CAJ | 학술논문

目的：观察沉默信息调节因子1（SIRT1）对肠缺氧上皮Caco-2细胞屏障功能的影响并探讨其分子机制。方法对Caco-2细胞分别进行1％O2浓度缺氧6 h（缺氧组）和常规处理（常规组）及加浓度为40μmol/L Resveratrol（SIRT1激动剂）预处理6 h后再缺氧6 h（观察组），分别检测3组肠上皮细胞跨上皮电阻（TER），PCR及Western blot分别检测3组SIRT1及紧密连接蛋白ZO-1、Occludin和Claudin-1的 mRNA与蛋白表达。结果缺氧组SIRT1 mRNA及其蛋白的相对表达量均明显低于常规组（分别为0．40±0．02比0．70±0．07，P＝0．001；0．37±0．03比0．76±0．03，P＝0．001），也明显低于观察组（0．50±0．02，P＝0．026；0．54±0．02，P＝0．011）。缺氧组3种紧密连接蛋白mRNA表达均低于常规组（P＜0．05），也低于观察组（P＜0．05），观察组的ZO-1及Claudin-1表达与常规组比较，差异无统计学意义（P＞0．05）。3种紧密连接蛋白的表达水平常规组最高，观察组次之，缺氧组最低，两两比较，差异均具有统计学意义（均P＜0．05）。常规组、缺氧组和观察组的TER分别为（142±7） Ohm/cm2、（94±3） Ohm/cm2和（119±7） Ohm/cm2；两两比较，差异均具有统计学意义（均P＜0．05）。结论缺氧条件下SIRT1水平降低，而SIRT1的激活可通过增加肠道紧密连接蛋白ZO-1、Occludin和Claudin-1 mRNA与蛋白的表达，从而减轻缺氧对肠上皮屏障功能的损伤。
목적：관찰침묵신식조절인자1（SIRT1）대장결양상피Caco-2세포병장공능적영향병탐토기분자궤제。방법대Caco-2세포분별진행1％O2농도결양6 h（결양조）화상규처리（상규조）급가농도위40μmol/L Resveratrol（SIRT1격동제）예처리6 h후재결양6 h（관찰조），분별검측3조장상피세포과상피전조（TER），PCR급Western blot분별검측3조SIRT1급긴밀련접단백ZO-1、Occludin화Claudin-1적 mRNA여단백표체。결과결양조SIRT1 mRNA급기단백적상대표체량균명현저우상규조（분별위0．40±0．02비0．70±0．07，P＝0．001；0．37±0．03비0．76±0．03，P＝0．001），야명현저우관찰조（0．50±0．02，P＝0．026；0．54±0．02，P＝0．011）。결양조3충긴밀련접단백mRNA표체균저우상규조（P＜0．05），야저우관찰조（P＜0．05），관찰조적ZO-1급Claudin-1표체여상규조비교，차이무통계학의의（P＞0．05）。3충긴밀련접단백적표체수평상규조최고，관찰조차지，결양조최저，량량비교，차이균구유통계학의의（균P＜0．05）。상규조、결양조화관찰조적TER분별위（142±7） Ohm/cm2、（94±3） Ohm/cm2화（119±7） Ohm/cm2；량량비교，차이균구유통계학의의（균P＜0．05）。결론결양조건하SIRT1수평강저，이SIRT1적격활가통과증가장도긴밀련접단백ZO-1、Occludin화Claudin-1 mRNA여단백적표체，종이감경결양대장상피병장공능적손상。
Objective To observe the effect of SIRT1 on intestinal barrier function of epithelial Caco-2 cells under hypoxia and investigate its mechanism. Methods Caco-2 cells were randomly divided into three groups: normoxia group (Nx), hypoxia group (Hx,1%O2 for 6 h) and hypoxia plus 40 μmol/L Resveratrol (agonist of SIRT1) group (Hx+Res). Transepithelial electrical resistance (TER) was determined. mRNA and protein expressions of SIRT1 and tight junctions (ZO-1, Occludin, Claudin-1) were examined by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting. Results Both mRNA and protein expressions of SIRT1 were significantly reduced in Hx group as compared with Nx group (0.40±0.02 vs. 0.70±0.07, P=0.001; 0.37±0.03 vs. 0.76±0.03, P=0.001). The mRNA and protein expressions of SIRT1 were significantly increased in Hx+Res group as compared with Hx group(0.50±0.02 vs. 0.40±0.02, P=0.026; 0.54±0.02 vs. 0.37±0.03, P=0.011). The expression levels of ZO-1 , Occludin and Claudin-1 in Hx group were lower than those in Nx group (P<0.05), however, pretreatment with Resveratrol could attenuate the decreased expression of above 3 molecules under hypoxia (P<0.05). TERs of Nx group, Hx group and Hx+Res group were (142±7) Ohm/cm2, (94±3) Ohm/cm2 and (119±7) Ohm/cm2 respectively. Compare with the Nx group, the TER of Hx group was significantly decreased (P<0.05). TER of Hx+Res group was significantly increased compare with Hx group, but it was still significantly lower than that in Nx group (P<0.05). Conclusions Expression of SIRT1 is significantly reduced under hypoxia. Activation of SIRT1 can maintain the epithelial barrier function through regulating the expression of tight juncions under hypoxia.