姥鲛烷诱导法建立小鼠狼疮肾炎模型

Development and study of experimental C57BL/6 mouse lupus nephritis model induced by pristane

目的:采用姥鲛烷诱导建立C57BL/6小鼠狼疮肾炎模型,并进行生物学鉴定和形成机制的探讨。方法:模型组小鼠单次腹腔注射0. 5 mL姥鲛烷,对照组同方式注射等量生理盐水。注射后监控生存状态;第10天采用流式细胞术分析脾脏巨噬细胞、粒细胞、树突状细胞及B细胞的活化情况;间接免疫荧光法每月检测血清抗核抗体(ANA)及抗双链DNA(dsDNA)抗体的滴度,Albustix试纸法分析尿蛋白含量;注射后8个月处死小鼠并取肾脏,经直接免疫荧光法观测免疫复合物沉积状况,免疫组化法分析干扰素γ(IFN-γ)与白细胞介素4(IL-4)的表达,并进行HE染色及透射电镜病理检查。结果:第10天时脾脏巨噬细胞、粒细胞、树突状细胞及B细胞呈不同程度活化(P<0. 05),B细胞表面标志物B7-1、B7-2及MHC-Ⅱ表达上调(P<0. 05);3~8个月时血清ANA和抗dsDNA抗体水平进行性升高(P<0. 05),3只小鼠出现腹水,1只消瘦并最终于7个月时死亡;4个月时3只小鼠出现蛋白尿,8个月时蛋白尿阳性率100%,尿蛋白含量1~20 g/L;8个月时腹腔内出现大量的异位淋巴组织,肾小球及系膜区免疫复合物沉积严重,肾组织中IFN-γ表达量升高,淋巴细胞侵润、组织坏死、电子致密物沉积等病理损伤明显,而对照组未见明显脾脏免疫细胞活化及自身抗体、蛋白尿产生,肾脏中IFN-γ与IL-4表达正常,无明显肾脏免疫复合物沉积及病理性改变。结论:采用姥鲛烷诱导法建立了与人类狼疮肾炎症状类似的C57BL/6小鼠狼疮肾炎模型。

AIM:To establish a C57BL/6 mouse lupus nephritis model induced by pristane and to explore the pathogenic mechanism of this model.METHODS:Thirty C57BL/6 mice were randomly divided into 2 groups.The mice in model group were intraperitoneally injected with 0.5 mL pristane,while the mice in control group were injected with 0.5 mL normal saline.The state of health and survival of the animals were monitored during the whole experiment.The ac⁃tivation of macrophages,granulocytes,dendritic cells and B cells in spleen and the markers on B cells were detected by flow cytometry on the 10th day after injection.The serum levels of antinuclear antibodies(ANA)and anti-double strand DNA antibodies(anti-dsDNA)were measured every month after injection by indirect immunofluorescence assay,and pro⁃teinuria was checked mouthy by Albustix test strips.Eight months after injection,all mice were killed,and the kidneys were slided and stained with HE or PE-labeled IgG to analyze the expression of interferon-γ(IFN-γ)and interleukin-4(IL-4).The evidence of glomerulonephritis was histopathologically observed,and the ultrastructural changes of the glomerulus were determined by transmission electron microscopy.RESULTS:Ten days after the pristane injection,the populations of macrophages,granulocytes,dendritic cells and B cells were increased significantly compared with control mice(P<0.05).The expression of B7-1,B7-2 and MHC-II on the B cells was also increased(P<0.05).The serum levels of ANA and anti-dsDNA were increased from 3 to 8 months(P<0.05).Meanwhile,part of the mice showed different degrees of as⁃cites,emaciation or even death,part of the mice had proteinuria 4 months later,and 8 months later,all of the model mice had proteinuria,with 1~20 g/L of urine protein.At the 8th month,a large number of heterotopic lymphoid tissues ap⁃peared in the abdominal cavity.The deposition of immune complexes in glomeruli and mesangial areas was serious.The expression of IFN-γin the renal tissues was increased.Pathological damages such as lymphocyte infiltration,tissue necro⁃sis and deposition of electron dense substances were obvious in the model mice.In control group,autoantibodies and pro⁃teinuria was not detected under the same experimental condition.Meanwhile,no evidence of glomerulonephritis in the con⁃trol mice was observed.CONCLUSION:The C57BL/6 mouse model of lupus nephritis induced by pristane is established successfully.