Research Project
10489981
Project Summary A key priority for the NIH is to limit disability caused by osteoarthritis (OA) and other chronic diseases that emerge with age. However, there is a knowledge gap regarding the mechanisms by which aging drives OA progression, which has limited the options for effective early intervention. One emerging strategy is to initiate apoptosis in senescent chondrocytes as a way to prevent the secretion of catabolic factors. The long-term goal of this work is to catalyze more effective treatments for OA by determining when and why chondrocytes become senescent. The objectives are to quantify the senescence burden, examine the causal role of cell division in senescence induction, and determine the effect of senolytic clearance in aging mice. Our central hypothesis is that cell division during the early phases of OA induces senescence in chondrocytes, and that the accumulation of senescent chondrocytes is a driver of OA progression during aging. Our approach uses p16tdTom senescence reporter mice to enable the flow cytometry analysis and physical separation of senescent chondrocytes. Further, we use a complementary set of iterative immunofluorescence, live-cell imaging, and computational techniques to interrogate the mechanisms of senescence induction in primary human chondrocytes. The first aim is to determine the temporal relationship between chondrocyte senescence and age-related OA, with the hypothesis that senescence emerges as a response to cartilage degradation in early OA. RNA-sequencing on chondrocytes sorted as p16tdTom-positive and p16tdTom-negative will be used to analyze changes to the senescent phenotype during aging and OA progression. The second aim is to determine the mechanisms by which mitogenic stimulation enhances senescence induction, with the hypothesis that chondrocytes harboring DNA damage will initiate senescence upon cell division through an extended duration of p38 activity. The dynamics of p38 within asynchronously dividing cell populations will be related to a final cellular state that is determined by a multi-dimensional protein signature. The third aim is to determine the efficiency of senescent cell clearance and the extent of protection from age-related OA, with the hypothesis that repeated intra-articular injection of navitoclax will mitigate OA progression. To enhance the dynamic range for assessing clearance and the protection from OA, a Jnk2 knockout genetic background with enhanced senescence and age-related OA will be used. The expected outcomes of this work include a better understanding of the timecourse of chondrocyte senescence, the role of cell division as a senescence trigger, and the extent to which removal of senescent cells alters age-related OA. This work is innovative in that it utilizes a sophisticated cohort of mice to isolate a well-defined population of senescent chondrocytes, analyzes senescence induction at the single-cell level, and employs a genetic background with enhanced senescence and OA for clearance studies. These contributions are expected to have a positive impact on society by stimulating more effective strategies to target senescent cells for the prevention and treatment of OA.
