Echinacoside targets HSC70 to inhibit osteoclastogenesis and ameliorate ovariectomy-induced osteoporosis.

Abstract

BACKGROUND: Echinacoside (ECH), a natural phenylethanoid glycoside isolated from Cistanche deserticola, exhibits promising therapeutic potential against osteoporosis (OP). Nevertheless, its specific molecular targets and the mechanisms involved are not yet fully understood. PURPOSE: This study integrates small-molecule affinity chromatography with both RANKL-induced osteoclastogenesis and ovariectomy (OVX)-induced rat models of OP to systematically investigate the molecular targets and mechanisms responsible for the anti-OP effects of ECH. METHODS: Small-molecule affinity chromatography and molecular dynamics simulations were employed to identify the direct molecular targets of ECH and their potential binding sites. Subsequently, combined with molecular biology techniques, the RANKL-induced osteoclastogenesis model was utilized to further elucidate the molecular mechanisms by which ECH regulates bone homeostasis. Finally, the anti-OP effect of ECH was confirmed in vivo using an OVX-induced rat models of OP. RESULTS: This study reveals that heat shock cognate 71 kDa protein (HSC70) serves as a key cellular target of ECH in modulating osteoclastogenesis. The amino acids ARG36 and ARG272 of HSC70 are confirmed as the primary binding sites for ECH. Notably, HSC70 is markedly upregulated in both OP models and clinical patient samples. Mechanistically, ECH suppresses osteoclastogenesis by promoting ubiquitination-mediated degradation of IKKβ. Furthermore, knockdown of Hsc70 demonstrated a similar biological effect as ECH treatment. In vivo studies validate that ECH, via targeting HSC70, markedly ameliorates bone loss and metabolic dysregulation in the OVX-induced rat OP model. Specifically, Hsc70 overexpression induces bone loss in normal rats. Conversely, Hsc70 knockdown alleviates bone loss in OVX rats. Moreover, both overexpression and knockdown of Hsc70 abolish the therapeutic effect of ECH in OVX rats. CONCLUSION: This work reveals HSC70 as a novel regulator of bone remodeling and proposes ECH, a new HSC70 inhibitor, as a promising therapeutic agent. This study provides important insights into the pathogenesis of OP and proposes an innovative targeting strategy for metabolic bone disorders.