不同极化类型巨噬细胞参与肾纤维化的动物实验研究

Effect of macrophage polarization on interstitial fibrosis in mouse unilateral ureteral obstruction model

目的 观察单侧输尿管结扎（UUO）模型小鼠肾脏间质纤维化程度,巨噬细胞浸润及极化的改变.方法 8～ 10周龄C57BL/6J雄性小鼠12只,采用单侧输尿管结扎的方法建立UUO模型,分别于手术后第7、14天处死动物,留取肾组织标本.Masson染色法检测胶原组织的沉积,实时定量PCR法检测α平滑肌肌动蛋白（α-SMA）、I型胶原（Col-1） mRNA表达;免疫荧光染色法检测肾间质巨噬细胞浸润程度及极化的表达变化.结果 Masson染色结果显示：与对照组相比,实验组小鼠肾组织胶原纤维沉积增多;与7d组相比,14 d组胶原纤维沉积增多（均P＜ 0.05）.免疫荧光染色结果显示：与对照组相比,UUO组小鼠肾组织α-SMA表达阳性的细胞增多;与7d组相比,14 d组小鼠α-SMA阳性的细胞增多（均P＜0.05）.实时定量PCR结果显示：与对照组相比,实验组肾组织α-SMA、Col-1 mRNA表达增加;与7d组相比,14 d组小鼠肾组织α-SMA、Col-1 mRNA表达增加（均P＜0.05）.与对照组相比,术后14 d组小鼠M2型巨噬细胞亦增多（P＜0.05）;两组M1型巨噬细胞浸润数目的差异无统计学意义.结论 UUO诱导的肾间质纤维化模型肾组织中的巨噬细胞浸润明显增加,主要以M2型巨噬细胞浸润为主,推测M2参与了肾脏纤维化的形成.

Objective To investigate the effect of macrophage polarization on tubulointerstitial fibrosis of mouse unilateral ureteral obstruction（UUO） model.Methods Twelve male C57BL/6J mice were employed,each of which with an age of 8 to 10 weeks.UUO model was established with these mice with the method of unilateral ureteral ligation.Mice were then sacrificed on the 7th and 14th day respectively after operation,and renal tissue specimens were obtained.The authors detected collagen deposition by Masson staining,and alpha smooth muscle actin （alpha SMA） as well as collagen type Ⅰ （Coll-1） mRNA by real-time quantitative PCR.The authors also detected the degree of renal interstitial macrophages infiltration and expression changes of polarization by immunofluorescence staining.Results Compared with the mice that were observed on the 7th day after operation,the degree of renal interstitial fibrosis in mice observed on the 14th day after operation was comparatively serious,the difference shown by semi-quantitative results was statistically significant （P ＜ 0.05）.Moreover,mice observed on the 14th day after operation have more M2 macrophages,the difference between two groups of mice was statistically significant （P ＜ 0.05）.On the contrary,there was no statistically significant difference in the degree of M1 macrophages infiltration between these two groups of mice.Conclusions In the renal interstitial fibrosis model induced by UUO,the degree of macrophage infiltration increased significantly,mainly resulted from M2 macrophage infiltration,suggesting that M2 macrophages were involved in the formation of renal fibrosis.