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目的探讨苦参碱对慢性移植物抗宿主病狼疮样小鼠肾组织单核细胞趋化蛋白1(MCP-1)、核因子B表达及肾小管-间质纤维化的影响。方法 6周龄Balb/c(B/C,H-2d)雌性小鼠与C57BL/6(B6,H-2b)雄性小鼠自行交配,获得杂交第一代雌性小鼠[(B/C×B6)F1(H-2d/b)]。无菌取出Balb/c雌性小鼠的胸腺、淋巴结和脾脏,分离淋巴细胞,按每只鼠5×107个活细胞,分别于实验的第1、4、7、10天免疫F1小鼠,末次免疫后,F1的小鼠被随机分为模型组、苦参碱组各23只,苦参碱组每天按30mg.kg-1.d-1腹腔注射苦参碱生理盐水,模型组、正常对照组予等量生理盐水注射,分别于实验的第8、16周末,随机取3组的小鼠各8只,收集24h尿量,采血取肾,检测24h尿蛋白定量和血肌酐,采用酶联免疫吸附法定量测定小鼠血清抗双链DNA(anti-ds-DNA)抗体、抗核抗体(ANA)、白细胞介素(IL)-6、肿瘤坏死因子(TNF)-α;用PAS染色、免疫组化技术检测肾小球和肾小管间质中的MCP-1、核因子(NF)-κB蛋白的表达和分布,采用HPIAS-1000型全自动彩色图像分析系统,对免疫组化染色阳性信号进行扫描,半定量分析各组阳性表达的光密度值。结果治疗组小鼠的24h尿蛋白定量、血清肌酐与同期病理组比较,有明显的改善(P<0.05);anti-dsDNA、ANA、IL-6、TNF-α显著低于同期病理组(P<0.01);肾组织MCP-1、NF-κB表达较正常肾组织显著增高,但较同期病理组弱(P<0.05);肾小球硬化指数和肾小管间质损伤指数低于病理组。结论慢性移植物抗宿主病小鼠模型,具有狼疮性肾炎的典型特征,苦参碱可降低血清anti-dsDNA、ANA、IL-6、TNF-α的水平,对杂交F1代小鼠自身免疫反应有一定的抑制作用,并抑制NF-κB的活化,下调IL-6、TNF-α、MCP-1的表达,延缓肾小球硬化和肾小管间质纤维化的进展。
Objective To investigate the effects of matrine on the expression of monocyte chemoattractant protein-1 (MCP-1), nuclear factor-B and tubulointerstitial fibrosis in renal tissue of chronic graft-versus-host-disease-resistant lupus-like mice. Methods Six female Balb / c (B / C, H-2d) female mice were crossed with C57BL / 6 male mice (B6, H-2b) B6) F1 (H-2d / b)]. Thymus, lymph nodes and spleens of Balb / c female mice were aseptically removed, lymphocytes were isolated, and 5 × 107 viable cells per mouse were used to immunize F1 mice on the 1st, 4th, 7th and 10th day respectively After immunization, F1 mice were randomly divided into model group, matrine group, each 23 matrine group by daily intraperitoneal injection of 30mg.kg-1.d-1 matrine saline, model group, normal control The rats were randomly divided into normal saline injection, 8 mice at the end of the experiment, 8 mice in each group were randomly selected, collected 24h urine volume, blood taken kidney, 24h urine protein and serum creatinine were detected, The anti-ds-DNA, anti-nuclear antibody (ANA), interleukin (IL) -6 and tumor necrosis factor (TNF) Immunohistochemistry was used to detect the expression and distribution of MCP-1 and NF-κB in glomeruli and tubulointerstitial tissues. The immunohistochemical staining was performed on HPIAS-1000 automatic color image analysis system The signals were scanned and semiquantitatively analyzed for the optical density value of each group. Results The 24h urine protein and serum creatinine in the treatment group were significantly improved compared with those in the same period (P <0.05), and the levels of anti-dsDNA, ANA, IL-6 and TNF- <0.01). The expressions of MCP-1 and NF-κB in renal tissue were significantly higher than those in normal kidney tissue, but were weaker than those in the same period (P <0.05). The glomerular sclerosis index and tubulointerstitial injury index were lower than those in pathological group. CONCLUSION: Chronic graft-versus-host disease is a typical model of lupus nephritis in mice. Matrine can reduce the level of anti-dsDNA, ANA, IL-6 and TNF- Inhibited the activation of NF-κB, down-regulated the expression of IL-6, TNF-α and MCP-1 and delayed the progression of glomerulosclerosis and tubulointerstitial fibrosis.