APJ Receptor Activation Clears Damaged Mitochondria to Halt Inflammation-Driven Bone Loss

Inflammatory conditions including rheumatoid arthritis, diabetes, and inflammatory bowel disease disrupt the bone immune microenvironment, triggering osteoclast overactivation and progressive bone loss. Current anti-inflammatory treatments such as glucocorticoids paradoxically worsen bone loss, leaving a significant unmet therapeutic need. This study investigated whether the Apelin receptor (APJ), a G-protein-coupled receptor with known roles in cardiovascular and neurological inflammation, could regulate the immune-bone axis in systemic inflammatory bone loss.

Using an LPS-induced systemic inflammatory bone loss mouse model, the authors divided C57BL/6 mice into sham, LPS, and LPS + Apelin-13 groups. Apelin-13 (the most bioactive APJ ligand isoform) was administered at 100 µg/kg intraperitoneally for seven days. Micro-CT analysis demonstrated that Apelin-13 treatment significantly preserved trabecular bone parameters. TRAP staining and histological analysis confirmed reduced osteoclast numbers. Importantly, macrophage M1 polarization markers (iNOS, CD86) were suppressed in treated animals, as measured by flow cytometry and immunohistochemistry.

In vitro mechanistic work used bone marrow-derived macrophages (BMDMs) stimulated with LPS, with APJ activity manipulated by Apelin-13 treatment or siRNA-mediated APJ knockdown. High-throughput RNA sequencing of these cells identified mitochondrial autophagy and the NOD-like receptor signaling pathway as the primary pathways regulated downstream of APJ, alongside AMPK and MAPK signal transduction cascades. Subsequent experiments confirmed that APJ activation upregulated the AMPK/BNIP3/PINK1/PARKIN mitophagy axis. Transmission electron microscopy revealed increased autophagic structures and improved mitochondrial morphology. JC-1 and MitoTracker assays showed restored mitochondrial membrane potential, while DHE and DCF-DA flow cytometry demonstrated a marked reduction in intracellular ROS.

By clearing dysfunctional mitochondria and reducing ROS accumulation, APJ activation suppressed NLRP3 inflammasome assembly (reduced NLRP3, ASC, and IL-1β levels), thereby blocking M1 macrophage polarization. In turn, the inflammatory cytokine milieu driving osteoclastogenesis was curtailed, and RANKL-induced osteoclast differentiation was significantly impaired. APJ knockdown by siRNA reversed these protective effects, confirming the receptor's necessity in the pathway.

These findings establish a mechanistic chain from APJ activation → AMPK phosphorylation → BNIP3 upregulation → PINK1/PARKIN-dependent mitophagy → ROS clearance → NLRP3 suppression → reduced M1 polarization → decreased osteoclastogenesis → bone protection. The study suggests APJ agonism as a novel therapeutic strategy that could simultaneously address inflammation and bone loss, avoiding the skeletal side effects of glucocorticoids.