissue damage to regenerate and heal organs and tissues. Unfortunately, as people age, fewer adult stem cells pare able to properly perform their function. This leads to the clinical scenarios associated with aging. Reversing the effects of aging in adult stem cells – re-booting them if you will – can potentially overcome this problem.

Andreadis and his coworkers have previously shown that the capacity of adult stem cells to form muscle and generate force declines with age. Specifically, Andreadis and others examined smooth muscle cells found in arteries, intestines and other tissues. In this new study, grad student Panagiotis Mistriotis introduced a gene called Nanog into aged stem cells. He found that Nanog activated two key cellular pathways that include Rho-associated protein kinase (ROCK) and Transforming growth factor beta (TGF-β). Activation of these two signaling pathways awakens dormant proteins like actin to build the new cytoskeletal networks that adult stem cells need to form contracting muscle cells. Force generated by these cells ultimately helps restore the regenerative properties that adult stem cells lose due to aging.

“Not only does Nanog have the capacity to delay aging, it has the potential in some cases to reverse it,” said Andreadis, who noted that introduction of the Nanog gene worked in three different models of aging: cells isolated from aged donors, cells aged in culture, and cells isolated from patients with Hutchinson-Gilford progeria syndrome.

Additionally, Andreadis and his group found that Nanog activated the central regulator of muscle formation, a signaling protein called serum response factor (SRF), which suggests that the same results may be applicable for skeletal, cardiac and other muscle types.

Andreadis and others are now examining potential drugs that can replace or mimic the effects of the Nanog gene. This will allow them to study the consequences of aging inside the body can also be reversed. This could have implications in an array of illnesses, everything from atherosclerosis, high blood pressure, and osteoporosis to Alzheimer’s disease.

This fascinating paper was published here: Panagiotis Mistriotis et al., “NANOG Reverses the Myogenic Differentiation Potential of Senescent Stem Cells by Restoring ACTIN Filamentous Organization and SRF-Dependent Gene Expression,” Stem Cells, 2016; DOI: 10.1002/stem.2452.

RepliCel Skin Rejuvenation and Tendon Repair Trials With Hair Follicle Stem Cells Underway


RepliCel Life Sciences has enrolled subjects for their skin rejuvenation and tendon repair trial.  The primary goal of these trials is to determine the safety of their cell therapeutic products.

NBDS-Cells-Follicle

In the first trial will test a product called RCS-01, which consists of cells derived from non-bulbar dermal sheath (NBDS) cells, which are taken from the outer regions of hair follicles.  NBDS cells express type 1 collagen, a protein that is steadily degraded in aged skin (hence the formation of wrinkles).  Therefore, RepliCel is confident that RCS-01 injections underneath the skin has the potential to rejuvenate aged or damaged skin.  The trial will examine male and female subjects, between 50-65 years old, and will address the inherent deficit of active fibroblasts required for the production of type 1 collagen, elastin and other critical extracellular dermal matrix components found in youthful skin. The trial will be conducted at the IUF Leibniz-Institut für umweltmedizinische Forschung GmbH in Dusseldorf, Germany.  Originally, RepliCel wished to enrolled 15 men and 15 women, but the large number of female subjects and paucity of men persuaded the company to move forward with the trial despite only enrolling a few men and all the projected women.

The second trial will test the safety and efficacy of RCT-01 in the repair of damaged Achilles tendons.  RCT-01 also consists of NBDS cells and this trial is a phase 1/2 clinical trial that examines the ability of NBDS cells to treat chronic tendinosis caused by acute and chronic tensile overuse.  This trial will take place at the University of British Columbia in Vancouver, BC, and will only treat 10 subjects.  Even though RepliCell wishes to originally test 28 participants, the company shorted the trial in order to have safety data by the end of 2016.

Darrell Panich, RepliCel Vice President of Clinical Affairs, said that the company had a late start on its trials, and therefore truncated the recruitment process in order to have safety data for analysis by the end of 2016.  Despite the small size of these trials (and they are small), the company is hopeful that their safety data will provide the impetus for moving forward with larger phase 2 trials.

Panich said, “We have adjusted our plans for the RCT-01 clinical trial in part because it started later in 2015 and enrolled slower than originally anticipated. While the trial did not meet projected enrollment targets, we are confident the safety and preliminary efficacy data obtained by year-end will provide a signal of the product’s potential to regenerate chronically injured tendon that has failed to respond to other treatments. This will allow our teams to effectively plan larger phase 2 trials in 2017 which are powered to be statistically significant for clinical efficacy (evidence the product works as intended).”

“Future trials involving products from our non-bulbar dermal sheath (NBDS) platform will be designed to investigate the efficacy of these products at different dose levels and treatment frequencies while continuing to collect other data that will be used to support eventual RCS-01 and RCT-01 marketing applications by our commercial partners.”

