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目的检测大鼠大脑中动脉栓塞(MCAO)后,瞬时受体电位(TRP)V4在梗死灶周围的表达变化,研究TRPV4在脑缺血损伤中的作用及机制。方法在大鼠MCAO模型中,免疫印迹法观察脑缺血后不同时间点梗死灶周围脑组织TRPV4表达的变化;在体外培养星形胶质细胞的氧糖剥夺(OGD)模型中,通过MTT实验和检测乳酸脱氢酶释放量,研究不同浓度TRPV4抑制剂钌红(RR)对OGD后细胞存活的影响;检测星形胶质细胞肿瘤坏死因子-α(TNF-α)和白介素(IL)-1βmRNA含量,研究不同浓度RR对OGD后细胞释放促炎因子的影响。结果 (1)大鼠MCAO后,梗死灶周围TRPV4表达迅速上调,于1 d迅速达到高峰并持续到3 d左右(P<0.01),之后开始下降,7 d时仍高于正常水平(P<0.05);(2)较小浓度RR(5μM)能增加OGD后星形胶质细胞的存活(P<0.05);(3)不同浓度的RR(5、10、20μmol/L)均能降低OGD后星形胶质细胞TNF-α和IL-1βmRNA水平(P<0.01)。结论 MCAO后大鼠脑组织TRPV4显著上调。在体外培养的星形胶质细胞中,TRPV4抑制剂能减轻OGD损伤,该作用可能通过抑制细胞因子而实现。
Objective To detect the expression of transient receptor potential (TRP) V4 around infarct in rats after middle cerebral artery occlusion (MCAO), and to study the role and mechanism of TRPV4 in cerebral ischemia injury. Methods The changes of TRPV4 expression in brain tissue of infarcted area at different time points after cerebral ischemia were observed by Western blotting in MCAO rat model. In OGD model of astrocyte cultured in vitro, MTT assay And the release of lactate dehydrogenase (LDH) were measured to investigate the effect of ruthenium red (TRPV4) inhibitor on the cell survival after OGD. The levels of tumor necrosis factor-α (TNF-α) and interleukin- 1βmRNA, to study the effect of different concentrations of RR on pro-inflammatory cytokines released from OGD cells. Results (1) After MCAO, the expression of TRPV4 in the infarct area rapidly increased and peaked at 1 d and continued until about 3 d (P <0.01), and then began to decline. At 7 d, TRPV4 was still higher than the normal level (P < 0.05). (2) RR (5μM) increased the survival of astrocytes after OGD (P <0.05). (3) Different concentrations of RR (5, 10 and 20μmol / L) Post-astrocyte TNF-α and IL-1β mRNA levels (P <0.01). Conclusion The expression of TRPV4 in rat brain after MCAO was significantly up-regulated. In cultured in vitro astrocytes, TRPV4 inhibitors can reduce OGD damage, which may be achieved by inhibiting cytokines.