通过RT-PCR检测小鼠胃黏膜感染海尔曼螺杆菌后细胞因子的表达

RT-PCR detection of cytokine expression in gastric mucosae of mice with Helicobacter heilmannii infection

目的通过比较海尔曼螺杆菌(Helicobacter heilmannii,H.heilmannii)感染BALB/c小鼠、C57BL/6小鼠、裸鼠前后以及不同时期胃黏膜IFN-γ、IL-4、IL-10及IL-1β的mRNA表达水平的变化,确定海尔曼螺杆菌感染小鼠胃黏膜诱发的免疫应答方式。方法采用小鼠体内传代保存的海尔曼螺杆菌株,分别灌胃3种小鼠,通过胃内接种,使海尔曼螺杆菌定植于小鼠胃内,建立长期稳定的小鼠感染模型。于第4、8、12、24、36周分别处死小鼠,取胃黏膜组织作半定量RT-PCR检测IFN-γ、IL-4、IL-10及IL-1β的mRNA水平。结果感染初期(4~12周),BALB/c、C57BL/6小鼠胃黏膜的IFN-γ、IL-4、IL-10、IL-1β的mRNA表达量较对照组均明显升高(P<0.05),其中IL-4、IL-10、IFN-γ的mRNA水平在第8~12周分别达到最大峰值,而在第4周IL-1β的mRNA水平已达高峰。12周后,BALB/c小鼠的IFN-γ表达明显降低,C57BL/6小鼠的变化不明显;感染后期(36周)的C57BL/6小鼠,IL-4的表达在感染组低于对照组(P<0.05)。免疫缺陷小鼠中IFN-γ、IL-4、IL-10、IL-1β的mRNA的表达量均较对照组增高,组间相比IL-1β、IL-4的mRNA表达差异有统计学意义(P<0.05)。结论细胞因子的表达随海尔曼螺杆菌感染时间的延长而增加,IFN-γ介导的Th1型细胞免疫在海尔曼螺杆菌感染免疫应答中占重要地位。不同品种小鼠对海尔曼螺杆菌感染的免疫应答方式有差别,C57BL/6小鼠对海尔曼螺杆菌感染主要呈Th1型细胞免疫应答,BALB/c小鼠为Th1/Th2混合免疫应答。

Objective To compare the changes in IFN-γ,IL-4,IL-10 and IL-1βexpression at mRNA level in gastric mucosae of BALB/c,C57BL/6 and nude mice at different stages of Helicobacter heilmannii(H.heilmannii)infection,and to investigate the types of induced immune responses.Methods Each kind of mice was randomly divided into two groups:infection(n=30)and control(n=6)groups.Those in the infection groups were intragastrically inoculated with H.heilmannii strains to establish long-term stable mouse infection models.Gastric mucosa tissues were collected at weeks 4,8,12,24 and 36 and analyzed by semi-quantitative RT-PCT to detect the expression of IFN-γ,IL-4,IL-10 and IL-1βat mRNA level.Results In the early stage of infection(weeks 4-12),INF-γ,IL-4,IL-10,and IL-1βexpression at mRNA level in the gastric mucosae of BALB/c and C57BL/6 mice were significantly increased compared with those of the control groups(P<0.05).IL-4,IL-10 and IFN-γexpression peaked at weeks 8-12,while IL-1βexpression reached the peak at week 4.After 12 weeks,IFN-γexpression at mRNA level in BALB/c mice was significantly decreased,but showed no significant change in C57BL/6 mice.IL-4 expression at mRNA level in C57BL/6 mice at the late stage of infection(week 36)was lower than that in the corresponding control group(P<0.05).Expression of IFN-γ,IL-4,IL-10 and IL-1βat mRNA level in nude mice were all higher than those in the control group,and there were significant differences in IL-1βand IL-4 expression between groups(P<0.05).Conclusions Expression of cytokines in H.heilmannii-infected mice increased over time.IFN-γ-mediated Th1 immune responses were the predominant immunity induced by H.heilmannii infection.Immune responses to H.heilmannii infection varied with the kinds of mice.C57BL/6 mice showed mainly Th1 cell immune responses,while Th1/Th2 mixed immune responses were induced in BALB/c mice.