CAJ | 학술논문

目的观察高压氧(HBO)对缺血再灌注(I/R)人类肠黏膜上皮Caco-2细胞间紧密连接(TJ)的影响.方法培养单层人类大肠黏膜上皮细胞Caco-2,应用0.5 mmol/L丁基过氧化氢(t-buooh)诱导离体肠黏膜上皮细胞I/R损伤.按数字表法随机分为5组(每组8个培养孔):对照组(N组)、I/R组、HBO预处理(HBO-P)组、缺血期HBO干预(HBO-I)组、再灌注期HBO干预(HBO-R)组.采用跨膜电阻抗检测法(TEER)检测肠道黏膜上皮细胞间紧密连接的开放程度并评价其通透性.用Rt-PCR检测细胞间紧密连接蛋白ZO-1转录水平.结果 TEER阻值N组最高,其次为HBO-P组,再次为HBO-R组,最后为HBO-I组.N组与其他各组差异均有统计学意义(P＜0.01);HBO-P组阻值与I/R组、HBO-I组及HBO-R组的阻值比较,差异均有统计学意义(P＜0.05);HBO-R组与I/R组、HBO-I组阻值比较差异有统计学意义(P＜0.05);L/R组高于HBO-I组,且差异有统计学意义(P＜0.05).ZO-I mRNA的Rt-PCR结果显示:与N组相比,其他各组ZO-1 mRNA含量均明显下降,且差异有统计学意义(P＜0.01);HBO-P组ZO-1 mRNA含量均高于L/R组、HBO-R组及HBO-I组(P＜0.01);HBO-R组ZO-1 mRNA含量高于I/R组和HBO-I组(P＜0.05);HBO-I组与I/R组ZO-1 mRNA含量接近,差异无统计学意义(P＞0.05).结论 HBO干预具有保护单层Caco-2细胞TJ功能作用,抑制TJ开放,机制可能与维持TJ的骨架蛋白ZO-1有关.缺血期前和再灌注期给予HBO干预治疗显示有益结果,HBO预处理疗效最佳,缺血期给予HBO无效.
목적 관찰고압양(HBO)대결혈재관주(I/R)인류장점막상피Caco-2세포간긴밀련접(TJ)적영향.방법 배양단층인류대장점막상피세포Caco-2,응용0.5 mmol/L정기과양화경(t-buooh)유도리체장점막상피세포I/R손상.안수자표법수궤분위5조(매조8개배양공):대조조(N조)、I/R조、HBO예처리(HBO-P)조、결혈기HBO간예(HBO-I)조、재관주기HBO간예(HBO-R)조.채용과막전조항검측법(TEER)검측장도점막상피세포간긴밀련접적개방정도병평개기통투성.용Rt-PCR검측세포간긴밀련접단백ZO-1전록수평.결과 TEER조치N조최고,기차위HBO-P조,재차위HBO-R조,최후위HBO-I조.N조여기타각조차이균유통계학의의(P＜0.01);HBO-P조조치여I/R조、HBO-I조급HBO-R조적조치비교,차이균유통계학의의(P＜0.05);HBO-R조여I/R조、HBO-I조조치비교차이유통계학의의(P＜0.05);L/R조고우HBO-I조,차차이유통계학의의(P＜0.05).ZO-I mRNA적Rt-PCR결과현시:여N조상비,기타각조ZO-1 mRNA함량균명현하강,차차이유통계학의의(P＜0.01);HBO-P조ZO-1 mRNA함량균고우L/R조、HBO-R조급HBO-I조(P＜0.01);HBO-R조ZO-1 mRNA함량고우I/R조화HBO-I조(P＜0.05);HBO-I조여I/R조ZO-1 mRNA함량접근,차이무통계학의의(P＞0.05).결론 HBO간예구유보호단층Caco-2세포TJ공능작용,억제TJ개방,궤제가능여유지TJ적골가단백ZO-1유관.결혈기전화재관주기급여HBO간예치료현시유익결과,HBO예처리료효최가,결혈기급여HBO무효.
Objective To investigate the effects of hyperbaric oxygen (HBO) on the tight junction (T J) of monolayer human intestinal epithelial cells Caco-2 in ischemia reperfusion (I/R) injury in vitro.Methods In the culture of human intestinal monolayer epithelial cells Caco-2,0.5 mmol/L t-buooh was used to induce I/R injury of intestinal epithelial cells in vitro.The monolayer Caco-2 cells were randomly divided into 5 groups (n =8 cases):the control group,the ischemia/reperfusion group (I/R),the hyperbaric oxygen preconditioning group (HBO-P),the HBO intervention during ischemia group (HBO-I) and the HBO intervention during reperfusion group (HBO-R).Transmembrane electrical impedance detection (TEER) was used to detect the opening of the tight junction between epithelial cells of the intestinal mucosa and evaluate its permeability.The transcription of tight ligandin ZO-1 between cells was detected by RT-PCR.Results Results indicated that the impedance as shown by TEER was in the following order:the impedance value of the control group was the highest,then came the HBO-P group,further down was the HBO-R group,and at the end was the HBO-I group.Statistical significance could be noted when comparisons were made between the HBO-Pgroup and all the other groups(P ＜ 0.01).There was also statistical significance,when the impedance value of the HBO-P group was compared with those of the I/R,HBO-I and HBO-R groups (P ＜ 0.05).Statistical significance could also be noted,when the impedance value of the HBO-R group was compared with those of the I/R and HBO-I groups(P ＜0.05).The impedance value of the I/R group was higher than that of the HBO-I group,with statistical significance(P ＜0.05).RT-PCR results of ZO-1 indicated that the ZO-1 mRNA levels of the I/R,the HBO-P,the HBO-R and the HBO-I groups were decreased significantly,as compared with that of the control group,with statistical significance (P ＜ 0.01).The ZO-1 mRNA level of the HBO-P group was higher than those of the I/R,the HBO-R and the HBO-I groups,with statistical significance (P ＜0.01).The ZO-1 mRNA level of the HBO-R group was higher than those of the I/R and the HBO-I groups,also with statistical significance (P ＜ 0.05).The ZO-1 mRNA level of the HBO-I group was close to that of the I/R group,but without statistical significance(P ＞ 0.05).Conclusions HBO intervention seemed to have a certain protective effect on the tight junction of monolayer Caco-2 cells,and inhibit the decrease of tight ligandin ZO-1,the mechanism of which might be associated with the maintenance of the level of ZO-1.HBO intervention before ischemia and during reperfusion seemed to have beneficial effects,and HBO preconditioning could produce better results.Nevertheless,HBO intervention during ischemia seemed to be ineffective.