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目的了解脂多糖(LPS)对大鼠脑微血管内皮细胞(BMECs)通透性的影响及其可能的机制。方法分离和培养原代大鼠BMECs,随机分为对照组和LPS干预组。利用跨内皮细胞电阻抗检测BMECs屏障功能,Pull-down法测定RhoA活性,Western blot检测p115RhoGEF和紧密连接蛋白ZO-1、occludin、claudin-5的蛋白表达量。结果与对照组(159.0±8.6Ω.cm2)比较,LPS作用3 h后,大鼠BMECs跨内皮细胞电阻抗值明显下降(108.3±4.2Ω.cm2),作用12 h后达最低(85.4±2.5Ω.cm2)。LPS作用5 min后RhoA明显活化,1 h后出现p115RhoGEF蛋白表达上调,3 h后紧密连接蛋白ZO-1、occludin和claudin-5的表达水平均不同程度下降,与对照组比较差异有统计学意义(P<0.05)。结论 LPS诱导大鼠BMECs中p115RhoGEF/RhoA信号通路活化,导致紧密连接蛋白表达水平降低,引起BMECs通透性增加。[中国当代儿科杂志,2011,13(11):908-911]
Objective To investigate the effect of lipopolysaccharide (LPS) on the permeability of rat brain microvascular endothelial cells (BMECs) and its possible mechanism. Methods Primary rat BMECs were isolated and cultured and randomly divided into control group and LPS intervention group. The barrier function of BMECs was detected by the trans-endothelial cell impedance assay. The RhoA activity was measured by Pull-down assay. The protein expression of p115RhoGEF, tight junction proteins ZO-1, occludin and claudin-5 were detected by Western blot. Results Compared with the control group (159.0 ± 8.6Ω.cm2), after 3 h of LPS treatment, the trans-endothelial cell resistance of BMECs decreased significantly (108.3 ± 4.2Ω.cm2) and reached the lowest level at 12 h (85.4 ± 2.5 Ω.cm2). The expression of p115RhoGEF protein was up-regulated after LPS treatment for 5 min, and the expression of tight junction protein ZO-1, occludin and claudin-5 decreased after 3 h, with significant difference compared with the control group (P <0.05). Conclusion LPS can induce activation of p115RhoGEF / RhoA signaling pathway in rat BMECs, resulting in decreased expression of tight junction protein and increased permeability of BMECs. [Chinese Journal of Contemporary Pediatrics, 2011,13 (11): 908-911]