Abstract: Background: Exosomes derived from bone marrow stem cells (BMSC-Exos) are considered as candidates for osteoporosis (OP) therapy. Estrogen is critical in the maintenance of bone homeostasis. However, the role of estrogen and/or its receptor in BMSC-Exos treatment of OP, as well as its methods of regulation during this process remain unclear.Methods:BMSCs were cultured and characterized. Ultracentrifugation was performed to collect BMSC-Exos. Transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and western blotting were used to identify BMSC-Exos. We examined the effects of BMSC-Exos on the proliferation, osteogenic differentiation, mineralization, and cell cycle distribution of MG-63 cells. The protein expression of estrogen receptor α (ERα) and the phosphorylation of extracellular signal-regulated kinase (ERK) were investigated through western blotting. We determined the effects of BMSC-Exos on the prevention of bone loss in female rats. The female SD rats were divided into three groups: the sham group, ovariectomized (OVX) group, and the OVX + BMSC-Exos group. Bilateral ovariectomy was performed in the OVX and OVX + BMSC-Exos groups, while a similar volume of adipose tissue around the ovary was removed in the sham group. The rats in OVX+BMSC-Exos group were given BMSC-Exos after 2 weeks of surgery. Micro-CT scanning and histological staining were used to evaluate the in vivo effects of BMSC-Exos.Results:BMSC-Exos significantly upregulated the proliferation, ALP activity, and the ARS staining in MG-63 cells. The results of cell cycle distribution demonstrated that BMSC-Exos significantly increased the proportion of cells in the G2+S phase and decreased the proportion of cells in the G1 phase. Moreover, PD98059, an inhibitor of ERK, downregulated the expression of ERα, which was promoted by administration of BMSC-Exos. Micro-CT scan showed that in the OVX+BMSC-Exos group, BMSC-Exos significantly promoted ERα expression, with ameliorated bone mineral density (BMD), bone volume/tissue volume fraction (BV/TV), trabecular number (Tb. N), and trabecular separation (Tb. Sp). Additionally, the microstructure of the trabecular bone was preserved in the OVX + BMSC-Exos group compared to that in the OVX group.Conclusion:BMSC-Exos showed an anti-osteoporotic role in OVX rats both in vitro and in vivo, which may involve the ERα/ERK signaling pathway.