巨噬细胞感染鼠伤寒沙门菌后NO表达及与胞内菌增殖的关系

目的探讨鼠巨噬细胞RAW264.7在感染鼠伤寒沙门菌野生株（ATCC 10248）后细菌在细胞内的增殖过程。方法分别采用鼠伤寒沙门菌活菌和灭活菌（细菌：细胞比例为20∶1）感染鼠巨噬细胞,于感染1、4、8、12、24 h时采用实时定量RT-PCR检测iNOS mRNA变化;采用免疫荧光染色法计数感染细胞内细菌数量;采用0.1%Triton X-100 PBS裂解活菌,计数感染细胞活菌数量。结果活的鼠伤寒沙门菌感染后,在前12 h细菌细胞内持续增殖;活菌和灭活菌感染均能引起感染的巨噬细胞iNOS mRNA表达增加（P均〈0.05）,尤以灭活菌感染者为著;细菌感染后细胞内NO生成增加（P均〈0.05）。结论鼠伤寒沙门菌感染能有效促进鼠巨噬细胞iNOS表达,细胞内活的鼠伤寒沙门菌可抑制细胞iNOS表达及NO生成。

Relationship between NO expression and Salmonella typhimurium proliferation in infected macrophages

Objective To explore the proliferation process of Salmonella typhimurium wild strain（ATCC 10248）in infected macrophages（RAW264.7）.Methods RAW264.7 macrophages were infected at a multiplicity of infection of 20 with living and inactivated Salmonella typhimurium for 1 h.Realtime RT-PCR was used to detect iNOS expression inside the host cell 1 h,4 h,8 h,12 h or 24 h after infection,respectively;immunofluorescence stain was employed to count the bacterial numbers in infected RAW264.7 macrophages;Triton X-100 PBS（0.1%）was used to lysis RAW264.7 macrophages infected with living bacteria at the given time point,then the living bacterial number was counted by bacterial culture.Results Salmonella typhimurium infected in macrophages continued to proliferate within 12 hours.The expression of iNOS in RAW264.7 macrophages was promoted by both living and inactivated Salmonella typhimurium（P〈0.05）,but to an increase mRNA level for inactivated bacterium as compared with living bacterium;Nitric oxide increased in macrophages after bacterial infection（P〈0.05）.Conclusions Salmonella typhimurium infection can effectively promote the iNOS expression of macrophages;living Salmonella typhimurium wild strain infected in macrophages can inhibit the expression of iNOS and the production of nitric oxide.