HIF-1α facilitates osteocyte-mediated osteoclastogenesis by activating JAK2/STAT3 pathway in vitro

Osteocytes, entrapped within the mineralized bone matrix, has been found to have numerous functions such as acting as an orchestrator of bone remodeling through regulation of both osteoclast and osteoblast activity and also functioning as an endocrine cell. Due to a specialized morphology and surrounding structure, osteocytes are more tolerant to hypoxia during osteoporosis, fracture, osteoarthritis, and orthodontic-orthognathic combination therapy. Hypoxia-inducible factor-1α (HIF-1α) is one of the master regulators of hypoxia reactions, playing an important role in bone modeling, remodeling, and homeostasis. This study aimed to investigate the pivotal functional role of HIF-1α in osteocytes initiating of bone remodeling under hypoxia. In the present study, the osteoclasts formation induced by RAW264.7 was significantly promoted in conditioned media (CM) from osteocytic MLO-Y4 exposed to hypoxia in vitro. Therefore, hypoxic MLO-Y4 cells simulated by 100 μmol/L CoCl₂ or 2% O₂ stably expressed HIF-1α proteins and upregulated the expression of receptor activator of nuclear factor-κB ligand (RANKL) at both the messenger RNA (mRNA) and protein level. Furthermore, with the Knockdown of HIF-1α, the expression of RANKL mRNA and protein decreased after transient transfection. In addition, the phosphorylation of Janus kinase 2 (JAK2) and signal transducer and activator of transcription (STAT3) was also correlated with HIF-1α and RANKL levels under hypoxia. Then AG490, a JAK2 inhibitor, inhibited p-JAK2, p-STAT3 and RANKL expression. It was possible that AG490 disturbed the contact of HIF-1α and RANKL by JAK2/STAT3 pathway, influencing osteoclastogenesis. Our findings suggested that HIF-1α promoted the expression of RANKL by activating JAK2/STAT3 pathway in MLO-Y4 cells, and enhanced osteocyte-mediated osteoclastic differentiation in vitro.