lncRNA-<i>H19</i>/<i>IGF1R</i>调控高血压脑出血后脑微血管内皮细胞自噬的机制研究

刘文萍; 刘丽丹; 彭颜晖

doi:10.16190/j.cnki.45-1211/r.2023.09.006

lncRNA-H19/IGF1R调控高血压脑出血后脑微血管内皮细胞自噬的机制研究

Mechanism of lncRNA-H19/IGF1R regulating autophagy of brain microvascular endothelial cells after hypertensive intracerebral hemorrhage

摘要

摘要: 目的：分析lncRNA-H19（H19）在高血压脑出血（HICH）患者体内的表达特征，并探讨H19 影响人脑微血管内皮细胞（HBMECs）自噬与氧糖剥夺（OGD）损伤的机制。方法：收集健康者、高血压患者和HICH患者血液样本，采用lncRNA array芯片检测lncRNA表达谱，RT-qPCR检测H19表达水平并分析其与脑出血量及NIHSS评分的关系；预测H19的靶蛋白，RT-qPCR检测其mRNA水平并分析其与H19的相关关系；采用OGD处理敲降或过表达H19及其互作蛋白的HBMECs，而后采用免疫荧光检测细胞自噬情况，western blotting检测自噬相关蛋白（LC3-Ⅰ和LC3-Ⅱ）以及缺氧诱导因子-1α（HIF-1α）和血管内皮生长因子（VEGF）。结果：H19 在高血压患者（t=0.021，P< 0.05）和HICH 患者（t=0.018，P< 0.05）中显著高于健康者；并随HICH分级升高而增加，与脑出血量及NIHSS评分呈正相关关系（R²=0.087，R²=0.113，P< 0.05）；随OGD处理时间增加，HBMECs中H19 水平增加（t=0.023，P< 0.05）；IGF1R mRNA 在HICH 患者体内低于健康者（t=0.015，P< 0.05），且随HICH 加剧而降低（t=0.043，P< 0.05）；HICH 患者体内H19 与IGF1R 呈负相关关系（R²=0.187，P< 0.05）；OGD 处理下调HBMECs 中IGF1R 蛋白，敲降H19可使OGD-HBMECs中IGF1R增加；在OGD-HBMECs中LC3自噬体、LC3-Ⅱ/LC3-Ⅰ、HIF-1α和VEGF均因敲降H19或过表达IGF1R而减少，但同时过表达IGF1R及H19后则均增加。结论：H19与HICH病情相关，下调H19可增加IGF1R，进而缓解OGD诱导的HBMECs自噬和损伤。

Abstract: Objective:To analyze the expression characteristics of lncRNA-H19(H19)in patients with hypertensive intracerebral hemorrhage (HICH) and explore the mechanism of H19 affecting autophagy and oxygen and glucose deprivation (OGD) injury of human brain microvascular endothelial cells (HBMECs).Methods:Blood samples of healthy people, hypertension patients and HICH patients were collected.The lncRNA array chip was used to detect the expression profile of lncRNA, and real-time fluorescence quantitative polymerase chain reaction(RT-qPCR)was used to detect the expression level of H19.The relationship between H19 expression and the amount of cerebral hemorrhage and NIHSS score was analyzed.The target protein of H19 was predicted, its mRNA expression level was detected by RT-qPCR, and its correlation with H19 was analyzed.HBMECs with knockdown or overexpression of H19 and its interacting proteins were treated with OGD.Autophagy was detected by immunofluorescence.Autophagy related proteins(LC3-Ⅰand LC3-Ⅱ), hypoxia inducible factor-1α(HIF-1α)and vascular endothelial growth factor(VEGF)were detected by western blotting.Results:The level of H19 in patients with HICH was significantly higher than that in healthy people (t=0.021, P< 0.05) and patients with hypertension(t=0.018, P< 0.05).The level of H19 increased with the increase of HICH grade, and was positively correlated with the amount of cerebral hemorrhage and NIHSS score (R²=0.087, R²=0.113, P< 0.05).The level of H19 in HBMECs increased with the increase of OGD treatment time(t=0.023, P< 0.05).IGF1R mRNA level in patients with HICH was lower than that in healthy controls people (t=0.015, P< 0.05), and decreased with the aggravation of HICH (t=0.043, P< 0.05).There was a negative correlation between H19 and IGF1R in patients with HICH (R²=0.187, P< 0.05).OGD treatment decreased the level of IGF1R protein in HBMECs, but knockdown of H19 increased IGF1R protein in OGD-HBMECs.In OGD-HBMECs, LC3 autophage, LC3-Ⅱ/LC3-Ⅰ, HIF-1α and VEGF decreased by knockdown of H19 or overexpression of IGF1R, but increased by overexpression of both IGF1R and H19 at the same time.Conclusion:H19 is related to HICH.Down regulation of H19 can increase IGF1R, and then futher alleviate OGD induced autophagy and injury of HBMECs.

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