摘要
目的:研究暴发性肝衰竭小鼠大肠上皮细胞间紧密连接蛋白occludin表达的变化.方法:BALB/c小鼠150只随机分为生理盐水对照组(n=30)、内毒素(LPS)对照组(n=30)、D-氨基半乳糖(D-GalN)对照组(n=30)和暴发性肝衰竭(LPS+GalN)组(n=60).采用D-GalN和LPS联合ip制备暴发性肝衰竭小鼠动物模型.应用免疫组织化学技术、Westernblot及实时定量PCR检测暴发性肝衰竭进程中,小鼠大肠上皮细胞间紧密连接蛋白occludin的定位、表达及mRNA的变化.结果:occludin蛋白主要沿小鼠大肠黏膜上皮细胞膜的顶端呈线状分布,在暴发性肝衰竭组小鼠,6h时occludin的阳性染色开始减少,9h时更为明显.Westernblot结果与免疫组织化学结果相一致,6h时开始下降(0.48±0.07),9h达到最低值(0.36±0.05),与生理盐水对照组(0.71±0.09)相比差异显著(P<0.05).实时定量PCR结果显示,暴发性肝衰竭小鼠2h时occludinmRNA即开始下降(0.85±0.12),6h时达到最低值(0.72±0.04),与生理盐水对照组(1.00±0.05)相比有明显差异(P<0.05),9h时有所恢复(0.93±0.10).结论:在暴发性肝衰竭过程中,大肠上皮细胞间紧密连接蛋白occludin表达下降,其mRNA也呈下调趋势.
AIM： To observe the changes of intestinal epithelial tight junction protein occludin in mice with fulrninant hepatic failure （FHF）, and explore the mechanisms of spontaneous bacterial peritonitis （SBP）.
METHODS： A total of 150 male BALB/c mice were divided into group A, B, C and D. The mice in group D were intraperitoneally injected with lipopolysaccharide （LPS, 10 ~g/kg） and D-galactosamine （GAIN, 800 mg/kg） to induce the model of FHF, while the mice in group A, B and C were intraperitioncally injected normal saline, LPS, and GalN, respectively. At the end of the 2^rd, 6^th, and 9^th, the mice were killed for the collection of liver and intestinal specimens. Immunohistochemistry, Western blot and quantitative real time polymerase chain reaction （QRT-PCR） were used to detect the distribution and expression of intestinal epithelial tight junction protein occludin.
RESULTS： Tight junction protein occludin was localized along the apical region of the lateral plasma membrane representing the region of tight junctions in surface and crypt epithelial cells. In mucosal tissues from mice with FHF 6 and 9 h after injection, occludin-positive staining was gradually weakened. Western blot demonstrated consistent and significant reduction of occludin expression in mice with FHF at the 6th （0.48±0.07） and 9th h （0.36 ± 0.05） （P 〈 0.05）, which was significantly lower than that in group A （0.71±0.09） （P 〈 0.05）. Furthermore, the expression of occludin mRNA was decreased starting from the 2^nd h （0.85±0.12）, and reached the lowest level at 6^th h （0.72 ± 0.04）, significantly lower than that in group A. At the 9^th h, occludin mRNA expression almost restored （0.93 ± 0.10） to the normal level.
CONCLUSION： The expression of tight junction protein occludin is decreased at both protein and mRNA level in intestinal epithelial cells of mice with FHF.
出处
《世界华人消化杂志》
CAS
北大核心
2006年第31期3008-3012,共5页
World Chinese Journal of Digestology
基金
国家自然科学基金资助项目
No.30670947~~
