PTH treatment prior to cyclic joint loading improves cartilage health and attenuates load-induced osteoarthritis development in mice
Osteoarthritis (OA) treatment is limited by the lack of effective non-surgical interventions to slow disease progression. Here, we examined the contributions of the subchondral bone properties to OA development. We used parathyroid hormone (PTH) to modulate bone mass prior to OA initiation and alendronate (ALN) to inhibit bone remodeling during OA progression. We examined the spatiotemporal progression of joint damage by combining histopathological and transcriptomic analyses across joint tissues. The additive effect of PTH pretreatment prior to OA initiation and ALN treatment during OA progression most effectively attenuated load-induced OA pathology. Individually, PTH directly improved cartilage health and slowed the development of cartilage damage, whereas ALN primarily attenuated subchondral bone changes associated with OA progression. Joint damage reflected early transcriptomic changes. With both treatments the structural changes were associated with early modulation of immunoregulation and -response pathways that may contribute to disease mechanisms. Overall, our results demonstrate the potential of subchondral bone-modifying therapies to slow the progression of OA. Overall design: Parathyroid hormone (PTH (1-34), 40mg/kg/day, SQ (66)) or vehicle (VEH, saline, same volume, SQ) pretreatment was administered for 8 weeks (5 days/week) to 18-week-old male C57BL/6J mice (202 animals total). Following pretreatment, cyclic mechanical loading was applied to the left tibia at a peak load of 9N for 0 (pretreatment only, no loading, Fig.1A), 1, 2, or 6 weeks (Fig.2A) (8, 29, 30). Concurrent with loading, pretreated mice received either alendronate (73mg/kg/day, intraperitoneal (18)) or vehicle (saline, same volume, intraperitoneal) treatment (5 days/week, Fig.2A). Mice were placed under general anesthesia during loading (2% isoflurane, 1.0L/min, Webster). Loading was applied to the left tibiae at 4Hz for 1200 cycles (5 min) per day, 5 days per week (8, 29, 30). The right tibiae served as contralateral controls. Following completion of the loading periods, mice were euthanized. In a subset of mice following 1, 2, and 6 weeks of loading, knee joints were harvested and fixed in 4% paraformaldehyde (PFA) overnight at 4° C for tissue level analysis (n=7-8/group). In the pretreatment only group, right limbs were harvested and fixed in 4% paraformaldehyde overnight at 4° C, and left limbs were frozen in PBS-soaked gauze immediately following dissection for nanoindentation analysis. In a separate group of mice following 0, 1, or 2 weeks of loading, different tissues (cartilage, inguinal lymph nodes, and metaphyseal cancellous and cortical bone, after bone marrow removal) were retrieved. Cartilage samples were stabilized in RNAlater while inguinal lymph node and bone samples were snap frozen in liquid nitrogen for for RNA isolation and bulk RNA sequencing analyses (RNA-seq, n=4-8/group). RNA was extracted from tibial cartilage, metaphyseal cortical bone, cancellous bone and inguinal lymph node tissues using protocols previously described (67, 68) (n=4-8/ea). Samples were not pooled. All experimental procedures were approved by the Institutional Animal Care and Use Committee at Cornell University and by the USAMRMC Animal Care and Use Review Office, and are reported following ARRIVE guidelines (69) and best practices to implement and report experiments using the tibial loading model (70).