巨噬细胞对单侧输尿管结扎小鼠肾纤维化的影响

Effect of macrophage on renal fibrosis in mouse model of unilateral ureter obstruction

摘要:

目的 检测不同亚型巨噬细胞在单侧输尿管结扎(UUO)模型小鼠肾组织中的表达及其对肾纤维化的影响。方法 雄性C57/BL小鼠39只,完全随机分为4组:假手术组(sham,行单侧输尿管分离术,n=9)、模型-3d组(UUO-3d,行单侧输尿管结扎术,n=10)、模型-7d组(UUO-7d,n=10)、模型-14d组(UUO-14d,n=10)。分别于UUO术后 3、7、14 d收取小鼠肾脏。病理染色观察假手术组及3个模型组小鼠肾间质纤维化及炎性细胞浸润变化。免疫荧光观察假手术组及3个模型组小鼠肾间质巨噬细胞浸润程度。流式细胞术检测肾脏巨噬细胞亚群(CD86和CD206)的比例变化。采用SPSS 20.0统计软件对数据进行分析。组间比较采用完全随机设计的单因素方差分析及SNK-q检验。结果 假手术组小鼠肾小管结构完整,肾间质清晰无炎性细胞浸润,无胶原蛋白沉积,巨噬细胞浸润数量少。与假手术组相比,模型组-3d、模型组-7d、模型组-14d小鼠肾间质胶原蛋白沉积面积[(7.8±1.5)% vs(14.1±2.7)%,(27.4±3.1)%,(39.3±2.7)%]及炎性细胞浸润[(169±16) vs(1068±164),(2159±432),(3536±318)]均增加(P<0.05)。随着UUO时间延长,巨噬细胞浸润显著增多[(20.8±2.7)%,(31.1±1.3)%,(49.5±2.1)%,(69.6±1.8)%；P<0.05],M2型巨噬细胞标志物(CD206)表达增高[(3.2±1.9)%,(34.1±2.1)%,(52.6±1.6)%,(76.7±2.3)%；P<0.05],而各组M1型巨噬细胞标志物(CD86)表达无明显差异[(76.4±3.6)%,(81.8±2.8)%,(80.6±4.4)%,(85.5±2.6)%；P>0.05]。结论 在肾纤维化发展过程中,M2型巨噬细胞浸润增加可能促进单侧输尿管结扎小鼠肾纤维化。

Abstract:

Objective To investigate the expression of macrophage subtypes in mouse model of unilateral ureter obstruction (UUO) and its role in renal fibrosis. Methods A total of 39 male C57/BL mice were divided into sham group (n=9), UUO-3d group (n=10), UUO-7d group (n=10) and UUO-14d group (n=10). Unilateral ureter separation was performed on the mice in the sham group, and unilateral ureteral ligation in the UUO groups. The kidney was obtained at 3,7 and 14 days after operation. Pathological staining was used to observe renal interstitial fibrosis and infiltration of inflammatory cells, and immunofluorescence to observe renal interstitial macrophage infiltration. Flow cytometry was performed to detect changes in the proportion of renal macrophage subtypes (CD86 and CD206). SPSS statistics 20.0 was used for data analysis, and SNK-q test for comparison. Results In the sham group, the renal tubules were structurally intact, and the renal interstitium was clear without infiltration of the inflammatory cells and collagen deposition and with infiltration of a small number of macrophages. Compared with sham group, all UUO groups (UUO-3d, -7d, -14) had larger collagen protein areas [(7.8±1.5) % vs (14.1±2.7)%, (27.4±3.1)%, (39.3±2.7)%] and more inflammatory cells [(169±16) vs (1068±164), (2159±432), (3536±318)] in renal interstitium (P<0.05). Prolonged obstruction time in the model group resulted in significantly increased infiltration of macrophages [(20.8±2.7)% vs (31.1±1.3)%, (49.5±2.1)%, (69.6±1.8)%; P<0.05]. The expression of M2 macrophage marker (CD206) also increased with prolonged UUO [(3.2±1.9)%, (34.1±2.1)%, (52.6±1.6)%, (76.7±2.3)%; P<0.05], but no significant difference was found between UUO groups in the expression of M1 macrophage marker (CD86) [(76.4±3.6)%, [JP+2](81.8±2.8)%, (80.6±4.4)%, (85.5±2.6)%; P>0.05].Conclusion Increased M2 macrophage may contribute to renal fibrosis in the mice with unilateral ureteral ligation.