ESPE Abstracts

Introduction: Glucocorticoid-induced osteoporosis is the most frequent cause of secondary osteoporosis. Glucocorticoids are known to diminish β-catenin activity via Wnt pathway signaling, leading to osteoporosis and osteonecrosis. Bone homeostasis depends on the regulation of β-catenin in osteoblasts. Disruption of bone homeostasis, oxidative stress and mitochondrial dysfunction are major causes of glucocorticoid-induced osteoporosis; therefore, alleviation of excess oxidative stress in osteoblasts is a potential therapeutic strategy for osteoporosis. Exosomes derived from bone marrow mesenchymal stem cell, as novel cell-free therapeutics, can modulate various biological processes, such as immunomodulation, reduce oxidative damage, and promote tissue repair as well as regeneration. The aim of this study is to evaluate the biological effects of bone marrow stem cell derived exosomes in an in vitro model of glucocorticoid induced osteoporosis model by using an experimental setting that resembles the in vivo conditions.

Methods: Primary osteoblasts and osteocyte cell line were used to reproduce an in vitro model of glucocorticoid induced osteoporosis by adding dexamethasone (1μM) for 24h. Cells were then treated with bone marrow derived exosomes for 24h and quantitative Polymerase Chain Reaction (qPCR) and western blot were performed to study whether exosomes activated the Wnt/β-catenin pathway. Dexamethasone challenge reduced bone formation in primary osteoblasts and bone mineralization in osteocytes; moreover, Wnt/β-catenin pathway was reduced following incubation with dexamethasone in both osteoblasts and osteocytes. Cell viability and proliferation were assessed using MTS. Osteogenic potential was evaluated through Western blotting, real-time PCR, alkaline phosphatase activity assay, and alizarin red staining. Differential expression of miRNAs was determined by high-throughput sequencing.

Results: In this study, we found that exosomes derived from bone marrow had a positive impact on the proliferation, differentiation, alkaline phosphatase activity, and the Alizarin red S staining in osteoblast and osteocyte cells. The results of cell cycle distribution demonstrated that exosomes increased the proportion of cells in the G2+S phase and decreased the proportion of cells in the G1 phase. Functionally, exosomes could promote the viability of osteoblasts, and significantly increase the expression level of annexin V. In addition, exosomes remarkably downregulated apoptosis-related genes and decreased apoptosis in osteoblasts. There was significant downregulation of IL-1β and TNF-α in osteoblasts, and there was upregulation of IL-1β and downregulation of TNF-α in osteoclasts. Importantly, we demonstrated that exosomes mimics could reverse dexamethasone-induced osteoporosis in vitro.

Conclusions: Exosomes can promote osteoblast proliferation by inhibiting cell apoptosis, and therefore may have a potential preclinical therapeutic effect on glucocorticoid-induced osteoporosis.