淫羊藿苷经miR-23b干预激素性骨微血管内皮细胞损伤的机制研究

Disordered circulation in bone is the key etiological factor for glucocorticoids-induced osteonecrosis of femoral head. Injury of bone microvascular endothelial cells hereby is the critical factor. miRNAs were found to play important roles in regulating vascular endothelial cells’ function. The applicant has studied the influence of glucocorticoids on bone microvascular endothelial cells, especially via transcriptome of miRNAs. The previous study was supported by the national natural science fund 81273972 and found that miR-23b in bone microvascular endothelial cells was significantly influenced by glucocorticoids. After information analysis with multiple databases including miRWalk, miR-23b was found to have significant effect on many intracellular signal transduction factors, such as Sprouty2, Sema6A, Sema6D, Apaf-1, TNFAIP1, TNFAIP2, TNFAIP3. These factors comprehensively affected bone microvascular endothelial cells and osteocyts, involving several signal transduction pathways such as apoptosis pathway and angiogenesis. As for the pathogenesis of glucocorticoids-induced damage to endothelial cells, miR-23b is supposed to play an important role in BMECs by certain signal pathways. In this study, we plan to qualify the effect of miR-23b on BMECs injury induced by glucocorticosteroids, along with its downstream coordinates. Therefore we would clarify the mechanism beneath the regulatory axis, i.e. miRNAs-target genes-differentially expressed proteins. Moreover, we will qualify the effect of icariin upon the expression levels of miR-23b and its downstream coordinates. Finally, we will define the regulation pathway of miR-23b and reveal the mechanism how icariin protect the bone microvascular endothelial cells from injury induced by glucocorticoids.

血供障碍是激素性股骨头坏死的病理基础，骨微血管内皮细胞(BMECs)损伤是关键环节，机制尚不明。我们前期研究发现，激素明显引起BMECs miR-23b下调，而有“补肾壮骨”功效的淫羊藿苷对激素性BMECs损伤有保护作用，且上调miR-23b表达。经靶点预测，miR-23b调控多种细胞因子,涉及细胞凋亡和血管生成等信号通路。淫羊藿苷是否经miR-23b调控上述通路起保护作用？值得深入研究。本项目拟用激素性BMECs损伤模型及兔激素性股骨头坏死模型，通过miR-23b高表达/敲减系统研究miR-23b对激素性BMECs损伤信号通路的调控作用,并通过萤光素酶报告基因系统进一步验证其分子靶点，揭示miR-23b调控激素性BMECs损伤的分子机制。通过淫羊藿苷干预BMECs损伤中miR-23b及其下游分子的研究，揭示淫羊藿苷保护激素性BMECs损伤的机制及靶点，诠释中医“肾主骨”理论的科学内涵。

骨微血管内皮细胞(BMECs)损伤是激素性股骨头坏死的密切相关，其机制尚不明。前期研究发现，激素明显引起BMECs miR-23b下调，而淫羊藿苷对激素性BMECs损伤有保护作用，且上调miR-23b表达。淫羊藿苷是否经miR-23b调控上述通路起保护作用值得深入研究。我们进行了BMECs全转录组测序分析，体外验证了ICA通过miRNA-222调控BMECs的成血管分化作用机制研究，连接介导和调控Y蛋白（JMY）在糖皮质激素（GC）诱导内皮细胞凋亡及迁移抑制中的作用机制，研究了miR-23b对激素性股骨头坏死发生和血管生成的影响，及对BMECs损伤的作用及相关信号通路。构建miR-23b调控的分子靶点的启动子重组质粒，通过萤光素酶报告基因系统，验证miR-23b对上述靶点的调控作用。我们初步筛选了具有表达差异的miRNA，lncRNA circRNA，明确了ICA通过miRNA-222调控BMECs的成血管分化作用机制，以及JMY在HUVEC经GC诱导损伤的发病机理，揭示了miR-23b在激素性BMECs损伤中的可能信号作用通路为VEGF-AKT通路，分析其对BMECs凋亡及血管生成的影响，通过研究淫羊藿苷干预激素性股骨头坏死模型中miR-23b及其下游分子的变化，揭示淫羊藿苷对激素性BMECs损伤起保护作用的分子机制及作用靶点。