小鼠肠上皮细胞岩藻糖基化与新生儿坏死性小肠结肠炎发生的关系  

Relations of the fucosylation of intestinal epithelial cells to the onset of neonatal necrotizing enterocolitis

作　　者：杜华 余香 余加林[1] 胡坤 贺雨[1] 肖洒[1] 艾青[1] 刘东 DU Hua;SHE Xiang;YU Jia-lin;HU Kun;HE Yu;XIAO Sa;AI Qing;LIU Dong(Department of Neonatology,Children's Hospital of Chongqing Medical University,Ministry of Education Key Laboratory of Child Development Disease Research,Chongqing Key Laboratory of Pediatrics,China International Science and Technology Cooperation BaseyorChild Development and Critical Disorders,Chongqing 400014,China;Department ofNeonatology,Shenzhen People's Hospital,Shenzhe,Cuang,long 518020,China)

机构地区：[1]重庆医科大学附属儿童医院新生儿科,儿童发育疾病研究教育部重点实验室,儿科学重庆市重点实验室,儿童发育重大疾病国家国际科技合作基地,重庆400014 [2]深圳市人民医院新生儿科,广东深圳518020

出　　处：《解放军医学杂志》2018年第10期828-833,共6页Medical Journal of Chinese People's Liberation Army

基　　金：国家自然科学基金(81571483,81370744);重庆医科大学附属儿童医院临床研究项目[(2014)254-lcyj2014-11];深圳市卫生和计划生育委员会博士创新项目(201605003)

摘　　要：目的观察肠上皮细胞岩藻糖基化水平在新生儿坏死性小肠结肠炎(NEC)小鼠模型中的变化,探讨其可能的机制。方法将42只10日龄的C57BL/6新生小鼠按随机数字表法分为NEC组(n=21,采用人工喂养+缺氧+冷刺激方法建立NEC模型)和对照组(n=21,母鼠喂养,不做处理),建模3d后处死。采用NEC病理损伤评分评估建模效果;采用流式细胞技术检测岩藻糖基化肠上皮细胞(F-ECs)和肠道固有层3型天然淋巴细胞(ILC3s)比例;采用实时荧光定量PCR检测肠上皮细胞白细胞介素-22受体(IL-22R)、岩藻糖基转移酶2(Fut2)和固有层淋巴细胞白细胞介素-22(IL-22)的表达水平;采用ELISA法检测固有层淋巴细胞IL-22蛋白的表达水平。结果成功建立NEC小鼠模型。流式细胞检测结果显示,NEC模型组F-ECs百分比低于对照组,差异有统计学意义(P<0.05);NEC模型组肠道固有层ILC3s百分比也明显低于对照组,差异有统计学意义(P<0.001)。实时荧光定量PCR结果显示,NEC模型组上皮细胞IL-22R、Fut2 mRNA表达水平明显低于对照组(P<0.001),固有层淋巴细胞IL-22 mRNA表达水平也明显低于对照组,差异有统计学意义(P<0.01)。NEC模型组固有层淋巴细胞IL-22蛋白表达水平低于对照组,差异有统计学意义(P<0.001)。结论肠上皮细胞岩藻糖基化参与了NEC小鼠模型的发病,其机制可能与ILC3s-IL-22-Fut2轴有关。Objective To investigate the changes of fucosylation level of intestinal epithelial cells in the mouse model of neonatal necrotizing enterocolitis(NEC), and explore it’s possible mechanism. Methods Forty-two 10-day old C57 BL/6 mice were randomly divided into NEC group(n=21, NEC model was established by artificial feeding, hypoxia and cold stimulation) and control group(n=21, mother feeding and no treatment), and mice were sacrificed 3 days after modeling. NEC pathological lesion scoring was performed to assess the modeling effect. Flow cytometry was used to detect the proportion of fucosylated intestinal epithelial cells(F-ECs) and intestinal lamina propria group 3 innate lymphocytes(ILC3 s). The levels of interleukin-22 receptor(IL-22 R), fucosyltransferase 2(Fut2) of intestinal epithelial cells and interleukin-22(IL-22) of lamina propria lymphocyte were detected by quantitative real-time PCR. The level of IL-22 protein in lamina propria was detected by enzyme-linked immunosorbent assay(ELISA). Results The NEC mouse model was successfully established. Flow cytometry results showed that the percentage of F-ECs was lower in NEC model group than in control group(P<0.05). The percentage of intestinal ILC3 s was also significantly lower in NEC model group than in control group with statistically significant difference(P<0.001). The results of quantitative real-time PCR showed that the mRNA levels of IL-22 R and Fut2 were significantly lower in epithelial cells of NEC model group than in control group(P<0.001). The mRNA level of IL-22 was also significantly lower in lamina propria lymphocytes of NEC model group than in control group(P<0.01). The expression level of IL-22 protein in lamina propria lymphocytes was lower in NEC model group than in control group(P<0.001). Conclusion Decreased fucosylation of intestinal epithelial cells is involved in the pathogenesis of NEC mouse model, and the mechanism may be related to the ILC3 s-IL-22-Fut2 axis.

关 键 词：岩藻糖基化 新生儿坏死性小肠结肠炎 3型天然淋巴细胞 岩藻糖基转移酶2 

分 类 号：R722.13[医药卫生—儿科]