Osteoarthritis (OA) is a common degenerative process that is a major health problem in the US population. While it is known that muscle strength and exercise can modulate OA symptoms, the mechanism by which skeletal muscle alters OA pathogenesis is only partly elucidated. The conventional thought is that muscle activity directly affects the joint biomechanically by modulating joint loading and altering bone strength, attenuating OA severity. However, muscle also secretes factors that can have paracrine effects. How such factors affect the joint and how they change with exercise and in muscle pathologies is not known. Using cell culture studies, we found that human muscle cells exercised in culture produce secreted factors that alter the expression of important genes in OA severity in human articular explants. We then modified an approach that allows circulatory exchange between two mice using catheters rather than a standard parabiosis technique. This approach allows the study of the role of circulating factors in one animal on the severity of OA in another animal, while the animals have different exercise activity levels. Our preliminary data shows the downregulation of genes important in OA severity in a sedentary animal when its circulation is shared with an exercised animal. Here we will develop approaches to identify these factors and will characterize an in-vivo strategy to test the function of such factors in rodents. We will undertake the following aims: Building on our data showing that conditioned media from exercised muscle produces factors that inhibit expression of genes important in OA severity in osteoarthritic cartilage explants, we will further evaluate these paracrine effects for different exercise regimes and ages of the muscle cells. To identify differentially secreted proteins, we will use mass spectroscopy secretome analysis. Using our modified catheters based approach to exchange circulating blood, allowing animals to share a circulation while undertaking different physical activity regimens, we will determine how the duration of exercise, number of blood exchange procedures, and plasma or cell fractions alter the OA phenotype associated with a surgical joint injury. Expression of the most differentially regulated proteins by exercise identified in the first aim will be compared between the serum from exercised and quiescent mice using ELISA. Identified proteins could be targeted pharmacologically and as such, our data may identify a therapeutic approach that could be used to attenuate the severity of OA. Furthermore, this work will define the specific contribution of biochemical effects of exercise on OA severity.