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辅助性T淋巴细胞22及其效应因子IL-22在自身免疫性肝炎小鼠模型中的表达及意义
Expression and significance of Th22 cells and its functional factor interleukin-22 in mice with autoimmune hepatitis
文章发布日期:2018年07月06日  来源:  作者:潘景,潘文婷,吴丽慧  点击次数:417次  下载次数:80次

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【摘要】:目的研究自身免疫性肝炎(AIH)小鼠模型中辅助性T淋巴细胞(Th)22细胞水平及其功能因子IL-22的表达情况,探索其在AIH发病中的意义。方法选取30只4周龄雄性C57BL/6小鼠随机分为对照组(n=6)、AIH组(n=14),剩余10只用于提取肝脏特异性抗原S100。在实验第1天和第7天通过腹腔注射法将新鲜提取的S100与等体积弗氏完全佐剂混合液注射到小鼠腹腔内,第28天成功建立AIH小鼠模型(造模期间AIH组有7只小鼠因注射不当、出现腹水、感染等原因死亡)后,将小鼠经腹腔注射5%水合氯醛(0.1 ml/10 g)麻醉后摘取眼球取血,收集小鼠脾脏采用流式细胞术检测Th22细胞数,RT-PCR检测AHR mRNA水平;收集肝组织用于HE染色观察肝脏病理变化,RT-PCR和免疫印迹法检测肝组织IL-22、IL-22R表达情况;ELISA法检测血清中IL-22细胞因子水平。2组间比较采用t检验。结果对照组小鼠肝组织排列整齐,肝小叶结构清晰,AIH组小鼠汇管区有大量炎症细胞浸润,肝组织出现点状甚至碎片状坏死。AIH组Th22细胞数较对照组明显升高[(0.083±0.052)% vs(1.555±0.812)%,t=-4.405,P<0.05]。AIH组小鼠脾脏中Th22转录因子AHR mRNA的相对表达量较对照组有明显升高(0.485±0.096 vs 1.268±0.366,t=-5.452,P<0.05)。AIH组肝组织中IL-22 mRNA和IL-22R mRNA的相对表达量均较对照组明显升高(IL-22 mRNA:1.146±0.227 vs 3.813±0.478,t=-12.458 ,P<0.001;IL-22R mRNA:0.276±0.037 vs 1.734±0.248,t=-15.333,P<0.001);AIH组肝组织的IL-22及IL-22R的蛋白相对表达量均明显高于对照组(1.040±0.261 vs 0.410±0.077,t=-6.324,P<0.05;1.432±0.304 vs 0.830±0.146,t=-6.316,P<0.05)。AIH组血清IL-22水平较对照组升高,且差异有统计学意义(t=-15.799,P<0.05)。结论AIH小鼠Th22细胞及IL-22因子的表达水平均明显升高,其可能在AIH的发病中起重要作用。
【Abstract】:ObjectiveTo investigate the expression of Th22 cells and its functional factor interleukin-22 (IL-22) in a mouse model of autoimmune hepatitis (AIH) and the significance of Th22 cells and IL-22 in the pathogenesis of AIH. MethodsA total of 30 male C57BL/6 mice aged 4 weeks were randomly divided into control group with 6 mice and AIH group with 14 mice. Another 10 mice were used to extract liver-specific antigen S100. On days 1 and 7 of the experiment, the freshly extracted S100 was injected into the abdominal cavity of the mice together with an equal volume of Freund′s complete adjuvant, and a mouse model of AIH was successfully established on day 28 (7 mice in the AIH group died during model establishment due to improper injection, presence of ascites, or infection). The mice were given intraperitoneal injection of 5% chloral hydrate (0.1 ml/10 g) for anesthesia, and then eyeballs were removed to collect blood samples. The spleen was collected, and flow cytometry was performed to measure the percentage of Th22 cells and RT-PCR was performed to measure the mRNA expression of aryl hydrocarbon receptor (AHR). The liver tissue was collected; HE staining was used to observe liver pathological changes, and RT-PCR and Western blotting were used to measure the expression of IL-22 and IL-22 receptor (IL-22R) in liver tissue. ELISA was used to measure the serum level of IL-22. The t-test was used for comparison between the two groups. ResultsHE staining showed that the mice in the control group had well-arranged liver tissue and clear structures of hepatic lobules, while those in the AIH group had massive inflammatory cell infiltration in the portal area and dotted and even piecemeal necrosis in liver tissue. The AIH group had a significant increase in the percentage of Th22 cells than the control group (0.083%±0.052% vs 1.555%±0.812%, t=-4.405, P<0.05). Compared with the control group, the AIH group had a significant increase in the relative mRNA expression of AHR (0.485±0.096 vs 1.268±0.366, t=-5.452, P<0.05). Compared with the control group, the AIH group had significant increases in the relative mRNA expression of IL-22 (1.146±0.227 vs 3.813±0.478, t=-12.458, P<0.001) and IL-22R (0.276±0.037 vs 1.734±0.248, t=-15.333, P<0.001) and significantly higher relative protein expression of IL-22 (1.040±0.261 vs 0.410±0.077, t=-6.324, P<0.05) and IL-22R (1.432±0.304 vs 0.830±0.146, t=-0.631, P<0.05) in liver tissue. The AIH group had a significantly higher level of IL-22 than the control group (t=-15.799, P<0.05). ConclusionAIH mice have significant increases in the percentage of Th22 cells and the expression of IL-22, which may play an important role in the pathogenesis of AIH.
【关键字】:肝炎, 自身免疫性; T淋巴细胞, 辅助诱导; 白细胞介素22; 小鼠, 近交C57BL
【Key words】:hepatitis, autoimmune; T-lymphocytes, helper-inducer; interleukin-23; mice, inbred C57BL
【引证本文】:factor interleukin-22 in mice with autoimmune hepatitis[J]. J Clin Hepatol, 2018, 34(8): 1745-1749. (in Chinese)
潘景, 潘文婷, 吴丽慧. 辅助性T淋巴细胞22及其效应因子IL-22在自身免疫性肝炎小鼠模型中的表达及意义[J]. 临床肝胆病杂志, 2018, 34(8): 1745-1749.

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