库普弗细胞在小鼠日本血吸虫肝病发展过程中的表型变化
发布时间：2019-01-09 09:22
【摘要】：目的研究库普弗细胞(Kupffer cell)在小鼠日本血吸虫(Schistosoma japonicum)肝病发展过程中的表型变化。方法将20只6周龄的BALB/c雄性小鼠经腹部皮肤感染16条日本血吸虫尾蚴,在感染后0、21、32、42、52 d分别处死小鼠,取肝脏组织,HE染色和Masson三色染色观察肝脏病理变化,实时定量PCR(q PCR)检测肝脏Th1型细胞因子γ干扰素(interferon-γ,IFN-γ)、α肿瘤坏死因子(tumor necrosis factor-α,TNF-α)、Th2型细胞因子白细胞介素-4(interleukin-4,IL-4)、IL-13、IL-10)以及库普弗细胞的M1型巨噬细胞分化标志物诱导型一氧化氮合酶(inducible nitric oxide synthetase,i NOS)、白细胞分化抗原16(cluster of differentiation 16,CD16)、IL-6和M2型巨噬细胞分化标志物精氨酸酶-1(arginase 1,Arg-1)、CD206、IL-10的表达。体外培养库普弗细胞系,分别给予0、5、25、50 ng/ml IFN-γ或IL-4刺激12 h,或者给予25 ng/ml IFN-γ或IL-4分别刺激0、12、24、36 h后,q PCR检测库普弗细胞M1型、M2型巨噬细胞分化标志物。结果HE染色和Masson三色染色结果显示,小鼠感染后32 d肝组织中开始有虫卵沉积,42 d有明显的肉芽肿和纤维化病变。q PCR结果显示,与感染后0 d相比,IFN-γ在32 d表达水平最高,相对表达量为29.243±3.245,52 d时迅速下降,为8.923±3.002。IL-4在42 d最高,为25.521±4.957。IL-13在52 d最高,为50.793±9.631(均P0.05)。i NOS在42 d表达水平上升,相对表达量为2.950±0.321,在52 d下降,为1.783±0.319。Arg-1在52 d最高,为2.003±0.152(均P0.05)。0、5、25、50 ng/ml IFN-γ刺激库普弗细胞12 h后,i NOS、IL-6和CD16的相对表达量分别为54.690~68.577、1.887~2.427、2.417~2.787(均P0.05)。25 ng/ml IFN-γ刺激0、12、24、36 h后,i NOS、IL-6和CD16的相对表达量分别为34.810~109.210、10.327~15.143、1.887~3.317(均P0.05),而Arg-1与对照组相比无明显变化。0、5、25、50 ng/ml IL-4刺激库普弗细胞12 h后,Arg-1、IL-10和CD206的相对表达量分别为9.153~24.253、1.923~3.687和37.770~72.133(均P0.05);25 ng/ml IL-4刺激0、12、24、36 h后Arg-1、IL-10和CD206的相对表达量分别为3.563~12.613、1.637~2.673和19.732~71.943(均P0.05),而i NOS无明显变化。结论在日本血吸虫感染早期,库普弗细胞以M1型为主;而在感染中晚期,库普弗细胞以M2型为主,表明库普弗细胞转变与肝脏免疫微环境密切相关。
[Abstract]:Objective to study the phenotypic changes of (Kupffer cell) in mouse Schistosoma japonicum (Schistosoma japonicum) liver disease. Methods Twenty 6-week-old male BALB/c mice were infected with 16 cercariae of Schistosoma japonicum through abdominal skin. The mice were killed on day 0, 21, 32 and 42 respectively. Liver tissue, HE staining and Masson trichrome staining were used to observe the pathological changes of the liver. Real time quantitative PCR (q PCR) was used to detect interferon- 纬 (IFN- 纬), (tumor necrosis factor- 伪 (TNF- 伪) and interleukin-4,IL-4 (Th2 type cytokine) in liver. IL-13,IL-10) and inducible nitric oxide synthase (inducible nitric oxide synthetase,i NOS),) leukocyte differentiation antigen 16 (cluster of differentiation 16 CD16 in Kupffer cells. Expression of IL-6 and M 2 macrophage differentiation marker arginase 1 (arginase 1 Arg 1) and CD206,IL-10. The Kupffer cell line was cultured in vitro and stimulated for 12 h with 0 ~ 5U 2550 ng/ml IFN- 纬 or IL-4, or with 25 ng/ml IFN- 纬 or IL-4 for 36 h., q PCR was used to detect the M1 type of Kupffer cell line. M 2 macrophage differentiation markers. Results the results of HE staining and Masson trichrome staining showed that there was egg deposition in the liver tissue of mice 32 days after infection, and obvious granuloma and fibrosis on 42 days after infection. The results showed that compared with 0 days after infection, there were obvious granuloma and fibrosis in the liver of mice. The expression level of IFN- 纬 was the highest at 32 days, and the relative expression of IFN- 纬 decreased rapidly at 52 days after 29.243 卤3.245 days, 8.923 卤3.002.IL-4 at 42 days, 25.521 卤4.957.IL-13 at 52 days. 50.793 卤9.631 (P0.05) the expression level of). I NOS increased at 42 days, the relative expression level was 2.950 卤0.321, decreased at 52 days, 1.783 卤0.319.Arg-1 was highest at 52 days, 2.003 卤0.152 (P 0.05). After stimulated by 2550 ng/ml IFN- 纬 for 12 h, the relative expression of I NOS,IL-6 and CD16 in Kupffer cells was 54.690, 68.5771.887 and 2.4272.4172.787, respectively (P0.05). 25 ng/ml IFN- 纬 stimulated, i NOS, for 36 h. The relative expression levels of IL-6 and CD16 were 34.81010 ~ 109.21010 ~ (10.327) ~ 15.143n ~ (1.887) ~ 3.317 (P0.05), respectively, but Arg-1 had no significant change compared with the control group. After 12 h stimulation of Kupffer cells with Arg-1, Arg-1, was observed. The relative expression levels of IL-10 and CD206 were 9.153 and 37.70 respectively (P 0.05). The relative expression levels of Arg-1,IL-10 and CD206 were 3.563 ~ 12.613 ~ 1.637 ~ 2.673 and 19.732n ~ (71.943), respectively, 36 h after 25 ng/ml IL-4 stimulation (P0.05), but I NOS did not change significantly. Conclusion in the early stage of Schistosoma japonicum infection, the Kupffer cells are mainly M1 type, while in the middle and late stage of infection, the Kupffer cells are mainly M2 type, which indicates that the Kupffer cell transformation is closely related to the liver immune microenvironment.
