过表达Fibulin-5对大鼠脑缺血再灌注损伤后血脑屏障的保护作用

Fibulin-5 over-expression protects blood brain barrier in rats after cerebral ischemia/reperfusion injury

目的探讨过表达Fibulin-5对大鼠脑缺血再灌注损伤后血脑屏障的保护作用及其可能的机制。方法将112只成年雄性SD大鼠分成4组:假手术组(S)、脑缺血组(I/R)、脑缺血加过表达Fibulin-5腺病毒载体干预组(I/R+AdFBLN)以及脑缺血加空腺病毒载体干预组(I/R+Ad-HK)。大鼠在大脑中动脉阻塞前7 d接受腺病毒注射,再灌注后24 h,采用Western blot、RT-PCR和免疫荧光检测各组大鼠脑组织Fibulin-5和紧密连接蛋白occludin的表达;伊文思蓝漏出评估血脑屏障的通透性;TTC染色检测脑梗死体积并进行神经功能缺损评分。结果脑缺血再灌注24 h,与S组比较,I/R组大鼠伊文思蓝明显漏出(P<0.01),伴有紧密连接蛋白occludin表达降低(P<0.01)。I/R+Ad-FBLN组Fibulin-5的mRNA水平和蛋白水平明显高于其他3组(P<0.01),伊文思蓝漏出明显减少(P<0.01),occludin的表达明显增高(P<0.01),TTC梗死灶明显缩小(P<0.01),神经功能缺损明显减轻(P<0.01)。而I/R+Ad-HK组与I/R组之间差异均无统计学意义(P>0.05)。结论过表达Fibulin-5减轻大鼠脑缺血再灌注损伤后血脑屏障的损害,Fibulin-5促进紧密连接蛋白occludin的表达可能是其保护作用的机制之一。

Objective To determine the effect of adenovirus mediated over-expression of Fibulin-5 on blood brain barrier leakage in rats after cerebral ischemia/reperfusion （I/R） injury and investigate the possible underlying potential mechanism. Methods A total of 112 adult male Sprague-Dawley rats were randomly divided into sham operation group, cerebral I/R group, I/R plus AD-Fibulin-5 group, and I/R plus AD-HK group. Adenovirus carrying fibulin-5 gene （AD-Fibulin-5） or blank adenovirus （AD-HK） were injected into the right cortex and hippocampus of the rats from the latter 2 groups. In 7 d after gene transfer, the rats, except for those of the sham operation group, underwent 90 min of ischemia and then were reperfused for 24 h. The expression of fibulin-5 and occluding in the brain tissue were detected by Western blotting, RT-PCR and immunohistochemical assay. Evan-blue extravasation was used to measure the permeability of blood brain barrier. TTC staining was employed to observe the cerebral infarct volume. Neurological function was also evaluated. Results Intervention of AD-Fibulin-5 resulted in significantly increased expression of fibulin-5 at protein and mRNA levels （P〈0.01）, smaller amount of Even-blue leakage （P〈0.01）, enhanced expression of occludin （P〈0.01）, obviously decreased infarct volumes （P〈0.01）, obviously milder neurological impairment （P〈0.01） when compared to the other groups. Conclusion Over-expression of Fibulin-5 attenuates blood brain barrier leakage in rats after cerebral I/R injury, which might be due to its enhancing the expression of occludin.