二甲双胍对镉诱导小鼠急性肝毒性的影响

doi:10.3969/j.issn.1004-616x.2023.01.001

摘要/Abstract

摘要： 目的: 探讨二甲双胍是否可以缓解镉所诱导的小鼠肝毒性。方法: 将雄性野生型(WT)小鼠和肝脏特异性敲除编码PP2AAα的PPP2R1A基因(HO)小鼠随机各分为4组，包括对照组(0.9%生理盐水)、镉染毒组(1.5 mg/kg CdCl₂)、二甲双胍处理组(100 mg/kg二甲双胍)、镉和二甲双胍联用组(1.5 mg/kg CdCl₂，100 mg/kg二甲双胍)。各组小鼠连续7 d 分别腹腔注射上述受试物，采用半自动血生化分析仪检测小鼠外周血谷丙转氨酶(ALT)、谷草转氨酶(AST)和甘油三酯(TG)浓度；HE及油红O染色观察肝脏组织病理学改变；实时荧光定量PCR(qPCR)检测肝组织中镉相关转运蛋白zip14、dmt1、mrp1、oct2、bsep和zip8的mRNA表达水平。L02人肝细胞分别用CdCl₂染毒和/或二甲双胍处理后，CCK-8试剂盒法检测细胞活力，荧光显微镜下检测Fluo-4 AM的荧光强度评估细胞内钙离子水平。结果: 镉染毒时，WT小鼠血浆ALT和AST浓度分别升高了86.40%和3.19%，出现肝细胞损伤、肝脏脂质变性与沉积等病理改变，HO小鼠的肝脏损伤比WT小鼠更加明显。与镉染毒组相比，镉和二甲双胍联用组小鼠的血浆ALT与AST浓度分别降低了 61.14%和 44.02%，肝细胞坏死、脂质变性与沉积的程度亦有所减轻。未染毒时，HO 小鼠肝脏中的 zip14 和 dmt1mRNA表达水平较WT小鼠分别升高16.48%和53.61%；镉染毒后，zip14和dmt1 mRNA表达水平分别升高47.32%和136.23%。镉染毒L02细胞的荧光强度增强了2.72倍，相应细胞活力降低45.99%。和单纯镉染毒细胞比，1和2 mmol/L二甲双胍处理导致细胞荧光强度分别降低了40.77%和54.17%，细胞活力则升高了59.28%和89.78%。结论: PP2A在CdCl₂诱导小鼠肝毒性中起作用，可能通过调控镉转运蛋白的表达来影响镉的肝毒性。二甲双胍可缓解镉所诱导的肝脏损伤，其可能机制是抑制镉离子进入肝细胞。

关键词: 氯化镉, 肝毒性, 蛋白磷酸酶2A, 二甲双胍

Abstract: OBJECTIVE: To investigate effects of metformin on alleviating cadmium-induced liver injury in mice. METHODS: Male wild-type (WT) and liver-specific Ppp2r1a knockout mice (HO) were randomly divided into 4 groups： control (solvent control)， cadmium treatment (1.5 mg/kg CdCl₂)， metformin treatment (100 mg/kg metformin)， and cadmium and metformin combined treatment (1.5 mg/kg CdCl₂， 100 mg/kg metformin). These mice were consecutively administrated for 7 days by intraperitoneal injection. The semiautomatic blood biochemical analyzer was used to examine levels of ALT，AST，and TG. H&E and oil red O staining were performed to observe pathological changes of liver tissues. Real-time quantitative fluorescence PCR (qPCR) was conducted to determine mRNA levels of cadmium-related transporters. After L02 human hepatocytes were exposed to CdCl₂ and/or metformin， cell viability was detected by CCK-8 kit， and intracellular calcium ion levels were evaluated by detecting the fluorescence intensity of Fluo-4 AM under a fluorescence microscope. RESULTS: Cadmium exposure increased the blood plasma ALT and AST levels in WT mice by 86.40% and 3.19%，respectively. These mice exhibited pathological changes such as hepatocyte injury，liver lipid degeneration and deposition，and liver injury. Moreover，these effects were more profound in HO mice than that in WT mice. Compared with the cadmium treatment group alone，metformin treatment led to the 61.14% and 44.02% decrease in the plasma ALT and AST levels，respectively. Correspondingly，the degree of hepatocyte necrosis，lipid degeneration and deposition were relieved. Compared with WT mice，the mRNA levels of zip14 and dmt1， which mainly mediate cadmium transport and absorption in liver， were increased in HO mice by 16.48% and 53.61%，respectively. Upon CdCl₂ exposure，the mRNA levels of zip14 and dmt1 were increased in HO mice by 47.32% and 136.23%，respectively，compared to that in WT mice. The fluorescence intensity in L02 cells treated with CdCl₂ was increased by 2.72 times，and the corresponding cell viability was decreased by 45.99%. Compared to that in CdCl₂-treatemnt alone，1 mmol/L and 2 mmol/L metformin treatment reduced the fluorescence intensity by 40.77% and 54.17% ， respectively， and the corresponding cell viability were increased by 59.28% and 89.78%. CONCLUSION: PP2A activities were involved in CdCl₂-induced liver injury in mice by regulating the expression of cadmium transporter. Metformin could alleviate CdCl₂-induced hepatotoxicity，partly by inhibiting absorption of cadmium ions in hepatocytes.

Key words: CdCl₂, hepatotoxicity, protein phosphatase 2A, metformin

中图分类号:

R114

谭明雪, 柳晓玲, 姜姝芸, 潘心红, 陈雯, 陈丽萍. 二甲双胍对镉诱导小鼠急性肝毒性的影响[J]. 癌变·畸变·突变, 2023, 35(1): 1-8.

TAN Mingxue, LIU Xiaoling, JIANG Shuyun, PAN Xinhong, CHEN Wen, CHEN Liping. EEffects of metformin on cadmium-induced acute hepatotoxicity in mice[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2023, 35(1): 1-8.

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