【作者单位】： 徐州医科大学病原生物学与免疫学教研室江苏省免疫与代谢重点实验室;第二军医大学热带传染病学教研室;
【分类号】：R532.21
，
本文编号：2405437
[Abstract]:Objective to study the phenotypic changes of (Kupffer cell) in mouse Schistosoma japonicum (Schistosoma japonicum) liver disease. Methods Twenty 6-week-old male BALB/c mice were infected with 16 cercariae of Schistosoma japonicum through abdominal skin. The mice were killed on day 0, 21, 32 and 42 respectively. Liver tissue, HE staining and Masson trichrome staining were used to observe the pathological changes of the liver. Real time quantitative PCR (q PCR) was used to detect interferon- 纬 (IFN- 纬), (tumor necrosis factor- 伪 (TNF- 伪) and interleukin-4,IL-4 (Th2 type cytokine) in liver. IL-13,IL-10) and inducible nitric oxide synthase (inducible nitric oxide synthetase,i NOS),) leukocyte differentiation antigen 16 (cluster of differentiation 16 CD16 in Kupffer cells. Expression of IL-6 and M 2 macrophage differentiation marker arginase 1 (arginase 1 Arg 1) and CD206,IL-10. The Kupffer cell line was cultured in vitro and stimulated for 12 h with 0 ~ 5U 2550 ng/ml IFN- 纬 or IL-4, or with 25 ng/ml IFN- 纬 or IL-4 for 36 h., q PCR was used to detect the M1 type of Kupffer cell line. M 2 macrophage differentiation markers. Results the results of HE staining and Masson trichrome staining showed that there was egg deposition in the liver tissue of mice 32 days after infection, and obvious granuloma and fibrosis on 42 days after infection. The results showed that compared with 0 days after infection, there were obvious granuloma and fibrosis in the liver of mice. The expression level of IFN- 纬 was the highest at 32 days, and the relative expression of IFN- 纬 decreased rapidly at 52 days after 29.243 卤3.245 days, 8.923 卤3.002.IL-4 at 42 days, 25.521 卤4.957.IL-13 at 52 days. 50.793 卤9.631 (P0.05) the expression level of). I NOS increased at 42 days, the relative expression level was 2.950 卤0.321, decreased at 52 days, 1.783 卤0.319.Arg-1 was highest at 52 days, 2.003 卤0.152 (P 0.05). After stimulated by 2550 ng/ml IFN- 纬 for 12 h, the relative expression of I NOS,IL-6 and CD16 in Kupffer cells was 54.690, 68.5771.887 and 2.4272.4172.787, respectively (P0.05). 25 ng/ml IFN- 纬 stimulated, i NOS, for 36 h. The relative expression levels of IL-6 and CD16 were 34.81010 ~ 109.21010 ~ (10.327) ~ 15.143n ~ (1.887) ~ 3.317 (P0.05), respectively, but Arg-1 had no significant change compared with the control group. After 12 h stimulation of Kupffer cells with Arg-1, Arg-1, was observed. The relative expression levels of IL-10 and CD206 were 9.153 and 37.70 respectively (P 0.05). The relative expression levels of Arg-1,IL-10 and CD206 were 3.563 ~ 12.613 ~ 1.637 ~ 2.673 and 19.732n ~ (71.943), respectively, 36 h after 25 ng/ml IL-4 stimulation (P0.05), but I NOS did not change significantly. Conclusion in the early stage of Schistosoma japonicum infection, the Kupffer cells are mainly M1 type, while in the middle and late stage of infection, the Kupffer cells are mainly M2 type, which indicates that the Kupffer cell transformation is closely related to the liver immune microenvironment.
【作者单位】： 徐州医科大学病原生物学与免疫学教研室江苏省免疫与代谢重点实验室;第二军医大学热带传染病学教研室;
【分类号】：R532.21
，
本文编号：2405437
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