高压氧联合凋亡信号调节激酶抑制剂NQDI-1对大鼠脑缺血半暗带区神经细胞及血脑屏障通透性的保护作用

Co-treatment effects of HBO and inhibitor NQDI-1 of ASK-1 on nerve cells and blood-brain barrier permeability in ischemia penumbra of cerebral ischemia reperfusion injured rats

目的通过高压氧(hyperbaric oxygen,HBO)联合凋亡信号调节激酶(apoptosis signal-regulating kinase-1,ASK-1)抑制剂NQDI-1治疗脑缺血再灌注(ischemia reperfusion,IR)损伤大鼠,研究HBO治疗与大脑皮层缺血半暗带(ischemia penumbra,IP)区域神经细胞凋亡、ASK-1信号通路之间的关系,探索大鼠脑IR损伤后HBO与NQDI-1联合治疗对IP区域的作用。方法体质量260300 g的雄性大鼠,建立大脑中动脉损伤模型模拟脑IR。随机分为六组:正常对照组(NC组)、假手术组(Sham组)、IR组、IR损伤后HBO治疗7 d组(IR-HBO组)、IR损伤后NQDI-1治疗组(IR-NQDI-1组)、IR损伤后HBO联合NQDI-1治疗组(IR-HBONQDI-1组),每组6只。应用伊文思蓝染色、苏木精-伊红(HE)染色、免疫荧光染色等方法测定IR损伤及HBO联合NQDI-1治疗7 d前后IP区域血脑屏障(BBB)通透性及ASK-1、硫氧还蛋白-1(thioredoxin-1,TRX-1)、促分裂原活化蛋白(mitogen-activated protein,MMP)-9等的表达量。结果与假手术组比较,IR组HE染色显示神经细胞凋亡显著,伊文思蓝染色面积显著增大,ASK-1表达量显著增高(P<0.05),TRX-1表达量显著降低(P<0.05),MMP-9表达量显著增高(P<0.05)。在HBO联合NQDI-1治疗后,与IR组比较,HE染色显示凋亡程度显著降低,伊文思蓝染色面积显著减小,ASK-1表达量显著降低(P<0.05),TRX-1表达量显著增高(P<0.05),MMP-9表达量显著降低(P<0.05)。结论脑IR损伤能够造成大鼠BBB通透性降低;ASK-1信号通路可能参与IR造成的神经细胞凋亡;HBO联合抑制剂NQDI-1能够进一步恢复BBB通透性,显著降低凋亡相关细胞因子的表达,改善神经细胞微环境,抑制神经细胞凋亡的发生。

Objective To study the relationship of hyperbaric oxygen( HBO) treatment with neuronal apoptosis and apoptosis signal-regulating kinase-1( ASK-1) signal pathway in ischemia penumbra( IP) region of cerebral ischemia reperfusion( IR) injured rats treated with HBO and inhibitorNQDI-1 of ASK-1 in order to explore the effect of HBO and NQDI-1 co-treatment on IP region of brain IR injured rats. Methods The models of middle cerebral artery injured rats were established in male rats weighing 260-300 g to simulate brain IR injury and were divided into control group( NC group),Sham group,IR group,HBO treatment for 7 days after IR( IR-HBO group),NQDI-1 treatment after IR( IR-NQDI-1 group),HBO + NQDI-1 treatment after IR( IR-HBO-NQDI-1 group),six in each group. Evans blue staining,HE staining and immunofluorescence staining were used to determine the permeability of blood-brain barrier( BBB) and the expression of ASK-1,thioredoxin-1( TRX-1) and mitogen-activated protein-9( MMP-9) in IP area of IR injured rats and HBO +NQDI-1 co-treatment rats. Results HE staining showed that neuronal apoptosis was significant in IR group rats,Evans blue staining area was significantly larger in IR group rats than that in Sham group,the expression of ASK-1 was significantly increased and TRX-1 was significantly decreased,and the expression of MMP-9 was significantly increased in the rats of IR group( P< 0. 05). After HBO +NQDI-1 co-treatment for IR rats,HE staining showed that the degree of apoptosis was significantly lower in IR-HBO-NQDI-1 group than that in IR group,Evans blue staining area was significantly smaller in IR-HBO-NQDI-1 group than that in IR group,the expression of ASK-1 was significantly decreased,the expression of TRX-1 was significantly increased,and the expression of MMP-9 was significantly decreased in IR-HBO-NQDI-1 group( P< 0. 05). Conclusion In the rats,IR damage can decrease the permeability of BBB. ASK-1 signal pathway may be involved in neuronal apoptosis caused by IR injury in IP region. Treatment of HBO combined with inhibitor NQDI-1 can further restore the permeability of BBB,and significantly reduce the expression of apoptosis-related cytokines,then improve nerve cell microenvironment and inhibit the occurrence of nerve cell apoptosis in the rats.