乳酸杆菌对牦牛大肠黏膜屏障的保护作用及Bax和Bcl-2蛋白表达的影响

Protective effects of Lactobacillus on the colorectal mucosal barrier of yak and its impact on Bax and Bcl-2 protein expression

为了探讨乳酸杆菌对犊牦牛大肠埃希氏菌(E.coli)所致大肠黏膜屏障的病理损伤,以及对凋亡关键蛋白Bax、Bcl-2的影响,试验将24头健康犊牦牛随机分为空白对照组、模型组、低LAB组、高LAB组,空白对照组牦牛每日每头灌服生理盐水10 mL,持续10 d;模型组牦牛前5 d灌服生理盐水,第6天灌服生理盐水配制的E.coli O78,菌落总量为5×10^(8)cfu,第7~10天每天灌服生理盐水;低LAB组牦牛第1~6天与模型组一致,第7~10天每天灌服生理盐水配制的乳酸杆菌10 mL,菌落总量为1×10^(7)cfu;高LAB组牦牛第1~6天与模型组一致,第7~10天每天灌服生理盐水配制的乳酸杆菌10 mL,菌落总量为1×10^(11)cfu。第11天宰杀牦牛,采集各组牦牛盲肠、结肠和直肠肠段组织,采用H.E.染色法分别观察犊牦牛盲肠、结肠病理变化,通过Western-blot检测牦牛盲肠、结肠和直肠组织中凋亡关键蛋白Bax、Bcl-2的相对表达量。结果表明:与空白对照组相比,模型组盲肠、结肠固有层有大量炎症细胞浸润,肠上皮细胞出现变性、坏死及脱落现象;高LAB组的肠组织结构完整,肠绒毛及肠腺较发达;与空白对照组相比,模型组Bax蛋白在盲肠、结肠和直肠中的相对表达量显著增加(P<0.05),Bcl-2蛋白的相对表达量显著降低(P<0.05);经乳酸杆菌干预治疗后,Bax蛋白相对表达量随着乳酸杆菌总量的增加呈依赖性下降且差异显著(P<0.05),Bcl-2蛋白相对表达量随着乳酸杆菌菌落总量的增加呈依赖性增加且差异显著(P<0.05);高LAB组Bax和Bcl-2蛋白相对表达量与空白对照组相比差异不显著(P>0.05)。说明菌落总量为1×10^(11)cfu的乳酸杆菌对E.coli O78所致的腹泻犊牦牛起到了保护作用。

In order to investigate the effect of Lactobacillus on the pathological damage to the colorectal mucosal barrier in calf yaks caused by Escherichia coli(E.coli),and its influence on the key apoptosis protein Bax and Bal-2,in the experiment,twenty-four healthy calves were randomly divided into blank control group,model group,low LAB group and high LAB group.Calves in the blank control group received 10 mL of saline daily for 10 days.Yaks in the model group received saline for the first 5 d;E.coli 078 prepared in saline totaling 5×10^(8) cfu on day 6,and saline daily on days 7 to 10.Yaks in the low LAB group were consistent with the model group from day 1 to 6,and were received 10 mL of Lactobacillus prepared in saline daily from day 7 to 10,totaling 1×10^(7) cfu.Yaks in the high LAB group were consistent with the model group from day I to 6,and received 10 mL of Lactobacillus prepared in saline every day on day 7 to 10,with a total amount of 1×10^(11)cfu.Yaks were slaughtered on the 1lth day;tissues of cecum,colon and rectum were collected from each group of yaks;pathological changes of cecum and colon of yaks were observed by H.E.staining method.The relative expression of apoptotic proteins Bax and Bcl-2 in the cecum,colon and rectum of calf yaks was detected by Western-blot.The results showed that compared with the blank control group,the model group had a large number of inflammatory cells infiltration in the cecum and colon lamina propria,and the intestinal epithelial cells appeared to be degenerated,necrotic and detached.The intestinal tissue structure of the high LAB group was intact,and the intestinal villi and intestinal glands were more developed.Compared with the blank control group,the relative expression of Bax protein in the cecum,colon and rectum of the model group was significantly increased(P<0.05),and the relative expression of Bcl-2 protein was significantly decreased(P<0.05).After treatment with Lactobacillus intervention,the relative expression of Bax protein decreased dependently and significantly with the increase of Lactobacillus dose(P<0.05),and the relative expression of Bcl-2 protein increased dependently and significantly with the increase of Lactobacillus dose(P<0.05).The relative expression of Bax and Bcl-2 proteins in the high LAB group was not significantly different from that of the blank control group(P>0.05).The results indicated that the Lactobacllus with 1×10^(11) cfu had a protective effect on the diarrhea of calves caused by E.coli 078.