Cervus elaphus sibiricus (deer antler) extract alleviates osteoporosis via dual modulation of osteoblast and osteoclast activity in ovariectomy-induced mice on network pharmacology.

Cervus elaphus sibiricus (deer antler; Cervi Parvum Cornu; Nokyong) refers to the unossified antlers of young male deer and has been traditionally used in East Asian medicine to tonify the kidney and strengthen bone, supporting musculoskeletal health (osteoporosis, age-related frailty). Despite its long-standing use, the pharmacological mechanisms underlying its anti-osteoporotic effects remain largely unelucidated. This study aimed to evaluate the anti-osteoporotic effects of a dual-extraction deer antler extract (PKDE) and to elucidate its mechanism of action through an integrative approach combining network pharmacology with in vivo and in vitro models. Network pharmacology (KEGG, GO, PPI) was used to identify bone-related targets, and active compounds were verified by HPLC-MS. An ovariectomy (OVX)-induced osteoporosis mouse model was used to evaluate the in vivo efficacy of PKDE (48, 96, and 144 mg/kg, p.o., 4 weeks). Bone mineral density (BMD), bone mineral content (BMC), collagen deposition, and adipocyte size were assessed. Serum levels of TRACP-5b, CTX, and osteocalcin were measured. Osteogenic and osteoclastic gene expression in tibial tissue was analyzed by RT-PCR. In vitro, the effects of PKDE on osteoclastogenesis and osteoblast differentiation were investigated using RAW264.7 and SaOS-2 cells, respectively. PKDE increased BMD and BMC, restored collagen, and reduced marrow adiposity. Serum levels of TRACP-5b and CTX decreased, whereas osteocalcin increased. Gene expression analysis revealed downregulation of osteoclast markers (RANKL/OPG ratio) and upregulation of osteoblast markers (Col1a1, Bmp2, Spp1) in tibial tissue. In vitro, PKDE suppressed osteoclastogenesis in RANKL-induced RAW264.7 cells, as evidenced by a reduction in TRAP-positive multinucleated cells and downregulation of osteoclast-related genes such as Nfatc1 and Ctsk. In contrast, PKDE promoted osteoblast differentiation in AA/β-GP-induced SaOS-2 cells, as demonstrated by enhanced mineralization via Alizarin Red S staining and upregulated expression of osteogenic markers including COL1A1, BMP2, Runx2, SPP1, and IBSP. No hepatotoxicity or nephrotoxicity was observed. PKDE exerts dual anti-osteoporotic effects by inhibiting osteoclastogenesis and promoting osteoblast differentiation via modulation of bone metabolism pathways, supporting the traditional use of PKDE and highlighting its potential as a natural-origin therapeutic for bone health.