MicroRNA-195 参与切应力对血管内皮细胞炎症调控作用的研究

Role of miR-195 in regulation of vascular endothelial cell inflammation by shear stress

摘要:

目的 研究microRNA-195（miR-195）是否参与切应力对血管内皮细胞（ECs）炎症的调控作 用。方法 ①验证生理切应力调节ECs miRNAs 的表达：15 dyne/cm2 的生理切应力处理人脐静脉内皮细胞 （HUVEC），24 h 后利用实时荧光定量聚合酶链反应（qRT-PCR）检测内皮细胞相关miRNAs（miR-27b、 miR-143/145 及miR-195 等）的表达变化；② miR-195 与内皮细胞炎症反应的关系：体外转染miR-195 mimics 及inhibitors 至HUVEC，通过Western blot、免疫荧光检测ECs 中炎症相关因子表达及miR-195 对内 皮细胞的炎症控制水平。结果 15 dyne/cm2 的生理切应力处理HUVEC，24 h 后miR-27b、miR-143/145 及 miR-195 等表达上调；过表达miR-195 对ECs 内皮型一氧化氮合酶（eNOS）的表达没有影响，但抑制了 p65 的磷酸化（P-p65）表达；采用miRNA inhibitors 抑制内源性的miR-195 后，ECs eNOS 的表达下调，抑 制了κB 抑制蛋白（IκBα）的表达水平。免疫荧光结果显示，在ECs 中过表达miR-195 抑制了NF-κB 的核转运，而抑制miR-195 可能促进了NF-κB 的核转运，促进炎症。结论 生理切应力诱导miR-195 表 达上调，miR-195 可能参与切应力对ECs 炎症的调控作用。

Abstract:

Objective To investigate whether miR-195 is involved in the regulation of vascular endothelial cell (EC) inflammation by shear stress. Methods To verify the regulation of EC miRNA expression by physiological shear stress, 15 dyne/cm2 of physiological shear stress were used to process human umbilical vein endothelial cells (HUVECs), after 24 hours, the expressions of miRNAs (miR-27b, miR-143/145, miR-195, etc.) in endothelial cells were detected by qRT-PCR. To study the relationship between miR-195 and endothelial cell inflammatory response, miR-195 mimics and inhibitors were transfected into HUVECs in vitro, the expressions of inflammatory factors in ECs and the effect of miR-195 in controlling endothelial cell inflammation were detected using Western blot and immunofluorescence technique. Results After processing HUVECs with 15 dyne/cm2 of physiological shear stress for 24 hours, the expressions of miR-27b, miR-143/145 and miR-195 were significantly up-regulated (P < 0.05). The over-expression of miR-195 did not affect the expression of eNOS, but significantly inhibited the expression of P-p65 (P < 0.05); while inhibition of endogenous miR-195 by miRNA inhibitors significantly reduced the expression of EC eNOS (P < 0.05) and inhibited the expression of IKBa (P < 0.05). Immunofluorescence results showed that over-expression of miR-195 in ECs inhibited nuclear translocation of NF-κB, while inhibition of miR-195 may promote the nuclear translocation of NF-κB and promote inflammation. Conclusions The expression of miR-195 is significantly up-regulated by physiological shear stress. miR-195 may be involved in the regulation of EC inflammation by shear stress.