Suppressing ERp57 diminishes osteoclast activity and ameliorates ovariectomy-induced bone loss via the intervention in calcium oscillation and the calmodulin/calcineurin/Nfatc1 pathway.

ABSTRACT

Background: Increased osteoclast activity constitutes the primary etiology of excessive bone erosion in postmenopausal osteoporosis. ERp57, otherwise referred to as protein disulfide isomerase A3 (PDIA3), plays a crucial role in the regulation of intracellular calcium signaling. This is documented to exert a profound impact on osteoclast differentiation and functionality. Methods: To ascertain the potential role of ERp57 in disease progression, prevention, and treatment, network pharmacology and bioinformatics analyses were conducted in relation to postmenopausal osteoporosis and ERp57 inhibitor (Loc14). Then, subsequent experimental verifications were employed in vitro on osteoclast and osteoblast, and in vivo on ovariectomy (OVX) mice models. Results: Multiple enrichment analyses suggested that the "calcium signaling pathway" may constitute a potential avenue for therapeutic intervention by Loc14 in the treatment of postmenopausal osteoporosis. In vitro experiments demonstrated inhibition of ERp57 could block osteoclast differentiation and function by interfering with the expression of osteoclast marker genes (Traf6, Nfatc1, and Ctsk). Further mechanisms studies based on calcium imaging, qPCR, and WB established that ERp57 inhibitor (Loc14) could obstruct calcium oscillation in osteoclast precursor cells (OPCs) by limiting the entry sources of cytosolic Ca2+ and interfering with calmodulin/calcineurin/Nfatc1 pathway. Evidence from Micro-CT scanning and double calcein labeling confirmed that the application of Loc14 in vivo could alleviate bone loss and partially reversed the osteogenic impairment caused by OVX in mice. Conclusions: Our findings proved the suppressive effects of Loc14 on osteoclastogenesis via attenuating calcium oscillation and associated singling pathways, providing ERp57 as a potential therapeutic target for postmenopausal osteoporosis.