水飞蓟宾治疗非酒精性脂肪性肝病的实验研究

Hepatoprotective effect of silybin on rats with non-alcoholic fatty liver disease

目的:观察水飞蓟宾干预对非酒精性脂肪性肝病(NAFLD)大鼠肝脏组织过氧化物酶体增殖物激活受体(PPARs)基因及其下游基因表达、线粒体膜流动性的影响,探讨水飞蓟宾防治NAFLD的可能机制。方法:采用高脂饮食建立NAFLD大鼠模型,分为模型组、罗格列酮组、水飞蓟宾组,并以正常大鼠为空白对照组。6周后用荧光偏振法检测肝脏线粒体膜流动性的变化,RT-PCR法检测大鼠血清PPAR-α、PPAR-γ及其下游基因包括长链酰基辅酶A脱氢酶(LCAD)、脂酰辅酶A氧化酶(AOX)和细胞色素P450A1酶(CYP450A1)、脂联素、脂联素受体(Ad ipoR)、抵抗素mRNA水平的表达,观察各组上述指标的差异。结果:模型组大鼠肝组织可见弥漫性肝细胞大泡或小泡性的脂肪变性,部分伴肝细胞坏死和炎性细胞浸润,罗格列酮组与水飞蓟宾组肝组织可见少量小泡性脂肪变性,未见明显肝细胞坏死和炎性细胞浸润。模型组大鼠肝细胞线粒体微黏度明显高于空白对照组(P<0.01);罗格列酮组及水飞蓟宾组大鼠肝细胞线粒体微黏度低于模型组(P<0.05或0.01),水飞蓟宾组明显低于罗格列酮组(P<0.05)。与空白对照组比较,模型组PPARα-、AOX、CYP450A1、PPARγ-、脂联素、Ad ipoR mRNA表达水平明显下降,抵抗素表达水平升高。罗格列酮组PPARγ-、PPAR-α、脂联素mRNA表达水平明显高于模型组(P均<0.01或0.05);水飞蓟宾组PPAR-α、AOX、CYP450A1,PPARγ-、脂联素mRNA表达水平均比模型组上升(P<0.05),且抵抗素mRNA表达水平比模型组下降(P<0.05)。结论:水飞蓟宾可以有效防治NAFLD,其机制可能与稳定并维持适当的肝脏线粒体膜流动性,减轻肝脏脂质过氧化以及改善胰岛素抵抗有关。

Objective： To investigate the expressions of peroxisome proliferator-activated receptors （PPARs） and their downstream genes as well as the change of mitochondria membrane fluidity in rats with non-alcoholic fatty liver disease （NAFLD） , and to explore the probable mechanism of hepatoprotective effect of silybin. Methods： Rats were fed with high fat diet for 6 weeks to establish NAFLD models. Mitochondria membrane fluidity was detec- ted by fluorescence polarization immunoassay （ FPIA ） The expression of PPAR-α, PPAR-γ, LCAD, AOX, CYP450A1, adiponectin, adiponectin releptor （adipoR） and resistin genes were measured by reverse transcription-polymerase chain reaction （RT-PCR）. Results： After consumption of high -fat diet for 6 weeks, the rat liver in model group showed typical microvesicle and macrovesicle steatosis accompanied with mild to moderate lobular inflammatory cell infiltration and hepatocyte necrosis. After intervened by rosiglitazone and silybin, however, the liver showed fewer microvesicle steatosis, without significant necrosis and inflammatory cell infiltration. Microviscosity of mitochondrial membrane was remarkably higher in model group than in control group （ P 〈 0. 01 ）, and showed a distinct fall in the two pharmaceutical intervention groups （P 〈0.05 or 0. 01 ）, with a more notable decrease in silybin group as compared with rosiglitazone group （ P 〈 0.05 ）. Inhibition of the expressions of PPAR-ot, AOX, CYP450A1, PPAR-γ, adiponectin and adipoR and enhancement of the expression of resistin could be seen in model group when compared with control group. The mRNA expression of PPAR-γ in rosiglitazone group was re- markably increased as compared with model group （ P 〈 0. 01 ）. The expressions of PPAR-α and adiponectin showed similar change tendency in rosiglitazone group when compared with model group （ P 〈 0. 05 ）. Compared with model group, the mRNA expressions of PPAR-α, AOX, CYP450A1, PPAR-γ and adiponectin were increased while the expression of resistin was decreased in silybin group （P 〈 0. 05）. Conclusion： Mitochondria membrane stabilization, oxidative stress inhibition as well as insulin resistance improvement may be the three probable mecha- nisms for hepatoprotective effect of silybin in the NAFLD.