H型血管内皮细胞外泌体通过影响BMSCs成骨与成脂命运参与骨代谢调控

Senile osteoporosis and bone fracture compromise the quality of life and lead to early death. The primary cellular basis of senile osteoporosis is the age-dependent reduction in osteogenesis of bone marrow mesenchymal stem cells (BMSCs) accompanied with increasing adipogenesis of BMSCs. However, the mechanisms regulating osteogenesis and adipogenesis fate of BMSCs remain unclear. We previously found that H type endothelial cells-derived exosomes stimulated osteogenesis of BMSCs and inhibited its adipogenesis. In this study, at first, we will treat BMSCs and senile osteoporotic mice with exosomes from H-ECs cultured in media with various concentrations of oxygen. Then, we can get the best culture condition for accumulation of exosomes. Secondly, we will use Solexa high throughput sequencing and bioinformatics technology to screen out the key exosome miRNAs which stimulating osteogenesis and inhibiting adipogenesis of BMSCs, then to predict and validate the target genes and signaling pathways of the key miRNAs. At last, we will validate the effects of exosomes from H-ECs with overexpression of the key miRNAs on BMSCs and senile osteoporotic mice, as well as the mechanisms involved. In conclusion, this study aims to reveal the stimulatory actions on bone regeneration and inhibitory effects on bone marrow fat formation of exosomes from H-ECs, and the mechanisms involved, then to provide new ideas for prevention and treatment of senile osteoporosis.

老年人骨质疏松症（OP）和骨折导致了生存质量的下降和早逝。骨髓间充质干细胞（BMSCs）成骨减少与成脂增多是老年OP发病的主要细胞学基础；然而，BMSCs成骨与成脂命运调控的模式与机制仍不清楚。我们前期研究发现H型血管内皮细胞（H-ECs）通过分泌外泌体促进BMSCs成骨分化并抑制其成脂分化。我们拟进一步开展以下研究：①用不同氧浓度条件下培养的H-ECs的外泌体干预BMSCs与老年OP小鼠，筛选出富集H-ECs外泌体的最佳培养条件；②用Solexa高通量测序和生物信息学技术筛选出H-ECs外泌体中促成骨与抑成脂的关键miRNAs，并预测和验证其靶基因与靶向信号通路；③验证过表达关键miRNAs的H-ECs外泌体调控BMSCs成骨与成脂命运和改善老年OP的功效与机制。该项目旨在揭示H-ECs外泌体促进骨再生和抑制脂肪形成的功能及其分子机制，为临床老年OP防治研究提供新的思路。

老年人骨质疏松症（OP）和骨折导致了生存质量的下降和早逝。骨髓间充质干细胞（BMSCs）成骨减少与成脂增多是老年OP发病的主要细胞学基础；然而，BMSCs成骨与成脂命运调控的模式与机制仍不清楚。该项目原计划探明H型（CD31highEmcnhigh）血管内皮细胞（H-ECs）外泌体促进骨形成和抑制脂肪形成的功能及其分子机制，然而，我们按计划进行H-ECs分离培养过程中，发现原代H-ECs无法被稳定传代以获得大量可供分析的目的细胞，但CD31−Emcn+单阳性细胞可被稳定培养。进一步鉴定结果表明，此CD31−Emcn+细胞并非内皮细胞，而是一种间充质干细胞，并且其外泌体促进常规方法提取的CD31−Emcn− BMSCs成脂分化，抑制其成骨分化，提示CD31−Emcn+细胞是脂肪细胞的“龛（Niche）”。我们率先鉴定出骨髓脂肪细胞的龛细胞，为老年OP防治开辟了新思路与新方向。