茵杞调脂饮对非酒精性脂肪肝大鼠脂代谢影响的实验研究
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摘要
目的研究茵杞调脂饮对非酒精性脂肪肝大鼠脂质代谢的影响及作用机制。方法 48只SD大鼠随机分为正常组、模型组、辛伐他汀组(2.08 mg/kg),以及中药低(8.28 g/kg)、中(16.56 g/kg)、高(33.12 g/kg)剂量组。除正常组外,其余5组采用高脂饲料饲养12周建立非酒精性脂肪肝大鼠模型。造模成功后,各药物组灌胃给予相应药物,正常组、模型组灌胃给予等量0.9%NaCl溶液。连续干预8周,称取大鼠体质量、肝湿重,计算肝指数,常规检测血清丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转氨酶(AST)、胆固醇(TC)、三酰甘油(TG)水平,ELISA法检测肝组织TC、TG、丙二醛(MDA)、超氧化物歧化酶(SOD)、游离脂肪酸(FFA)水平,苏木素-伊红(HE)染色、油红O染色观察肝组织病理学情况,PCR检测肝组织肝X受体α(LXRα)、固醇调节元件结合蛋白-1c(SREBP-1c)相对表达量。结果①与正常组比较,模型组大鼠体质量、肝湿重、肝指数增加(P<0.05),血清ALT、AST、TC、TG水平升高(P<0.05),肝组织TC、TG、MDA、FFA水平及LXRα、SREBP-1c mRNA相对表达量升高(P<0.05),SOD水平降低(P<0.05)。②与模型组比较,辛伐他汀组、中药各剂量组大鼠体质量、肝湿重、肝指数减少(P<0.05),血清ALT、AST、TC、TG水平降低(P<0.05),肝组织TC、TG、FFA、MDA水平及LXRα、SREBP-1c mRNA相对表达量降低(P<0.05),SOD水平升高(P<0.05)。③与辛伐他汀组比较,中药中、高剂量组大鼠血清ALT、AST水平降低(P<0.05),中药高剂量组大鼠体质量、肝湿重、肝指数,肝组织FFA、MDA水平及LXRαmRNA、SREBP-1c mRNA相对表达量降低(P<0.05),SOD水平升高(P<0.05)。④中药各剂量组组间比较,高剂量组大鼠体质量、肝湿重、肝指数水平较低剂量组明显下降(P<0.05);中剂量组较低剂量组血清ALT、AST水平下降(P<0.05),高剂量组较低剂量ALT、AST、TC、TG水平下降(P<0.05);中药各剂量组组间比较,肝组织TC、TG(中剂量组较低剂量组)及FFA、MDA、SOD(高剂量组较低、中剂量组)水平随剂量的增加明显降低;中药高剂量组大鼠肝组织LXRαmRNA、SREBP-1c mRNA相对表达量较中药低剂量组减少(P<0.05)。结论茵杞调脂饮可明显降低非酒精性脂肪肝大鼠的血脂和肝脂水平,抑制脂质蓄积,减轻氧化应激,保护肝功能,改善肝脏病理损伤;其机制可能与抑制LXRα、SREBP-1c的表达有关。
Objective To investigate the effects of Yinqi Tiaozhi Decoction on lipid metabolism in rats with nonalcoholic fatty liver disease(NAFLD) and the mechanism of action. Methods 48 SD rats were randomly assigned into normal group, model group, simvastatin(2.08 mg/kg) group, low-dose(8.28 g/kg), middle-dose(16.56 g/kg) and high-dose(33.12 g/kg) Yinqi Tiaozhi Decoction groups. Except the normal group, high-fat diet was given to rats in the other 5 groups for 12 weeks to establish the NAFLD rat model. After successful modeling,corresponding drugs were administered by gavage to the drug groups,and 0. 9% Na Cl solution of equal volume was given by gavage to the normal and model groups. At 8 weeks of treatment,wet weight of the liver and liver index were calculated,the levels of alanine aminotransferase( ALT),aspartate aminotransferase( AST),cholesterol( TC) and triglyceride( TG) in serum were detected; the levels of TC,TG,malondialdehyde( MDA),superoxide dismutase( SOD) and free fatty acid( FFA) in liver tissue were determined by ELISA; hematoxylin-eosin( HE) staining and oil red O staining were performed to observe the