The broad objective is to investigate the role of mitochondrial Damage-Associated Molecular Patterns (mDAMPs) in traumatic joint injury and the development of osteoarthritis (OA) in the horse.

There is arguably no more urgent health issue to the short- and long-term welfare of equine athletes than traumatic joint injury. Cartilage provides near-frictionless joint surfaces and cushioning to protect underlying bone. Even mild cartilage damage can impair its ability to dissipate loads, exposing bone to repeated micro-trauma, which can ultimately lead to catastrophic fracture and/or osteoarthritis. Therefore, understanding how cartilage responds to loading and perpetuates damage signals throughout the joint is critical to preventing joint injury and disease.

Mitochondria are best known as the "powerhouses" of cells, producing the energy required for normal tissue function and repair. Remarkably, these organelles also act as mechanotransducers, sensing physical forces and converting those signals into biological responses. Our recent work revealed that mitochondrial dysfunction is one of the earliest responses of cartilage to overloading. In other tissues, injury-induced mitochondrial dysfunction causes cells to release danger signals, or mDAMPs (mitochondrial Damage Associated Molecular Patterns). These mDAMPs act as molecular triggers, inducing inflammation and perpetuating tissue damage. However, the role of mDAMPs has not been investigated in joint trauma or osteoarthritis.

Broadly, our goal is to investigate the types of signals that initiate extracellular mDAMP release by cartilage. More specifically, Aim 1a will test the hypothesis that mitochondrial dysfunction triggers mDAMP release. Chondrocytes grown in culture will be stressed with several compounds, including a general inflammatory stimulus (IL-1β), an oxidant (hydrogen peroxide), and three specific inducers of mitochondrial dysfunction (oligomycin, rotenone, FCCP). Culture media will be analyzed for three mDAMPs; 1) mitochondrial DNA, 2) the mitochondrial protein cytochrome C, and 3) the mitochondria-specific phospholipid cardiolipin. Aim 1b will test the hypothesis that mDAMPs are released from cartilage in response to mechanical overloading. Our previous work revealed that mitochondrial dysfunction is an immediate response of cartilage to impact-injury. In that study, live cartilage explants were injured, maintained in culture for one week, and media was harvested and frozen. This banked media will be analyzed for mDAMPs, as described above. Results of Aim 1 will provide insight into how mDAMP release is triggered, and therefore how it may be manipulated therapeutically.

These studies will provide the foundation for a new area of research. The next step will be to investigate mDAMPs in vivo; we plan to measure mDAMPs in banked joint fluid from horses with experimental joint trauma, as well as equine patients with naturally-occurring joint injury. The ultimate goal is to determine if mDAMPs in joint fluid are a useful marker of early articular injury, and develop targeted therapies to halt ongoing inflammation and joint destruction after trauma.