基于p38 MAPK/Drosha信号通路探讨脑出血患者继发性脑损伤及神经细胞凋亡的机制

Investigation to the Mechanism of Secondary Brain Injury and Nerve Cell Apoptosis in Patients with Intracerebral Hemorrhage Based on p38 MARK/Drosha Signaling Pathway

目的探讨p38丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)/Drosha信号通路对脑出血(intracerebral hemorage,ICH)继发性脑损伤(secondary brain injury,SBI)及神经细胞凋亡的影响。方法通过胶原酶注射建立的ICH小鼠模型。同时,将经氧合血红蛋白(oxyhemoglobin,OxyHb)处理的SH-SY5Y细胞用作体外ICH模型。小干扰RNA(siRNA)用于抑制ICH小鼠和SH-SY5Y细胞中p38表达。分别通过苏木精-伊红(hematoxylin-eosin,HE)染色法和Nissl染色观察血肿形成及Nissl小体形成情况。Western blot分析p38 MAPK/Drosha信号通路表达情况。流式细胞术评估细胞凋亡情况。结果siRNA介导的p38敲除有效缓解了ICH小鼠血肿形成(P<0.05),减少了海马组织CA1和皮质区域的Nissl小体数量(P<0.05),增加了脑干重/湿重比值(P<0.05),并降低了神经行为评分(P<0.05)。此外,p38的敲除显著增加了ICH小鼠和OxyHb处理的SH-SY5Y细胞中Drosha蛋白水平(P<0.05),显著降低了OxyHb诱导的神经元凋亡率(P<0.01)。结论p38上调通过抑制miRNA生物合成中的关键酶Drosha的表达介导ICH后SBI发展。

Objective To investigate the effect of p38 mitogen-activated protein kinase(MAPK)/Drosha signaling pathway on the secondary brain injury(SBI)after intracerebral hemorage(ICH)and nerve cell apoptosis.Methods The ICH mouse model was established by collagenase injection.Meanwhile,SH-SY5 Y cell treated with oxyhemoglobin(OxyHb)were used as the in vitro ICH model.Short interfering RNA(siRNA)were used to inhibit ICH mice and p38 expression in SH-SY5 Y cells.Hematoma formation and Nissl body formation were observed by hematoxylin-eosin(HE)staining and Nissl staining respectively.The expression of p38 MAPK/Drosha signaling pathway were analysed by Western blot.Cell apoptosis was evaluated by flow cytometry.Results siRNA mediated p38 knockout effectively alleviated the formation of hematoma in ICH mice(P<0.05),which reduced the number of Nissl corpuscles in CA1 and cortex of hippocampus(P<0.05),increased the ratio of brain stem weight/wet weight(P<0.05)and decreased the neurobehavioral score(P<0.05).In addition,the knockout of p38 significantly increased the Drosha protein level in ICH mice and SH-SY5 Y cells treated with OxyHb(P<0.05),which significantly reduced the apoptosis rate induced by OxyHb(P<0.01).Conclusion Up-regulation of p38 may contribute to SBI mediating ICH by suppressing the expression of Drosha,a key enzyme,in miRNA biogenesis.