Human cartilage-bone-synovium microphysiological system to study ptoa pathogenesis and treatment on earth and in space

Abstract

Purpose: Acute joint injuries from sports, exercise or accident-related trauma often progress to post-traumatic Osteoarthritis (PTOA), comprising ∼12% of all OA cases. Such joint injuries can cause mechanical damage to cartilage combined with an inflammatory response associated with the synovial lining of the joint capsule. Together, these events lead to cartilage and subchondral bone pathologies including cell death, tissue degradation, neoangiogenesis, osteophyte and cyst formation and synovial fibrosis. Astronauts on International Space Station-National Laboratory (ISS-NL) sustain a higher rate of exercise-related injuries, including those compromising joints, during their training and mission period. Acknowledging the urgent need to identify disease-modifying therapeutics to ameliorate degenerative evolution of OA/PTOA, we carried out this study to (1) simulate aspects of acute joint injury biology on earth and in space using a human Cartilage-Bone-Synovium coculture model (CBS-MPS), (2) to investigate the potential of dexamethasone (Dex) in reducing cell death and ECM degradation, and (3) to utilize a Multi-Use Variable-g Platform (MVP) to study PTOA-related pathogenesis and management on the ISS. Methods: Osteochondral plugs (3.5mm diameter and 5mm thick, including 1.5-2mm thick cartilage) harvested from condyles and trochlear grooves of cadaveric human distal femurs (n=8 knees, Collins grade 1/2, 64-82 yr) were co-cultured with 4-5mm diameter synovium explants harvested from synovial joint capsule of the same knees. All procedures were approved by the Rush University Medical Center Institutional Review Board and the Committee on the use of Humans as Experimental Subjects at MIT. PTOA-like mechanical trauma was simulated by subjecting the cartilage surface of the osteochondral plugs to a single injurious unconfined compression of 60% strain rate to a peak stress of 5MPa using an incubator-housed loading apparatus. The study design included the following groups: (1) Mechanical injury (INJ) - injured osteochondral plugs were cultured in presence of pro-inflammatory cytokines (including TNF-α, IL-6 and IL-8) that were released by synovial explants and quantified by multiplex-readout in situ, (2) Treatment group (INJ + DEX): 100nM Dex was added to the cocultures of injured osteochondral plugs and synovial tissues immediately after injury. A preliminary science validation test was performed to identify a suitable mechanism to adapt CBS cocultures to the custom experiment module and the MVP facility. Engineering validation testing was then carried out to build logistical protocols for the spaceflight including transport of cultures and simulation of tasks preceding launch operations. Results: The levels of pro-inflammatory cytokines such as IL-6, IL-8 and TNF-α were upregulated in CBS cocultures. The combination of mechanical injury and increased release of inflammatory cytokines from synovial tissues caused extensive cell death and biochemical changes in both bone and cartilage. In the INJ group, cartilage tissue showed widespread cell death [Fig1 a], and subchondral bone showed increased cell death as well, indicated by numerous empty lacunae in the matrix. Biochemical analysis showed aggravated release of matrix component (including GAGs) into the medium, associated with cartilage degradation [Fig 1c]. Addition of Dex significantly reduced the injury-induced cell death [Fig 1b] and GAG loss by week 2 [Fig 1c]. For spaceflight preparation, tissue cards [Fig 1d] were optimized for long-term holding and maintenance of CBS co-cultures, which were placed into custom culture chambers [Fig 1e]. Multiple tests using this instrumentation were used to guide the design and operation of automated MVP facility [Fig 1f]. Investigation of disease biology and acute injury management is currently being tested in preparation for eventual spaceflight experiments in microgravity aboard ISS-NL. Conclusions: By providing a platform to study cartilage, bone and synovium together, our CBS MPS can potentially enable a more physiologically relevant understanding of tissue interactions that may lead to PTOA pathogenesis. This system can also be used to test the efficacy of therapeutics. In Space, this MPS will be used to explore the role of therapeutic agents to ameliorate cartilage degradation in microgravity to support astronaut health during long space missions.