毛稔果提取物对小鼠化学性肝损伤的保护作用及机制研究

Study on protective effects and mechanism of Melastoma sanguineum Sims fruit extract on chemical liver injury in mice

目的研究毛稔果提取物对乙醇、对乙酰氨基酚(APAP)及四氯化碳(CCl_(4))诱导的化学性肝损伤小鼠的肝脏保护作用,探讨其作用机制。方法将96只小鼠按随机数字表法分为正常对照组、乙醇模型组、乙醇+联苯双酯对照组及乙醇+毛稔果提取物高、中、低剂量组,APAP模型组、APAP+联苯双酯对照组及APAP+毛稔果提取物高、中、低剂量组,CCl4模型组、CCl_(4)+联苯双酯对照组及CCl_(4)+毛稔果提取物高、中、低剂量组,每组6只。除正常对照组,其余各组分别制备乙醇模型、APAP模型、CCl4模型。毛稔果提取物高、中、低剂量组小鼠灌胃毛稔果提取物100、50、25 g/kg,联苯双酯对照组腹腔注射15 mg/ml联苯双酯溶液75 mg/kg,正常对照组腹腔注射等体积生理盐水/花生油溶液,1次/d,连续给药25 d。检测小鼠血清中GPT、GOT、TBIL水平;肝组织中SOD、GSH-Px活性及MDA水平;应用qRT-PCR法检测TNF-α、IL-6、乙醇脱氢酶(ADH)、乙醛脱氢酶(ALDH)mRNA表达;Western blot法检测肝组织中细胞色素P450 CYP1A1、CYP1A2、CYP3A蛋白表达;HE染色观察肝组织病理改变。结果与相应的乙醇、APAP及CCl4模型组比较,乙醇+联苯双酯对照组及乙醇+毛稔果提取物高、中剂量组,APAP+联苯双酯对照组及APAP+毛稔果提取物高、中剂量组,CCl_(4)+联苯双酯对照组及CCl_(4)+毛稔果提取物高、中剂量组小鼠血清GPT、GOT、TBIL水平降低(P<0.01或P<0.05),肝组织SOD、GSH-Px活力升高(P<0.01或P<0.05),MDA水平降低(P<0.01或P<0.05),IL-6、TNF-αmRNA水平降低(P<0.01);乙醇+毛稔果提取物高、中、低剂量组ADH、ALDH mRNA水平降低(P<0.01);与APAP模型组比较,APAP+毛稔果提取物组CYP1A1、CYP1A2表达升高(P<0.01),CYP3A蛋白表达降低(P<0.05);与CCl4模型组比较,CCl_(4)+毛稔果提取物组CYP1A1、CYP1A2和CYP3A蛋白表达降低(P<0.05)。结论毛稔果提取物对乙醇、APAP及CCl4所致的小鼠化学性肝损伤具有保护作用,其机制可能与抗氧化应激、抑制炎症反应及调节细胞色素P450相关酶系表达有关。

Objective To study the protective effects and mechanism of Melastoma sanguineum Sims fruit extract(MSE)on chronic chemical liver injury induced by ethanol,acetaminophen and carbon tetrachloride in mice;To discuss it mechanism.Methods Totally 96 mice were divided into normal control group,ethanol model group,ethanol+bifendate control group and ethanol+MSE high-,medium-and low-dosage groups,APAP model group,APAP+bifendate control group and APAP+MSE high-,medium-and low-dosage groups,CCl4 model group,CCl_(4)+bifendate control group and CCl_(4)+MSE high-,medium-and low-dosage groups,with 6 mice in each group.Except for the normal control group,the other groups were respectively prepared for the ethanol model,the APAP model and the CCl4 model.The mice in the MSE high-,medium-and low-dosage groups were intragastrically administrated with 10,5 and 2.5 g/kg of MSE,respectively;the bifendate control group was intraperitoneally injected with 15 mg/ml bifendate solution at 75 mg/kg;the normal control group was intraperitoneally injected with equal volume of normal saline/peanut oil solution once a day for 25 consecutive days.The levels of GPT,GOT and total bilirubin(TBIL)in serum were detected;the activities of SOD and GSH-Px and the content of MDA in liver tissue were detected;the mRNA expressions of TNF-α,IL-6,alcohol dehydrogenase(ADH)and acetaldehyde dehydrogenase(ALDH)were detected by qRT-PCR;the protein expressions of cytochrome P450 CYP1A1,CYP1A2,and CYP3A in liver tissue were detected by Western blot;the pathological changes of the liver tissue were observed by HE staining.Results Compared with the corresponding ethanol,APAP and CCl4 model groups,the serum GPT,GOT and TBIL levels of mice in the ethanol+bifendate control group and ethanol+MSE high-and medium-dosage groups,the APAP+bifendate control group and APAP+MSE high-and medium-dosage groups,and the CCl_(4)+bifendate control group and CCl_(4)+MSE high-and medium-dosage groups decreased(P<0.01 or P<0.05),the activities of SOD and GSH-Px in the liver tissue increased(P<0.01 or P<0.05),and the MDA level decreased(P<0.01 or P<0.05),and the mRNA levels of IL-6 and TNF-αdecreased(P<0.01);the mRNA levels of ADH and ALDH in the ethanol+MSE high-,medium-,and low-dosage groups decreased(P<0.01).Compared with the APAP model group,the expressions of CYP1A1 and CYP1A2 in the APAP+MSE groups increased(P<0.01),and the expression of CYP3A protein decreased(P<0.05);compared with the CCl4 model group,the expressions of CYP1A1,CYP1A2 and CYP3A proteins in the CCl_(4)+MSE groups decreased(P<0.05).Conclusion MSE has a protective effect on chronic chemically-induced liver injury induced by ethanol,APAP,and CCl4 in mice,and its mechanism may be related to antioxidant stress,inhibition of inflammatory response,and regulation of the expression of cytochrome P450-related enzymes.