pathology of liver tissue;and the relative expression of liver X receptor α( LXRα) and sterol regulatory element-binding protein-1 c( SREBP-1 c) in liver tissue was detected by polymerase chain reaction( PCR). Results(1) Compared to the normal group,the body mass,wet weight of the liver and liver index increased( P<0.05); the levels of ALT,AST,TC and TG in serum increased( P<0.05),the levels of TC,TG,MDA and FFA,the relative expression of LXRαand SREBP-1 c mRNA in liver tissue increased( P<0.05) and the SOD level decreased( P<0.05) in the model group.(2)Compared to the model group,the body mass,wet weight of the liver and liver index decreased( P<0.05); the levels of ALT,AST,TC and TG in serum decreased( P<0.05),the levels of TC,TG,MDA and FFA and the relative expression of LXRα and SREBP-1 c mRNA in liver tissue decreased( P<0.05) and the SOD level increased( P<0.05) in the simvastatin group and Yinqi Tiaozhi Decoction groups.(3)Compared to the simvastatin group,the levels of ALT and AST in serum decreased in the middle-dose and high-dose Yinqi Tiaozhi Decoction groups( P<0.05),the body mass,wet weight of the liver and liver index decreased,the levels of FFA and MDA and the relative expression of LXRα mRNA and SREBP-1 c mRNA decreased( P<0.05),and the SOD level increased( P<0.05) in the high-dose Yinqi Tiaozhi Decoction group.(4)The body mass,wet weight of the liver and liver index decreased significantly in the high-dose group compared to the low-dose group( P<0.05); the levels of ALT and AST in serum decreased in the middle-dose group compared to the low-dose group( P<0.05),and the levels of ALT,AST,TC and TG decreased in the high-dose group compared to the low-dose group( P<0.05).The levels of TC and TG( middle-dose group vs. low-dose group) and of FFA,MDA and SOD( high-dose group vs. low-and middle-dose groups)decreased significantly with increase in dose; and the relative expression of LXRα mRNA and SREBP-1 c mRNA in liver tissue decreased in the highdose group compared to the low-dose group( P<0.05). Conclusion Yinqi Tiaozhi Decoction reduces significantly the levels of blood and liver lipid in in rats with NAFLD,inhibits lipid accumulation,reduces oxidative stress,protects liver function,and improves pathological injury of the liver. The mechanism of action may be related with inhibition of the expression of LXRα and SREBP-1 c.
