双歧杆菌对肠上皮细胞生长和白介素-8分泌功能的影响

Effect of Bifidobacterium on cell growth and IL-8 secretion in IECs

目的探讨双歧杆菌对人肠上皮细胞株HT29生长及其IL-8分泌水平的影响。方法HT29细胞在96孔板上生长24 h后分为正常细胞对照组、高剂量双歧杆菌共培养组(细菌终浓度为1×1010CFU/ml)、低剂量双歧杆菌共培养组(细菌终浓度为1×106CFU/ml)、轮状病毒感染对照组,分别加入不同剂量双歧杆菌和感染轮状病毒共培养,继续培养24 h,光镜下观察细胞生长状态,MTT比色法检测细胞活性情况,ELISA检测细胞培养上清中IL-8表达水平。结果光镜下观察到双歧杆菌与HT29细胞共培养后细胞形态无明显改变,共培养24 h后MTT检测双歧杆菌对HT29细胞增殖和调亡无明显影响,但轮状病毒感染对照组细胞病变脱落,活细胞数量明显减少。共培养6 h,其余3组细胞培养上清中IL-8分泌较正常细胞对照组增加(P<0.05),高剂量双歧杆菌组增加较低剂量双歧杆菌组差异有显著性(P<0.05),但两个剂量组均明显低于轮状病毒感染阳性对照组的IL-8分泌增加水平(P<0.05);感染后24 h,细胞培养上清中IL-8分泌水平高于正常细胞对照组(P<0.05),但高、低剂量双歧杆菌组之间差异无显著性(P>0.05),两个剂量组IL-8分泌增加水平均明显低于轮状病毒感染阳性对照组(P<0.01)。结论两歧双歧杆菌共培养不影响HT29细胞的生长,双歧杆菌能够促进HT29细胞分泌细胞因子IL-8,但明显低于致病微生物刺激引起的细胞因子分泌水平改变,这种促进作用无时间—剂量依赖关系,提示双歧杆菌与肠道内致病微生物对肠道免疫功能的影响不同,双歧杆菌促进肠上皮细胞分泌IL-8可能与其参与的肠道黏膜免疫系统发育成熟相关。

Objective To study the effect of Bifidobacterium bifidum（B, bifidum） on cell growth and IL-8 secretion in intestinal epithelial cell line HT29. Methods B. bifidums were anaerobically cultivated. HT29 ceils were divided into four groups：normal control group （ NC group） ,low dose B. b/fidum co-cultivated group （ LB group） and high dose B. bifidum co-cultivated group （ HB group） and RV infected control group （PdC group）. Inactivated B. b/fidum were respectively added into LB group and HB group with final concentration of 1 ×10^6 CFU/ml and 1 ×10^9 CFU/ml and HT29 cells were infected with RV in PdC group. The state of cell was observed and the number of living cell were evaluated by MTT after 24 h;Cell supernatants were collected 6 h,24 h post cocultivation, and examined for secretion level of IL-8 by ELISA. Resuits There were no change in LB group and HB group 24 h post cocultivation, but the RIC group showed CPE at 24 h and the number of living cell decreased significantly compared with NC group, LB group and HB group. The other three groups had a higher IL-8 level （ P 〈 0.05 ） than NC group 6 and 24 h post coeultivation （ P 〉 0.05 ）, in which, HB group increased significantly compared with LB group. But markedly lower level of both two than RIC group （P 〈 0.05 ）. Conclusions B. bifidum could induce the secretion of cytokines IL-8 in HT29 cells with no damage of cell, but the change significantly little than which the pathogenic microorganism induced. It displays the different effect of B. bifidum and pathogenic microorganism on IECs, the promotion of IL-8 secretion may involve in the development and maturation of mucosal immunity.