Researchers in the United States and Japan have discovered a new mechanism that links age-related cartilage tissue stiffening with the repression of a key protein associated with longevity, and the development of age-related knee osteoarthritis (KOA). The study results showed that increased stiffening of the extracellular matrix (ECM)—a network of proteins and other molecules that surround and support tissues in the body—led to a decrease in a “longevity protein” called Klotho (α-Klotho) in knee cartilage, brought about by epigenetic changes. This decrease in Klotho resulted in chondrocyte damage in healthy cartilage.

Stiffening of the extracellular matrix is a defining feature of cartilage aging, and the study findings provide new insights into the mechanisms that lead to the deterioration of joints and thus osteoarthritis, demonstrating the role Klotho plays, and offering up potential new potential treatment targets to restore cartilage health. The researchers noted that their results may provide new insights into understanding the toll that epigenetic factors caused by aging take on other tissues throughout the body. The results could also point to future therapeutic strategies.

“This research enhances our mechanistic understanding of why osteoarthritis happens in the first place, and it paves the way for the development of therapeutics to prevent these changes,” said Fabrisia Ambrosio, PhD, MPT, inaugural director of the Atlantic Charter Discovery Center for Musculoskeletal Recovery of the Schoen Adams Research Institute at Spaulding Rehabilitation Network, and member of the faculty of Physical Medicine and Rehabilitation at Harvard Medical School. “Such therapeutics are important because there are currently no disease-modifying treatments for osteoarthritis; the best we can do for now is minimize pain and disability. Since matrix stiffening is a feature of aged tissues throughout the body, we anticipate that these findings may also have implications beyond cartilage repair for the field of aging research.”

Ambrosio, colleagues at the University of Pittsburgh, at the Japan Society for the Promotion of Sciences, and collaborators, reported on their work in Nature Communications, in a paper titled, “Age-related matrix stiffening epigenetically regulates α-Klotho expression and compromises chondrocyte integrity,” in which they concluded, “These findings are likely to have broad implications even beyond cartilage for the field of aging research.”

Osteoarthritis occurs when cartilage in a joint stiffens and begins to break down, which then damages the underlying bone, resulting in pain, swelling, and feelings of stiffness. Osteoarthritis is the most common form of arthritis, affecting approximately 32.5 million individuals in the United States. Rates are expected to rise with the aging population and trends in obesity. Osteoarthritis can significantly interfere with a person’s ability to perform routine daily tasks, and about half of adults with the condition are of working age, so it can impact their ability to earn a living.

There are currently no treatments to reverse cartilage stiffening and resulting damage. While treatments such as exercise, weight loss, physical therapy, medications, injections, and joint replacement surgery aim to reduce pain and improve mobility, much is still unknown about the molecular causes of this damage and how to treat it. These unknowns are especially germane to knee osteoarthritis, where no single event causes the cartilage damage, and the greatest predictive risk factor is aging.“Extracellular matrix stiffening is a quintessential feature of cartilage aging, a leading cause of knee osteoarthritis,” the authors wrote. “Yet, the downstream molecular and cellular consequences of age-related biophysical alterations are poorly understood … Although we have known of the poor healing capacity of cartilage for over 250 years, this limited capacity is both poorly understood and inadequately treated in the clinic.”

Increasingly, researchers have sought to better understand the role that epigenetics, or how changes in behaviors and environment as people age alter how genes work, can impact tissues and disease processes throughout the body.

For their reported study, using advanced mass spectrometry technology, the researchers mapped out the trajectory of structural and protein changes in mice with KOA over the course of their lifetimes, and according to sex. They then compared their findings to the current understanding of knee osteoarthritis in humans. “We next examined mechanistic drivers of KOA via a series of genetic and pharmacologic manipulations,” they further explained.

Through the newly reported work, the researchers found that Klotho was heavily involved in the molecular process that led to osteoarthritis. The study was an extension of previous researchers showing that klotho protects mitochondria within skeletal muscle and plays a key role in skeletal muscle regeneration following injury. As people age, their Klotho levels go down, hence why it’s referred to as a longevity protein.

The new analysis revealed that when knee cartilage tissue became stiffer, the gene that codes for Klotho was repressed. The team verified this in models of young and old chondrocyte cells responsible for cartilage formation, which were seeded in environments designed to mimic young and old tissue stiffness. Young chondrocyte cells looked old when put on a stiff surface due to the loss of Klotho, but when the researchers protected the cells from the stiffness in their environment, they observed improved chondrocyte health. “we discovered that increased matrix stiffness drove Klotho promoter methylation, downregulated Klotho gene expression, and accelerated chondrocyte senescence in vitro,” they wrote. “In contrast, exposing aged chondrocytes to a soft matrix restored a more youthful phenotype in vitro and enhanced cartilage integrity in vivo … pharmacological reduction of cartilage stiffness in aged mice increased α-Klotho levels and restored cartilage integrity.”

“These results provide a compelling new paradigm that will be important for the field in terms of understanding the connection between age-related tissue stiffening and risk for osteoarthritis with aging,” said Hirotaka Iijima, PhD, PT, assistant professor at the Institute for Advanced Research and Graduate School of Medicine in Nagoya University. The authors further concluded, “Taken together, these findings suggest that age-related alterations in matrix stiffness initiate pathogenic mechanotransductive signaling cascades, leading to epigenetic repression of the Klotho promoter and compromised chondrocyte health.”

The study results also revealed that the incidence of osteoarthritis increased in male mice with age, while female mice showed no onset of the disease and their cartilage tissue was generally preserved. This unexpected finding differs from the response observed in humans, for which it is post-menopausal women who are significantly more likely to develop severe knee osteoarthritis than men. The findings warrant further study, according to the authors, and a project is underway in Ambrosio’s lab to examine the effects that menopause has on knee osteoarthritis at the molecular level.

Given the latest findings, the researchers plan to study whether there are ways to intervene with the disease process that leads to osteoarthritis, such as by blocking the pathway that represses Klotho, even in the face of a stiff extracellular matrix environment. They hope that their discoveries can be used to develop treatments for osteoarthritis and other conditions caused by aging. “Together, these findings suggest that preventing or reversing age-related matrix stiffening and the resulting pathogenic mechanotransductive signaling may serve as a promising therapeutic target to attenuate or even reverse the development of agerelated KOA,” the team stated.

“We’re interested in evaluating whether epigenetic regulation of Klotho and other longevity factors by the extracellular matrix may help explain functional decline of tissues throughout the system,” said Ambrosio. The authors further noted: “While this study focused on cartilage as a model, a major conceptual innovation of our work is the demonstration of the novel mechanistic link between age-related increases in ECM stiffness and epigenetic repression of α-Klotho … we anticipate that these studies may have broader implications  in the field of aging research even beyond cartilage.”

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