“The delivery of clinical data when promised is important to management”, said R. Lee Buckler, President & CEO, RepliCel Life Sciences Inc. “We have made critical decisions to keep our commitment to the financial community and we believe the data from these trials will facilitate us closing a licensing and co-development deal on one or both of these products similar to the kind we have in place with Shiseido Company for our RCH-01 product,” he added.

RepliCel is confident that their NBDS fibroblast platform will address numerous indications where impaired tissue healing has been stalled due to a paucity of active fibroblasts, which are required for tissue remodeling and repair.  NBDS fibroblasts, isolated from the hair follicles of healthy individuals, are a rich source of fibroblasts and are unique in their ability to express high levels of type 1 collagen and elastin to push-start the healing process.

RepliCel is also developing products from this same platform to address larger commercial markets in the areas of musculoskeletal and skin-related conditions.

Killing off Senescent Cells Extends the Life and Improves the Health of Laboratory Mice


Our bodies contain a mixture of cells that grow and others whose growing days are long since over. Such cells are called “senescent cells” and there is some indication that accumulations of worn out cells that have given up the ghost can contribute to the onset of age-related diseases.

A new study from the Mayo Clinic in Rochester, Minnesota has shown that eliminating senescent cells can extend the healthy lives of lab mice. These results constitute one of the first direct demonstrations that treatments that specifically target these deadbeat cells, either by killing them off of blocking their deleterious effects, might provide a new strategy to combat age-related diseases in human patients.

Aging is a fact of life for animals. Aging bodies contain cells that have lost the ability to divide. These senescent cells build up throughout our bodies and release molecules that can potentially harm nearby tissues. Diseases of advanced age, like type 2 diabetes, kidney failure, and heart disease, have been linked to the presence of large numbers of senescent cells.

Two Mayo Clinic molecular biologists, Darren Baker and Jan van Deursen, devised an ingenious way to test the relationship of accumulations of quiescent cells with age-related diseases. They engineered laboratory mice with a construct they called “ATTAC.” This construct expresses the FK506-binding-protein–caspase 8 (FKBP–Casp8) fusion protein and green fluorescent protein (GFP) under the control of a promoter that is only active when cells are senescent. The ATTAC construct is not harmful to cells, but if ATTAC-containing cells that have become senescent are hit with a drug called AP20187, they die. Baker and van Deursen injected one-year-old mice with AP20187 twice a week starting at one year of age. They injected a control group of mice with buffer only.

The AP20187 treatments extended the median lifespan in both male and female mice. To make sure that nothing strange was going on with the genetic backgrounds of these mice, they conducted these experiments in mice strains with two distinct genetic backgrounds, but the results were the same. The engineered mice that were injected with AP20187 showed consistent clearance of quiescent cells and extended, healthier life spans.

The AP20187-injected mice showed clearance of fat cells, their kidneys functioned at a higher level, and their hearts were more resilient to stress. These same mice also showed more energetic behaviors, since they explores their cages more and they developed cancers at later ages. There was also evidence of less inflammation in the AP20187-injected mice.  These mice had their lifespans extended by 20–30%. These results were published in the journal Nature on February 3rd of 2016.

Despite the rather fancy genetics involved with this experiment, the design is somewhat easy to follow and the results have a ring of credibility to them. “We think these [quiescent] cells are bad when they accumulate. We remove them and see the consequences,” says Baker. “That’s how I try to explain it to my kids.”

In 2011, Baker and van Deusen and others investigated mice that harbor a mutation that greatly accelerates aging. This mutation mimics the human genetic disease “progeria,” which is a rare, fatal genetic condition characterized by accelerated aging in children. The name of the condition, progeria, comes from Greek and means “prematurely old.” Classic progeria is also called “Hutchinson-Gilford Progeria Syndrome,” which was named after Dr. Jonathan Hutchinson and Dr. Hastings Gilford, who first described it.

In their 2011 Nature paper, Baker and others showed that removing senescent cells in mice with an engineered type of progeria benefited those mice. Therefore, this 2016 paper is a follow-up to that 2011 study.

On the coattails of these experiments, Baker, van Deusen and others in their laboratories are beating the bushes for drugs that can directly eliminate senescent cells, or, at least, stop them from secreting the damaging factors that do so much damage to nearby tissues. In fact, van Deursen has co-founded a company that has licensed patents to develop such drugs.

According to Dominic Withers, a clinician-scientist who studies ageing at Imperial College London, the experiments and Baker, van Deusen and their colleagues, “gives you confidence that senescent cells are an important target.” Withers also said: “I think that there is every chance this will be a viable therapeutic option.”