Objective: To observe the expressions of complement 3 (C3) and endothelial cell injury-associated proteins before and after cathepsin L (CTSL) blockade in renal injury of trichloroethylene (TCE) -sensitized mice. Methods: In June 2018, 41 SPF female BALB/c mice were divided respectively into blank control group (n=5) , vehicle control group (n=5) , TCE group (n=15) and TCE+CTSLi group (n=16) to establish trichloroethylene-sensitized mice model by pretreating the mice with intraperitoneal injection of CTSL inhibitor (CTSLi) and using TCE for the first and last challenge. According to the skin sensitization score, the mice were divided into positive group and negative group. 72 hours after the last challenge, the renal function indexes of the mice were detected, the pathological changes of mice kidneys were observed, and the glomerular C3 and endothelial cell damage-related proteins [vascular cell adhesion molecule 1 (VCAM-1) , tight junction protein 5 (Claudin-5) and Syndecan-1] expression levels were detected. Results: The sensitization rates of mice in TCE group and TCE+CTSLi group were 53.3% (8/15) and 50.0% (8/16) , respectively, and there was no significant difference between the two groups (P>0.05) . Compared with vehicle control group and the corresponding TCE negative group, the serum creatinine (CRE) and blood urea nitrogen (BUN) levels of mice in the TCE positive group was increased, while the TCE positive group were higher than the TCE+CTSLi positive group (P<0.05) . Pathological examination showed obvious vacuolar degeneration and cellular edema in the mice kidney of the TCE positive group. In the TCE+CTSLi positive group, the above pathological damage was significantly improved. Immunohistochemical results showed that the expression of glomerular C3 fragment and VCAM-1 in TCE positive group were significantly higher than that of the vehicle control and TCE negative group (P<0.05) , while TCE+CTSLi positive group was significantly lower than that of TCE positive group (P<0.05) . Western blot test results showed that the relative expression levels of Claudin-5 and Syndecan-1 protein in the mice glomeruli of TCE positive group were significantly lower than those in the vehicle control group and TCE negative group (P<0.05) . Compared with the TCE positive group, the Claudin-5 protein was increased in the kidney of the TCE+CTSLi positive group, but the difference was not statistically significant (P>0.05) , while the Syndecan-1 protein was significantly increased in the TCE+CTSLi positive group (P<0.05) . Conclusion: CTSL may mediate the glomerular structural damage by cutting complement C3, activating the complement system, damaging endothelial cell structural protein Syndecan-1 and overexpressing adhesion molecule VCAM-1 in TCE-sensitized mice. Inhibiting the expression of CTSL may be an effective way to protect the glomerular integrity of structure and function in pharmacology.
目的： 观察组织蛋白酶L（CTSL）阻断前后三氯乙烯（TCE）致敏小鼠肾小球中补体片段3（C3）和内皮细胞损伤相关蛋白的表达情况，探讨CTSL在TCE致敏小鼠肾脏损伤中的作用。 方法： 于2018年6月，将41只SPF级雌性BALB/c小鼠随机分为空白对照组（n=5）、溶剂对照组（n=5）、TCE组（n=15）和TCE+CTSLi组（n=16），建立TCE致敏小鼠模型，通过腹腔注射CTSL抑制剂（CTSLi）对小鼠进行预处理，采用TCE对小鼠进行初次激发和末次激发，根据皮肤致敏反应评分将小鼠分为致敏阳性组和致敏阴性组。于末次激发后72 h检测小鼠肾功能指标，观察小鼠肾脏组织病理学改变情况，并检测肾小球C3和内皮细胞损伤相关蛋白[血管细胞黏附分子1（VCAM-1）、紧密连接蛋白5（Claudin-5）和多配体蛋白聚糖1（Syndecan-1）]的表达水平。 结果： TCE组和TCE+CTSLi组小鼠致敏率分别为53.3%（8/15）和50.0%（8/16），两组间差异无统计学意义（P>0.05）。TCE致敏阳性组小鼠血清肌酐（CRE）、尿素氮（BUN）水平明显高于溶剂对照组和TCE致敏阴性组（P<0.05），而TCE+CTSLi致敏阳性组小鼠血清CRE、BUN水平明显低于TCE致敏阳性组（P<0.05）。TCE致敏阳性组小鼠肾脏可见明显空泡变性及细胞水肿，TCE+CTSLi致敏阳性组小鼠上述病理损害程度明显改善。TCE致敏阳性组肾小球C3片段和VCAM-1表达明显高于溶剂对照组和TCE致敏阴性组（P<0.05），而TCE+CTSLi致敏阳性组C3片段和VCAM-1表达明显低于TCE致敏阳性组（P<0.05）。Western blot检测结果提示，TCE致敏阳性组小鼠肾小球Claudin-5、Syndecan-1蛋白相对表达水平较溶剂对照组和TCE致敏阴性组明显降低（P<0.05）。与TCE致敏阳性组比较，TCE+CTSLi致敏阳性组小鼠肾脏Syndecan-1蛋白相对表达水平明显升高，差异有统计学意义（P<0.05）；Claudin-5蛋白表达水平升高，但差异无统计学意义（P>0.05）； 结论： CTSL可能通过切割补体C3，激活补体系统，通过破坏内皮细胞结构蛋白Syndecan-1及过表达黏附分子VCAM-1，介导TCE致敏过程中小鼠的肾小球结构和功能损害，抑制CTSL的表达可能是保护小鼠肾小球结构和功能完整性的有效手段。.
Keywords: Cathepsin L; Complement C3; Syndecan-1; Tight junction prolein 5 (Claudin-5); Trichloroethylene; Vascular cell adhesion molecuve1 (VCAM-1).