轮状病毒致新生鼠胆道闭锁肝内外各亚型炎症细胞表达初探

Inflammatory cellular profiles in the liver and spleen of biliary atresia induced by rotavirus

目的研究轮状病毒感染致新生鼠胆道闭锁不同时段肝内外各亚型炎症细胞浸润状态，并初步探讨调控新生鼠胆道闭锁肝内外炎症反应的主要炎症类型。方法选取健康新生Balb／C小鼠，出生24h内使用轮状病毒腹腔注射为实验组，注射等量的轮状病毒培养液（DMEM）为对照组。注射病毒后第7d、14d分别处死新生鼠并获取胆管、肝脏及脾脏组织标本。根据新生鼠皮肤黄染、体重变化将实验组分为致胆道闭锁组（BA＋）与未致胆道闭锁组（BA-），再根据胆管HE染色结果，验证分组的准确性。利用流式细胞技术，分别检测各组在不同时段肝脏及脾脏组织内CIN＋Th、CD8＋Tc、NK细胞、巨噬细胞（Mac）及T淋巴调节细胞（Treg）的浸润状态，并进行统计学分析。结果脾脏组织：三组间炎症细胞浸润的类型及数量均无明显统计学差异（P〉0．05）；肝脏组织：轮状病毒感染7d后BA一组CD4’Th、Mac及BA＋组CIN’Th、CD8’Tc、NK、Mac、Treg与对照组相比表达明显升高（P〈0．05），BA＋组CI）8’Tc、NK、Treg表达又明显高于BA一组（P〈0．05）；感染14d后＆BA-组CIN-Th及BA＋组CD4＋Th、CD8＋Tc、NK、Treg与对照组相比表达显著升高（P〈0．05），BA＋组CD8’Tc、NK、Treg表达亦明显高于BA-组（P〈0．05）；CIN’Th在BA-组及BA＋组均表现为持续升高；CD8’Tc、Treg仅在BA＋组表现为持续升高，而Mac表现为先升高后降低。结论胆道闭锁新生鼠肝内存在特异性渐进性炎症反应，CIN’T淋巴辅助细胞、CD8＋T淋巴杀伤细胞、NK细胞、巨噬细胞及T淋巴调节细胞广泛激活并可能共同参与了这一特异性炎症反应。

Objective To explore the inflammatory cellular profiles in the liver and spleen in the rotavirus （RRV） mouse model of biliary atresia （BA）. Methods Balb/c neonatal mice were used in this experiment. Within the first 24 hours after birth, mice were intraperitoneally injected with RRV to induce BA. The control mice were injected with RRV culture medium DMEM. The mice were sac- rifieed 7 or 14 days after RRV infection. Extrahepatic bile duct, liver and spleen were harvested for further study. The RRV infected mice were divided into the biliary atresia group （BA） and non-biliary atresia group （nBA）. The diagnosis of BA was confirmed by obliteration of bile duct on H^E stai- ning. The inflammatory cellular profiles including CD4＋ T lymphocytes helper cells （CD4＋ Th）, CD8＋ cytotoxie T lymphoeytes （CD8＋ Tc）, NK cells and Maerophages （Mac） and regulatory T cells （Treg） were determined by flow eytometry. Results In the spleen, there was no significant differences of CD4＋ Th, CD8＋ Tc, Mac and Treg between the BA mice and control mice either 7 or 14 days after RRV infection （all P^0. 05）. In the liver, 7 days after RRV infection, CD4＋ Th, CD8＋ Tc, NK, Mac and Treg in BA mice were signifieantly increased than those of control mice and the nBA mice （all, P^O. 05）. Mac of the nBA mice was increased significantly than that of control mice （P~ 0. 05）. In the liver, 14 days after RRV infection, CD4 ＋ Th, CD8 ＋ Tc, NK, Mac and Treg in BA mice were significantly increased than those of control mice （all P〈0. 05）. CD8 ＋ Tc, NK, Mac and Treg in BA mice were significantly increased than those of control mice and the nBA mice （all P〈 0. 05）. CD4 ＋ Th of the nBA mice was also increased significantly than that of control miee （P〈 0. 05）. At the day 14 after infection, CD8 ＋ Tc, NK, and Treg in BA group were significantly higher than those of control group and nBA group （P%0. 05）. CD4 ＋ Th in nBA group and BA group was significantly higher than that of control group （P^0. 05）. In RRV infected mice, CD4 ＋ Th was con- sistently increased until after 14 days. However, CD8 ＋ Tc and Treg were increased until 14 days only in BA mice. Mac peaked on the 7 day after RRV infection, and then decreased. Conclusions The in- flammatory cellular profile in the live is dynamically changed in the course of BA development, which may be associated with the pathogenesis of hiliarY atresia.