黄体酮促进脑出血后轴突再生及其相关机制

Progesterone promotes axonal regeneration after cerebral hemorrhage and related mechanisms in rats

目的观察黄体酮对脑出血(intracerebral hemorrhage, ICH)后大脑基底节区神经元轴突再生的影响,探讨其具体的机制。方法将SD大鼠分为脑出血组、黄体酮小剂量组(8 mg/kg)、黄体酮大剂量组(16 mg/kg),假手术组为对照组。观察各组术后1、3、7、14 d神经功能缺损、脑组织含水量、病理变化及轴突生长相关蛋白表达变化,并分析术后3 d各组脑血肿体积、氧化应激相关指标。联用大剂量黄体酮和PI3K/Akt通路抑制剂LY294002作为抑制剂组,检测术后3 d各组轴突相关蛋白及通路蛋白p-Akt、RhoA表达量的变化,并采用双标荧光对GAP43、p-Akt/RhoA进行定位。结果脑出血组大鼠在各时间点的神经功能障碍和脑水肿程度最重,脑血肿体积最大,氧化应激产物水平增高最明显,黄体酮干预后在各时间点损伤情况缓解,大剂量组比小剂量组的效果更明显,差异具有统计学意义(P<0.05);Western blot检测结果显示,黄体酮大剂量组比黄体酮小剂量组和脑出血组GAP43、NF200表达增加,MAG、Nogo-A表达降低,组间差异具有统计学意义(P<0.05);与黄体酮大、小剂量组比较,抑制剂组GAP43、NF200及p-Akt表达下调,MAG、Nogo-A、RhoA表达量增高,差异均有统计学意义(P<0.05);双标荧光结果显示,与脑出血组比较,黄体酮大剂量组神经元细胞上p-Akt表达上升,RhoA表达降低,差异具有统计学意义(P<0.05)。结论黄体酮可改善脑出血后神经功能障碍,促进损伤轴突的再生修复,该机制可能与激活PI3K/Akt通路从而抑制RhoA表达有关。

Objective To determine the effect of progesterone on axon regeneration of cerebral ganglia neurons after intracerebral hemorrhage(ICH),and explore the underlying mechanism.Methods SD rats were randomly divided into ICH group,low-(8 mg/kg)and high-dose progesterone(16 mg/kg)groups,and sham operation group.Combined treatment of high-dose progesterone and PI3 K/Akt pathway inhibitor LY294002 was given to the rats from the inhibitor group.The neurological deficits,brain water content and pathological changes and expression of axon growth-related proteins were observed at 1,3,7 and 14 d after operation.And the cerebral hematoma volume and oxidative stress related indexes were analyzed at 3 d after operation.The expression levels of axonal-associated proteins and pathway proteins p-Akt and RhoA were detected at 3 d after operation,and the locations of GAP43 and p-Akt/RhoA were identified with immunofluorescence staining.Results The ICH group had severest neurological dysfunction and cerebral edema at each time point,and largest volume of cerebral hematoma and highest levels of oxidative stress products.After progesterone intervention,the damage was relieved at each time point,with the efficacy of high-dose more obvious than low-dose(P<0.05).Western blot results showed that the high-dose progesterone intervention increased the expression levels of GAP43 and NF200 and decreased those of MAG and Nogo-A when compared to the low-dose progesterone and ICH groups(P<0.05).Western blot analysis indicated that the expression of GAP43,NF200 and p-Akt was down-regulated while that of MAG,Nogo-A and RhoA was increased in the inhibitor group compared with the high-and low-dose progesterone groups(P<0.05).The results of double-labeled fluorescence staining showed that the expression of p-Akt was increased and that expression of RhoA was decreased in neuronal cells of the high-dose progesterone group compared with ICH group(P<0.05).Conclusion Progesterone improves neurological dysfunction after cerebral hemorrhage and promotes regeneration and repair of damaged axons,which may be related to activation of PI3 K/Akt pathway and inhibition of RhoA expression.