复元醒脑汤对糖尿病脑梗死大鼠BMECs的影响及miRNA-320的调控机制研究

[目的]在细胞水平观察复元醒脑汤对糖尿病脑梗死大鼠缺血脑组织脑微血管内皮细胞(BMECs)的影响,从微小RNA-320(miRNA-320)调控途径探讨复元醒脑汤治疗糖尿病脑梗死的作用机制。[方法]进行复元醒脑汤大鼠灌胃,制备含药血清和对照血清。对糖尿病脑梗死大鼠的缺血脑组织进行BMECs的分离培养,分别加入对照血清、含药血清、转染miRNA-320 inhibitor、中药血清混合转染miRNA-320 inhibitor;实时荧光定量PCR(qPCR)法检测各组BMECs的miRNA-320、胰岛素样生长因子-1(IGF-1)和胰岛素样生长因子受体-1(IGF-1R)基因表达水平,通过甲臜化合物(MTS)检测复元醒脑汤对BMECs增殖能力的影响;通过Transwell实验观察复元醒脑汤对BMECs迁移能力的影响;观察复元醒脑汤对BMECs小管形成能力的影响。[结果]与对照血清组相比,含药血清组、miRNA-320 inhibitor组、含药血清混合miRNA-320 inhibitor组均能使BMECs的增殖、迁移以及小管形成能力显著提高(P0.01),同时显著降低了miRNA-320的表达水平,显著提高了IGF-1的表达水平。[结论]复元醒脑汤能够抑制糖尿病脑梗死大鼠缺血脑组织中miRNA-320表达,增加IGF-1的表达,进而促进BMECs的增殖、迁移及成管能力。

Mechanism research of regulation Fuyuan Xingnao Decoction to BMECs and expression microRNA-320 in diabetic cerebral infarction

[Objective] To explore the function of Fuyuan Xingnao Decoction on the BMECs of diabetic cerebral infarction and to explore specific mechanisms for the treatment of diabetic cerebral infarction from the microRNA-320 regulation pathways.[Methods] Rats were fed with Fuyuan Xingnao Decoction or PBS to prepare medicated serum and control serum. BMECs separated from diabetic rats'' cerebral infarction tissue. The control serum, medicated serum, microRNA inhibitors, medicated serum mixed microRNA inhibitors were added in to BMECs respectively. The gene expression of microRNA-320, IGF-1 and IGF-1R was observed with the method of qPCR. After the treatment of Fuyuan Xingnao Decoction, the MTS test was used to detect BMECs proliferation ability; Trans-well assay was used to observe BMECs migration ability; and tubule formation ability was also observed.[Results] Comparing with the control serum group, microRNA-320 inhibitor group, medicated serum group, medicated serum mixed with inhibitor control group all can improve the capacity of proliferation, migration and tubular formation in BMECs (P<0.01), decrease the expression of microRNA-320 and improve the expression of IGF-1.[Conclusions] Fuyuan Xingnao Decoction increased the protein secretion of IGF-1 by inhibiting the expression of microR-320, thus enhanced the capacity of proliferation, migration and tubular formation in BMECs.