Retro-inverso TAT-Beclin-1 induces synovial fibrosis and does not protect cartilage from degeneration in a mouse model of OA

Abstract

Background: Beclin-1 is a component of the autophagy pathway necessary for formation of autophagosomes, contributing to autophagy-mediated cellular homeostasis. Enhancing autophagy through inhibition of mTOR activity, either via genetic deletion in chondrocytes or intra-articular injection of rapamycin, attenuates progression of surgically-induced models of osteoarthritis (OA). Retro-inverso TAT-Beclin-1 is a cell-permeable peptide which competes for binding to the endogenous Beclin-1 inhibitor GAPR-1, thus promoting autophagy. It is unknown whether activation of Beclin-1 is sufficient to protect joints from osteoarthritis progression. Objectives: For this study, we sought to determine if retro-inverso TATBeclin-1 could attenuate OA progression in a surgically-induced mouse model. Methods: Eight-week old C57BL/6 mice underwent destabilization of the medial meniscus (DMM) surgery to induce OA, or sham surgery as a control. Mice were injected intra-articularly with retro-inverso TAT-Beclin-1 (2 mg/kg in 5ml) twice weekly for 2 or 9 weeks. Mice were sacrificed at 10-weeks post-surgery. Knee joints were stained with Safranin-O/Fastgreen to evaluate cartilage degeneration and Masson’s trichrome to determine degree of synovitis using OARSI scoring for mice. Sections were stained for a-SMA (myofibroblast) and CD45 (hematopoietic-origin cell) to evaluate changes in markers of fibrosis and inflammation, respectively. Results: As opposed to the effects of mTOR deletion in cartilage or rapamycin treatment in joints, injection of retro-inverso TAT-Beclin-1 for 2 into knee joints of mice with DMM-induced OA had no effect on the degree of articular cartilage degeneration in the tibia or femur as compared to PBS-injected controls. However, in both sham and DMM mice, retro-inverso TAT-Beclin-1 for 2 or 9 weeks of treatment induced a pronounced thickening of the synovium with increased cell numbers and collagen deposition compared to PBS-treated mice. The increased number of synovial cells in 9-week treated mice did not show substantial expression of a-SMA+ or CD45+ cells, suggesting the increased number of cells and matrix in the synovium was independent of myofibroblast differentiation or inflammatory influx. Conclusion: Contradictory to our expected results, retro-inverso TATBeclin-1 did not attenuate cartilage degeneration. Rather, it promoted substantial synovial thickening that likely involved cell proliferation and collagen deposition. This severe fibrotic phenotype appears independent of myofibroblast differentiation or inflammation, normally associated with typical fibrotic responses. To evaluate the potential for dose responses with retro-inverso TAT-Beclin-1 in synovial joints, we are currently modifying our dosing strategy in an effort to determine possible disease modifying effects of this novel Beclin-1 activator. Disclosure of Interests: None declared DOI: 10.1136/annrheumdis-2019-eular.2680