摘要
目的探究mi R-200b对肠上皮紧密连接的影响及作用机制。方法利用阴性对照慢病毒及表达mi R-200b的慢病毒感染Caco-2细胞形成NC株和200b株,以10 ng/m L肿瘤坏死因子(TNF-α)处理建立体外肠上皮损伤模型,并分为NC组、NC+TNF-α组、200b组和200b+TNF-α组。测量4组细胞对肠上皮跨膜电阻抗(TEER)及对异硫氰酸荧光素-右旋糖酐(FITC-dextran)的通透性;Western blot检测肌球蛋白轻链激酶(MLCK)及磷酸化肌球蛋白轻链(P-MLC)表达。结果与NC组相比,NC+TNF-α组TEER值降低、对FITC-dextran通透量显著增高、MLCK和P-MLC表达上调,差异均有统计学意义(P?<0.05)。与NC+TNF-α组相比,200b+TNF-α组TEER显著升高、FITC-dextran通透量显著降低、MLCK和P-MLC表达显著下调,差异均有统计学意义(P?<0.05)。结论 mi R-200b可调控MLCK/P-MLC信号通路修复TNF-α导致的肠上皮紧密连接损伤。
Objective To explore the impact and mechanism of miR-200b on intestinal epithelial tight junction. Methods The negative-lentivirus and human-miR-200b-lentivirus were employed to infect the Caco-2 cell thus establishing two stable cell lines which were then stimulated by 10 ng/mL human tumor necrosis factor-a （TNF-a） to establish the model of the intestinal epithelial injury. Those Caco-2 cells were divided into NC, NC＋TNF-α, 200b, and 200b＋TNF-α groups.The tight junction permeability was detected by transepithelial electrical resistance （TEER） and Fluorescein isothioeyanate-labeled dextran （FITC- dextran）. The protein alterations myosin light chain kinase （MLCK）/phosphorylated myosin light chain （P-MLC） pathways were measured by Western blot analysis. Results Compared to NC group, NC＋TNF-α group had lower TEER, higher FITC-dextran, and up-regulated expressions of MLCK and P-MLC proteins （P〈0.05）. Compared to NC＋TNF-ct group, 200b＋TNF-α group had higher TEER, lower FITC-dextran and down-regulated expressions of MLCK and P-MLC proteins （P〈0.05）. Conclusion miR- 200b ameliorated TNF-α-induced intestinal epithelial tight junction disruption via regulation MLCK/P-MLC pathway.
出处
《临床儿科杂志》
CAS
CSCD
北大核心
2016年第7期538-541,共4页
Journal of Clinical Pediatrics
基金
国家自然科学基金项目(No.81370485)
