摘要
目的研究内毒素和烧伤脓毒症大鼠血清培养的巨噬细胞中Nrf2的表达。方法在动物模型中,将大鼠分为正常对照组、LPS组、单纯烧伤组、烧伤合并铜绿假单胞菌脓毒症组、烧伤合并金黄色葡萄球菌脓毒症组。先将后4组制备成40%面积的Ⅲ度烧伤创面,分别将金黄色葡萄球菌,铜绿假单胞菌及LPS注入大鼠腹腔内,在验证脓毒症动物模型后,感染8 h心脏穿刺收集血液制备血清备用。然后取RAW 264.7细胞为靶细胞,以不同组血清干扰8 h后,分别运用RT-PCR和Western blotting技术检测Nrf2在靶细胞中的表达情况。结果所有组Nrf2明显高于正常对照组。RT-PCR示Nrf2 mRNA表达水平在单纯烧伤组中最高(P<0.01),其次分别是内毒素组(P<0.01)、假单胞菌组(P<0.01)和葡萄球菌组(P<0.01)。Western印记示Nrf2蛋白表达在内毒素组中最高(P<0.01),其后分别是单纯烧伤组(P<0.01)、绿脓杆菌和葡萄球菌组(P<0.05)。结论当受到细菌或内毒素及内源性抗氧化系统对抗氧化应激损害干扰后,内源性抗氧化系统出现于巨噬细胞RAW264.7中,并可能作为治疗目标应用于严重烧伤合并MODS、脓毒症休克等患者中。
[ Objective] To study the expression of Nrf2 in macrophage （RAW 264.7） induced by rat bum serum infected with endotoxins and bacteria. [Methods] The experiments were divided into animal model and cell model. For animal model, Wistar rats were divided into 5 groups： normal group, LPS group, Burn group, Psedomonas group and Staphyllococcus group. The last four groups were made 40% 3 degree burn TBS. Lethal dose of staphylococcus aureus, Pseudomonas aeroginosa and LPS was then injected on burnt mouse peritoneum according to above group. Blood collected from all groups by cardiac puncture at 8 hours and centrifuged to collect serum. For cell model experiment, Macrophage 264.7 was target cell and was interfered with 8 hours blood serum and separately done RT-PCR and western blotting to study the expression of Nrf2 in that target cells. [ Results] There is prominent expression of Nrf2 antibody in both RT-PCR and western blotting experiment in 4 groups compared to normal group. In RT-PCR Nrf2 mRNA expression level is high- est in Bum group （81.19±3.83247）, then LPS group （71.8825±2.24292）, Pseudomonas group （65.70752±97473）, Staphylococcus group （60.525±3.5116） and Normal group （40.985±3.09207） respectively （P〈0.05）. In western blotting experiment Nrf2 protein expression is highest in LPS group （97.733±3.274）, then burn group （89.8±2.9）, pseudomonas group （80.066±2.059）, staphylococcus group （73.333±3.082）and normal group （68.333±3.521） respectively （P 〈0.05）. [ Conclusion] The expression of Nrf2 shows that endogenous antioxidative system is present inside the macrophage RAW264.7 cell interfered with bacteria and LPS and protect against oxidative stress injury through endogenous antioxidant system. And it can be a therapeutic goal for severe burn patient complicating MODS, septic shock etc.
出处
《中国现代医学杂志》
CAS
CSCD
北大核心
2012年第35期32-37,共6页
China Journal of Modern Medicine
关键词
NRF2
RAW264
7
脓毒症
内毒素
氧化应激损伤
内源性抗氧化系统
Nrf2
RAW264.7
endotoxin
staphylococcus aureus
pseudomonas aeroginosa
oxidative stress injury
endogenous antioxidant system
