April 15, 2017 (Vol. 37, No. 8)

Looking to Epigenetics and the Hydra to Facilitate Healing and Slow Aging

Abnormal tissue repair has a serious impact on the health and quality of life of those affected. It also incurs healthcare costs reaching tens of billions of dollars per year in the U.S., with similar costs in Europe.

Both insufficient wound healing and excessive fibrosis or scarring occur when the normal tissue-repair mechanisms present in an organism are malfunctioning. Recent advances suggest that environmental factors can influence tissue repair and regeneration by triggering epigenetic changes that influence the way cells that are responsible for wound healing behave.

The environment also impacts on tissue repair via mechanical factors, often overlooked in regenerative medicine. These factors are of key importance when considering the design of new therapeutics and also in building functional tissue equivalents such as organoids. In a similar fashion, understanding the role of the different cell types that are involved with tissue repair (for example, in the connective stroma) can give valuable insights to researchers.

While regeneration still occurs in some tissues, humans lost a large amount of regenerative ability during evolution. In contrast, some simple organisms are still able to regenerate whole organs. Age has a strong influence on tissue repair and cellular regeneration, and model organisms with high regenerative ability are being used to discover how tissue regeneration changes during the aging process.

These and other strategies to turn abnormal tissue repair into faultless regeneration will be discussed at Tissue Repair and Regeneration, a Gordon Research Conference to be held June 4–9 in New London, NH.

Epigenetics is a critical factor in the context of tissue repair and regeneration as it is involved in all aspects of this process. [Wavebreakmedia/Getty Images]

Impact of Epigenetics

It is important to consider epigenetics in the context of tissue repair and regeneration, as it is involved in all aspects of this process. “Environmental factors that influence tissue repair and regeneration operate by triggering epigenetic changes that determine the activities of cells that are responsible for wound healing,” explains Derek Mann, Ph.D., professor of hepatology at Newcastle University. “As an example, alcohol can induce epigenetic changes in wound-healing cells in the liver that promote excessive scar formation.

“In addition, the process of generating wound-healing myofibroblasts is under the control of epigenetic mechanisms that determine the phenotype, activities, and fate of these cells.”

Dr. Mann and his team have a focus on developing new therapies for liver fibrosis. They demonstrated that epigenetic regulator proteins, such as MECP2, EZH2, and ASH1, change gene expression at the chromatin level and promote fibrosis by stimulating the myofibroblast cells to produce collagen. If these proteins are “turned off,” then it is possible to stop progression of fibrosis in the diseased liver.

At present, the main method of confirming liver fibrosis is through invasive liver biopsy. However, Dr. Mann and colleagues showed that quantification of DNA methylation in circulating cell-free DNA can provide a surrogate biomarker for liver fibrosis.

“We are anticipating that our continuing work on epigenetics in fibrosis will lead to the development of new biomarkers that are diagnostic and prognostic for fibrosis progression in patients with chronic liver disease,” says Dr. Mann.

Elisabeth Zeisberg, M.D., Ph.D., a group leader at the Medical University of Göttingen, also works on the epigenetics of tissue fibrosis. “Just like in cancer,” she says, “where tumor-suppressor genes have been studied for decades, we now understand that there are also ‘fibrosis-suppressor genes’ which can be silenced or reactivated by epigenetic mechanisms.”

Dr. Zeisberg and co-researchers have identified a gene called RASAL1, which when silenced through epigenetic control contributes to kidney, heart, and liver fibrosis. They found that reactivating this gene can stop the fibrotic process.

Repurposing of drugs already approved for one condition to treat another is being investigated in a number of clinical areas. Dr. Zeisberg and her team discovered that the antihypertensive drug hydralazine, used to lower blood pressure in pregnant women, can improve outcomes in experimental models of kidney and heart fibrosis by activating genes involved with fibrosis.

“Due to its safeness, it provides an attractive opportunity for clinical translation,” states Dr. Zeisberg. “We are currently planning a prospective clinical trial to test its effectiveness in treating heart fibrosis.”

Both Dr. Mann and Dr. Zeisberg are searching for DNA methylation-based biomarkers that can help predict who will develop fibrosis, as well as indicate a patient’s prognosis once the condition is detected.

“This is important since fibrosis is a silent pathology that often only presents at a late stage where treatments may not be possible,” insists Dr. Mann. “It would provide greatly improved personal clinical management for patients and enable selection of patients for future clinical trials with antifibrotic drugs.”

Dr. Zeisberg explains that her team is investigating the potential for developing biomarker-stratified, targeted therapies. “Our main focus,” she elaborates, “is to develop Cas9-directed enzymes to specifically de-methylate select genes.”

Researchers at the Medical University of Göttingen have identified an epigenetic mechanism, transcriptional suppression of RASAL1 through aberrant promoter methylation, that contributes to cardiac fibrosis. The aberrant methylation can be reversed through Tet3-mediated hydroxymethylation. Here, confocal microscopy shows the expression pattern of DNA methyltransferase (DNMT) and Tet3 (TET) in cardiac endothelial cells.

