巨噬细胞浸润在肾小管间质纤维化和上皮间质转化中的作用及意义

发布时间：2018-06-16 18:49

本文选题：IgA肾病 + 肾小管间质纤维化　；参考：《安徽医科大学》2016年硕士论文

【摘要】：目的：探讨巨噬细胞在肾小管间质纤维化(Tubulointerstitial fibrosis, TIF)及上皮细胞间充质转化(Epithelial-Mesenchymal Transition, EMT)中的作用及可能机制,为临床上肾脏疾病针对TIF的治疗提供理论依据。方法：(1)收集安徽医科大学第二附属医院病理科56例肾脏穿刺活检诊断为IgA肾病患者标本,用HE和特殊染色PAS、PASM、Masson染色法检测肾间质病理变化,根据肾间质纤维化及肾小管萎缩程度将56例病例分为轻、中、重度三组,另收集5例肾肿瘤远离肿瘤的正常肾组织为实验对照组。采用免疫组化方法,用CD68标记巨噬细胞,CD206标记M2型巨噬细胞,同时检测EMT相关蛋白TGF-β1、Vimentin、SMA、E-Cadherin的表达,并收集患者详细临床资料：(2)人肾小管上皮细胞HK-2与不同浓度巨噬细胞U937共培养。将实验分为五组：空白对照组(HK-2细胞单独培养),实验1组(HK-2细胞与1×104个/ml浓度的U937细胞共培养),实验2组(HK-2细胞与1×105个/ml浓度的U937细胞共培养),实验3组(HK-2细胞与1×106个/ml浓度的U937细胞共培养),实验4组(HK-2细胞与1×107个/ml浓度的U937细胞共培养)。倒置显微镜下观察细胞形态变化,采用Western blot方法检测EMT相关蛋白α-SMA、 Vimentin及E-cadherin的表达。结果：(1) CD68、CD206阳性细胞在TIF重度组高于中度组,在TIF中度组高于TIF轻度组(P0.05)；CD68阳性细胞、CD206阳性细胞与TGF-β1、SMA、Vimentin蛋白的表达呈正相关(P0.05)；与E-Cadherin蛋白的表达负相关(P0.05)；与肾功能指标血肌酐、尿素氮及尿蛋白正相关。TGF-β1、SMA、Vimentin蛋白的表达与肾小管间质纤维化程度及肾功能指标血肌酐、尿素氮及尿蛋白正相关(P0.05);E-Cadherin的表达与肾小管间质纤维化程度及上述肾功能指标呈负相关(P0.05)。(2)与U937细胞共培养24h后,HK-2细胞形态由铺路石样变为长梭形,细胞间隙明显增宽；与对照组比较,与U937细胞共培养后HK-2细胞α-SMA、Vimentin蛋白的表达明显增加,E-cadherin蛋白的表达明显下降,差异具有统计学意义(P0.05)。结论：巨噬细胞在肾脏疾病中可能具有促进肾间质纤维化及肾小管上皮细胞EMT的作用。
[Abstract]:Objective: to investigate the role and possible mechanism of macrophages in tubulointerstitial fibrosis (TIFs) and epithelial mesenchymal transition (EMTT). Methods the specimens of 56 patients with IgA nephropathy diagnosed by renal biopsy were collected from Department of Pathology, second affiliated Hospital of Anhui Medical University. The pathological changes of renal interstitium were detected by HE and PAS-PASMMA-Masson staining. According to the degree of interstitial fibrosis and atrophy of renal tubules, 56 cases were divided into three groups: mild, moderate and severe. M 2 macrophages were labeled with CD68 and CD206, and the expression of EMT associated protein TGF- 尾 1 and VimentinnSMAE-Cadherin was detected by immunohistochemistry. The detailed clinical data of human renal tubular epithelial cells (HK-2) were collected and co-cultured with different concentrations of macrophages (U937). The experiment was divided into five groups: the blank control group was cultured alone, the experimental group 1 was co-cultured with 1 脳 104 / ml U937 cells, the second group was co-cultured with 1 脳 105 / ml U937 cells, and the third group was treated with U937 cells at a concentration of 1 脳 105 / ml. The cells were co-cultured with 1 脳 106 / ml U937 cells and 1 脳 107 / ml U937 cells. The expression of EMT related proteins 伪 -SMA, vimentin and E-cadherin were detected by Western blot. Results (1) the positive cells of CD68 and CD206 were positively correlated with the expression of TGF- 尾 1 SMAMA-Vimentin protein and negatively correlated with the expression of E-Cadherin protein in TIF severe group, and in TIF moderate higher than that in TIF mild TIF group (P 0.05), and the expression of TGF- 尾 _ 1SMAMA-Vimentin protein was negatively correlated with the expression of E-Cadherin protein (P0.05A) in TIF group and TGF- 尾 _ 1 SMAMA-Vimentin protein. The expression of Vimentin protein was positively correlated with serum creatinine, urea nitrogen and urine protein. The expression of Vimentin protein was correlated with the degree of renal tubulointerstitial fibrosis and the serum creatinine of renal function index. The expression of E-Cadherin in U937 cells and U937 cells were negatively correlated with the degree of renal tubulointerstitial fibrosis and the above renal function indexes. After 24 hours of co-culture with U937 cells, the morphology of HK-2 cells changed from paving stone to long fusiform, and the intercellular gap was obviously widened. Compared with the control group, the expression of 伪 -SMA-Vimentin protein in HK-2 cells increased significantly after co-culture with U937 cells, and the expression of E-cadherin protein decreased significantly (P 0.05). Conclusion: macrophages may play a role in promoting renal interstitial fibrosis and EMT in renal tubular epithelial cells.
【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R692

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