The role of mechanosensing and mechanotransduction in joint homeostasis and osteoarthritis susceptibility - SUMMARY Osteoarthritis (OA) is a debilitating disease characterized by loss of joint space, degeneration of cartilage at articular surfaces, remodeling of bone and other joint tissues, and inflammation. Although it is the major cause of disability in the aged population, not a single disease modifying drug is available. The functions and identities of the biological processes that pose vulnerabilities to OA are unknown. We have identified a novel association between OA and PIEZO1 activity exhibiting heightened susceptibility to age-associated OA. Our analyses of 151 families with dominantly inherited OA identified four independent OA-susceptibility alleles affecting the mechanosensitive cation channel, PIEZO1. An independent indication that PIEZO1 is critically involved in OA comes from a recent genome wide association study (GWAS) that identified a dominant rare coding allele of PIEZO1 associated with reduced OA progression. We do not know how these mutations affect PIEZO1 activity and OA susceptibility. PIEZO1 responds to mechanical cues to regulate diverse biological processes in maintaining musculoskeletal homeostasis. However, its role in OA susceptibility is unresolved. Here we combine single channel biophysical analyses with cellular and molecular studies using mice harboring gain- and loss- of function human PIEZO1 mutations to determine the role of PIEZO1 in joint homeostasis and OA. Our preliminary electrophysiological studies indicate that all four familial OA alleles are hypomorphic, reducing the open probability of the channel in response to mechanical stimulus, while the GWAS allele is hypermorphic. Mice carrying the dominant familial OA-associated Piezo11398W allele, introduced by genome editing, have significantly increased inflammatory gene expression in the knee joint. We hypothesize a reduction in Piezo1 activity is a central component of the homeostatic signaling networks and cell processes that maintain the synovial joint. In Aim 1 we identify how mutations in PIEZO1 alter single channel activity, trafficking, and cellular Ca2+ levels in chondrocytes expressing WT and mutant proteins. In Aim 2 we test the hypothesis that mice expressing Piezo11398W will have reduced susceptibly to injury induced OA while Piezo12484L will accelerate OA. Mouse models of age-dependent OA are rare and changes that indicate early stages of OA are unknown. Our preliminary data indicate that PIEZO1 has a different role in injury- vs age-associated OA. Therefore, it is crucial to define the role of PIEZO1 during aging and establish aging models of OA. In Aim 3 we establish i) that reduced Piezo11398W activity promotes OA-associated gene expression in multiple joints during aging, ii) and causes accelerated onset of histologically recognizable age-dependent OA in the joints that are correlated with use/weight-bearing activity. RNA-seq analyses will be used to uncover changes in gene expression and cell populations in the joint that tracks development of OA. Our work will have direct clinical impact, informing efforts to identify biomarkers of susceptibility or early stages of OA, which in turn help develop better therapies for OA.
