活血解毒通络法对急性脑皮质下白质梗死ephrin-A5信号通路的神经重塑作用机制

In recent years, neurologists notice that subcortical white matter stroke due to a very serious clinical consequences. The clinical pathology exists in the subcortical white matter and brainstem, thalamus and basal ganglia lacunar infarction. It is a sign of worsening disease progression that appearing loss of blood stasis toxin syndrome after acute subcortical white matter stroke 3-5 days.This subject intends to prove ephrin-A5 signaling pathway was activated afteracute cerebral ischemia, the axon and glial cell were damaged too; activating blood，removing toxic substance dredging method can adjust the signal path to repair the blood brain barrier and reshape damaged white matter. Under the guidance of Chinese Medicine theory, we may establish rat model of loss of blood stasis toxin syndrome of subcortical white matter, observe changes of ephrin-A5 expression at different timepoint, clearly know the changes rule of ephrin-A5 in acute subcortical white matter stroke, reveal the dynamic changes of ephrin-A5 signaling pathway. We study on the influence activating blood，removing toxic substance dredging method of ephrin-A5 and its signaling pathway downstream molecules. We research the relevant between loss of blood stasis toxin syndrome in.acute stroke and ephrin-A5 signaling pathways on protein molecular level, and promote the remodeling mechanism of the activating blood ，removing toxic substance dredging method to ephrin-A5 signaling pathway.

近年来，脑皮质下白质梗死因有非常严重的临床后果而引起人们的注意。其临床病理为存在于皮质下白质和脑干，丘脑和基底节腔隙性梗死。急性脑缺血后3-5天时出现的血瘀毒损脑络证是病情进展恶化的标志。本课题拟证实急性脑缺血后启动和活化ephrin-A5信号通路，轴突和胶质细胞受损；活血解毒通络法可以通过调节该信号通路，从而修复血脑屏障，重塑受损的脑白质神经细胞。本课题在中医病证结合的理论指导下，建立急性脑皮质下白质梗死血瘀毒损证大鼠模型，观察急性脑白质梗死后不同时点ephrin-A5表达的变化，明确大鼠急性白质梗死后ephrin-A5的动态变化，揭示ephrin-A5信号通路的动态变化规律。研究活血解毒通络法对ephrin-A5及其信号通路下游分子激活情况的影响。从蛋白分子层面研究急性脑梗死血瘀毒损证与ephrin-A5信号通路相关性，及活血解毒通络法对ephrin-A信号通路的神经重塑作用机制。

ephrin-A5信号通路是急性脑缺血后血脑屏障破坏的重要环节之一。急性缺血性卒中后启动和活化ephrin-A5信号通路，抑制ephrin-A5信号通路可以显著改善脑缺血的预后。研究表明活血解毒通络法可以显著改善急性脑缺血后血脑屏障，重塑受损的神经细胞，但机制不清楚。本研究采用急性脑缺血血瘀毒损证大鼠模型，观察大鼠急性脑缺血后ephrin-A5信号通路的动态变化，证实活血解毒法对急性脑缺血后神经细胞和血脑屏障的作用，从蛋白分子层面研究急性脑梗死血瘀毒损证与ephrin-A5信号通路相关性，及活血解毒通络法中药古碟子对急性脑皮质下白质梗死ephrin-A5信号通路的神经重塑作用，进一步阐述急性白质梗死“毒损脑络”病因病机的分子生物学基础。研究结论1.急性脑皮质下白质梗死后，缺血海马、皮质Ephrin-A5及EphB2表达变化，Ephrin-A5信号通路的激活是血脑屏障破坏的主要原因之一。2．急性脑缺血后1h海马区Ephrin-A5及EphB2过度表达，6h皮质区Ephrin-A5及EphB2过度表达，表明海马的缺血敏感性可能早于皮质。3.本研究中基因的表达水平与细胞中对应蛋白质的表达没有完全一致，可能与以下原因有关：a.真核基因表达的转录和翻译发生的时间和位点存在时空间隔;b.mRNA和蛋白质半衰期不同 c.mRNA转录后可以调控不同的蛋白质 d.基因表达的部位与时间与相应蛋白质的活化时间及执行功能的部位有所不同。4.苦碟子通过调节急性脑缺血后Ephrin-A5信号通路起到神经重塑作用。苦碟子干预后，Ephrin-A5及EphB2过度表达明显好转，推测可能是通过干预ephrin-A5信号转导系统，抑制星形细胞反应，阻碍胶质瘢痕形成，促进轴突萌发及神经功能恢复，进而起到神经重塑作用。5.苦碟子对ephrin-A5信号转导通路的调节作用可以为活血解毒法中药治疗急性脑皮质下梗死的作用靶点。6.Meta分析显示：苦碟子注射液联合常规治疗急性脑梗死在改善总有效率、神经功能缺损程度、Barthel指数、血清炎性因子等指标上皆明显优于常规治疗，证明苦碟子注射液对发病72小时内超早期或早期急性脑梗死患者具有良好的治疗效果，显示了临床应用苦碟子注射液的有效性；且不良反应发生率低。