IL-10基因转化的大肠杆菌对小鼠结肠炎的治疗作用

Treatment of murine colitis with IL-10-secreting Escherichia coli

目的:观察白介素-10(interleukin-10,IL-10)基因转化大肠杆菌(E.coli/mIL-10)对葡聚糖硫酸钠(DSS)诱导小鼠结肠炎的影响,并探讨其相关机制.方法:将小鼠IL-10基因序列转化至大肠杆菌(E.coli/mIL-10),阴性组为空质粒转化大肠杆菌(E.coli0).60只♀balb/c小鼠随机分成6组:正常组对照组,DSS组,DSS+E.coli/mIL-10组,DSS+E.coli0,E.coli/mIL-10组和E.coli0组.建立小鼠急性DSS结肠炎模型.自小鼠模型建立第1天开始,DSS+E.coli/mIL-10组和正常鼠+E.coli/mIL-10分别给予E.coli/mIL-10灌胃至实验结束,DSS+E.coli0组和E.coli0组分别给予E.coli0灌胃至实验结束(1×108cfu/天/只),正常对照组以及DSS组给予同等培养基灌胃至实验结束.每天观察各组疾病活动指数(DAI),并在实验结束后检测各组小鼠炎症肠段肿瘤坏死因子(TNF)、髓过氧化物酶(MPO)和核因子(NF)-κBP65的表达.结果:自实验第4天开始DSS+E.coli/mIL-10组小鼠DAI明显低于DSS组和DSS+E.coli0组.实验结束时DSS+E.coli/mIL-10组结肠组织中TNF(172.46±66.71pg/g组织)、MPO活性(2.35±0.39U/g组织)比DSS组和DSS+E.coli0组[(237.85±47.01)和(239.81±50.38)pg/g组织]、[(4.15±0.77)和(3.5±1.23)U/g组织]均明显降低;结肠组织NF-κB阳性表达减少.正常小鼠给予重组大肠杆菌灌胃后未出现结肠黏膜损伤.结论:利用经IL-10基因转化的大肠杆菌可以明显缓解DSS小鼠的结肠炎症损伤,减低MPO活性、抑制炎症肠段炎症细胞NF-κB的活化及炎性细胞因子的分泌.利用基因工程技术结合肠道共栖菌表达IL-10可以为IBD治疗提供一个新的方法.

AIM： To evaluate the therapeutic effect of interleukin-10 （IL-10）-secreting Escherichia coli （E. coli） on murine colitis. METHODS： Sixty 6-8-wk-old female BALB/c mice were divided into six groups： normal control group, dextran sulfate sodium （DSS） group, empty plasmid-transformed E.coli （E.coliO） group, IL-10 gene-transformed E.coli （E.coli/ mIL-10） group, DSS ＋ E.coliO group, and DSS ＋ E.coli/mIL-lO group. The DSS, DSS ＋ E.coliO, and DSS ＋ E.coli/mIL-lO groups were fed 5 % DSS solution for 7 d to induce colitis, while the other groups were given normal tap water. TheE.coliO and DSS ＋ E.coliO groups were intragastrically given E.coliO; the E.coli/mIL-lO and DSS ＋ E.coli/mIL-10 groups were given recombinant E.coli/mIL-lO; and the DSS and normal control groups were given LB （Luria-Bertain） medium. The treatment lasted for 8 days. Disease activity index （DAI） was recorded each day. The expression of TNF-α, NF-κB p65 and myeloperoxidase （MPO） in the colon was measured at the end of the experiment. RESULTS： Mice in the DSS ＋ E.coli/mIL-l0 group showed a lower DAI than those in the DSS and DSS ＋ E.coliO groups from day 4 to the end of the experiment. The levels of TNF-cz and MPO in the colon and the expression of NF- κB P65 in the nuclei of inflammatory cells were lower in the DSS ＋ E.coli/mIL-l0 group （172.46 pg/g ± 22.23 pg/g, 2.35 U/g ± 0.15 U/g） than in the DSS （237.85 pg/g ± 14.86 pg/g, 4.15 U/g ± 0.29 U/g） and DSS ＋ E.coliO groups （239.81 pg/g ± 50.38 pg/g, 3.5 U/g ±1.23 U/g） at the end of experiment. No colonic injury was observed in mice in the normal control, E.coli/IL-l0 and E.coliO groups. CONCLUSION： Local delivery of IL-10 genetransformed E.coli ameliorates DSS-inducd murine colitis possibly by decreasing proinflammatory cytokine production and inhibiting NF-κB activation. Gene therapy strategies using engineered E.coli encoding immunoregulatory cytokines may provide a potential approach for treatment of inflammatory bowel disease.