Importance of Mechanics and Positioning

While epigenetic changes tend to occur in response to biochemical triggers, a healthy physical environment is equally important for healthy tissue repair and regeneration.

“People have learned in the last 20 years that cells do much more than just respond to biochemical factors. Instead, it is well documented today that the way in which cells anchor themselves to their surroundings and what they feel when they pull on those anchors has an impact on gene expression,” comments Viola Vogel, Ph.D., professor and deputy chair of the department of health science and technology, and head of the Laboratory of Applied Mechanobiology at ETH Zürich.

She believes that a lot of tissue-engineering research has been simplistic and has yet to adequately address the forces by which cells stretch the surrounding tissue fibers, forces that alter the biochemical display of these fibers, which then in a feedback loop alter cell and tissue functions.

However, “the feedback between engineers, biologists, and the medical community is steadily increasing. Engineers learn from biologists how to better mimic aspects of real tissue, and biologists learn from engineers how to prepare better 3D model systems to study cell function in complex environments,” she explains.

“There are no good tools to map localized forces in real tissues at high resolution; for example, to find out how much force cells generate in healthy versus diseased tissues, and how much they stretch the fibers that surround them,” notes Dr. Vogel. “Currently, we are trying to develop such probes.”

Yuval Rinkevich, Ph.D., head of a laboratory at the Helmholtz Zentrum München, also believes it is important to reflect the natural tissue environment as closely as possible: “We know a lot of what is happening in cell culture doesn’t really reflect what’s happening in vivo, and so we try to keep it as close as possible to the native settings where the cells operate.”

Dr. Rinkevich and colleagues are using dermal fibroblast lineages to study a number of cell types involved in wound repair, particularly the cells of the supportive stroma. “It’s the supportive stroma that’s primarily involved in the majority of the fibrotic or scarring outcomes that usually develop after injury or disease,” he explains.

“Something that we didn’t perhaps appreciate several years ago, is that the population of cells we collectively refer to as fibroblasts or stromal cells [consists of cells that] are very different from one another,” Dr. Rinkevich clarifies. Each of the cells within this heterogeneous group, he continues, has its own unique characteristics.

He explains that knowledge of the cells in the stromal compartment tissue lags behind that of other areas, such as the blood: “I think this is the very beginning of this field, where it is becoming apparent that there is a remarkable heterogeneity of cell types within the stroma and that it would be very exciting to learn what each one of those cell types does and how it impacts disease or health.”

Aging and Regeneration

Brigitte Galliot, M.D., Ph.D., associate professor of the department of genetics and evolution and vice-dean of the faculty of sciences at the University of Geneva, studies the freshwater Hydra polyp as a model organism with which to research regeneration and tissue repair.

The Hydra is one of a group of organisms that have strong regenerative capacity. They are also well known and widely studied for their apparent lack of aging.

Dr. Galliot and her colleagues have been investigating links between aging and regenerative capacity in these animals. “Recently, we found an animal that aged,” she says, “and one of our questions was, when they age, do they lose regenerative capacity?”

“What we notice in the animals that are aging is that they lose regeneration,” she continues. “There was a progressive, irreversible decrease of cell proliferation, and we think that this is directly due to deficient autophagy.

“There are some examples in the hematopoietic system that autophagy has an important impact on the self-renewal of stem cells; so it’s a developing idea at the moment, not only in Hydra, but in other systems, too.”

She explained that autophagy has two main advantages for cells. First, nutrients and energy are produced. Second, all the molecular aggregates—for example, the protein and lipid aggregates that are toxic for your cells—are eliminated.

Looking forward, Dr. Galliot offers the following comment: “We want to see whether Hydra would be a good model system for degenerative diseases such as Alzheimer’s disease, where cells that do not divide accumulate a lot of aggregates.”

The freshwater polyp Hydra, a champion of regeneration,is being studied by scientists at the University of Geneva. They recently found that in the Hydra, cells of the epithelial type modify their genetic program by overexpressing a series of genes, including genes that are involved in diverse nervous functions.

Environmental Impact

The environment has a key role to play in regulating both a cell’s and an organism’s ability to regenerate and to repair tissue in response to injury. Whether in the form of biochemical cues that lead to epigenetic changes, such as consumption of alcohol and liver fibrosis, or in the form of physical changes in the environment that result in physiological responses being triggered, the environment exerts a powerful effect.

While we are still in the relatively early stages of the tissue engineering revolution, it is important that these influences are taken into account to ensure the in vivo environment is accurately represented. The influences of the environment can also be seen in model organisms such as the Hydra polyp, the study of which suggests that there is a link between autophagy and the organism’s aging and regenerative abilities. It is important for the future of regenerative medicine to collect such information and put it to good use in developing future therapies that reflect the natural tissue environment as closely as possible.

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