Objective To investigate the effects of Yinqi Tiaozhi Decoction on lipid metabolism in rats with nonalcoholic fatty liver disease(NAFLD) and the mechanism of action. Methods 48 SD rats were randomly assigned into normal group, model group, simvastatin(2.08 mg/kg) group, low-dose(8.28 g/kg), middle-dose(16.56 g/kg) and high-dose(33.12 g/kg) Yinqi Tiaozhi Decoction groups. Except the normal group, high-fat diet was given to rats in the other 5 groups for 12 weeks to establish the NAFLD rat model. After successful modeling,corresponding drugs were administered by gavage to the drug groups,and 0. 9% Na Cl solution of equal volume was given by gavage to the normal and model groups. At 8 weeks of treatment,wet weight of the liver and liver index were calculated,the levels of alanine aminotransferase( ALT),aspartate aminotransferase( AST),cholesterol( TC) and triglyceride( TG) in serum were detected; the levels of TC,TG,malondialdehyde( MDA),superoxide dismutase( SOD) and free fatty acid( FFA) in liver tissue were determined by ELISA; hematoxylin-eosin( HE) staining and oil red O staining were performed to observe the pathology of liver tissue;and the relative expression of liver X receptor α( LXRα) and sterol regulatory element-binding protein-1 c( SREBP-1 c) in liver tissue was detected by polymerase chain reaction( PCR). Results(1) Compared to the normal group,the body mass,wet weight of the liver and liver index increased( P<0.05); the levels of ALT,AST,TC and TG in serum increased( P<0.05),the levels of TC,TG,MDA and FFA,the relative expression of LXRαand SREBP-1 c mRNA in liver tissue increased( P<0.05) and the SOD level decreased( P<0.05) in the model group.(2)Compared to the model group,the body mass,wet weight of the liver and liver index decreased( P<0.05); the levels of ALT,AST,TC and TG in serum decreased( P<0.05),the levels of TC,TG,MDA and FFA and the relative expression of LXRα and SREBP-1 c mRNA in liver tissue decreased( P<0.05) and the SOD level increased( P<0.05) in the simvastatin group and Yinqi Tiaozhi Decoction groups.(3)Compared to the simvastatin group,the levels of ALT and AST in serum decreased in the middle-dose and high-dose Yinqi Tiaozhi Decoction groups( P<0.05),the body mass,wet weight of the liver and liver index decreased,the levels of FFA and MDA and the relative expression of LXRα mRNA and SREBP-1 c mRNA decreased( P<0.05),and the SOD level increased( P<0.05) in the high-dose Yinqi Tiaozhi Decoction group.(4)The body mass,wet weight of the liver and liver index decreased significantly in the high-dose group compared to the low-dose group( P<0.05); the levels of ALT and AST in serum decreased in the middle-dose group compared to the low-dose group( P<0.05),and the levels of ALT,AST,TC and TG decreased in the high-dose group compared to the low-dose group( P<0.05).The levels of TC and TG( middle-dose group vs. low-dose group) and of FFA,MDA and SOD( high-dose group vs. low-and middle-dose groups)decreased significantly with increase in dose; and the relative expression of LXRα mRNA and SREBP-1 c mRNA in liver tissue decreased in the highdose group compared to the low-dose group( P<0.05). Conclusion Yinqi Tiaozhi Decoction reduces significantly the levels of blood and liver lipid in in rats with NAFLD,inhibits lipid accumulation,reduces oxidative stress,protects liver function,and improves pathological injury of the liver. The mechanism of action may be related with inhibition of the expression of LXRα and SREBP-1 c.
引文
[1]ARA U'JO A R,ROSSO N,BEDOGNI G,et al.Global epidemiology of non-alcoholic fatty liver disease/non-alcoholic steatohepatitis:What we need in the future[J].Liver Int,2018,38(Supple1):47-51.
[2]李生鹏,王全楚.非酒精性脂肪性肝病的流行病学进展[J].胃肠病学和肝病学杂志,2017,26(10):1085-1087.
[3]KWAK M S,KIM D.Non-alcoholic fatty liver disease and lifestyle modifications,focusing on physical activity[J].Korean J Intern Med,2018,33(1):64-74.
[4]王鹏,严妍,于文君,等.他汀类药物不良反应的研究进展[J].中国医院药学杂志,2016,36(22):2035-2039.
[5]吕宝伟,冯春青,孙建光.补肾降浊饮对非酒精性脂肪肝大鼠胰岛素抵抗的影响[J].中国实验方剂学杂志,2018,24(12):107-113.
[6]丁婧,王辉,余诗灏,等.肥胖大鼠模型的建立及其脂代谢相关分子机制研究[J].中国实验动物学报,2012,20(5):20-24.
[7]闫蓉,牛春燕,于璐,等.非酒精性脂肪性肝病细胞模型中自噬与脂质代谢的相互调节[J].临床肝胆病杂志,2017,33(10):1981-1986.
[8]YU D,CHEN G,PAN M,et al.High fat diet-induced oxidative stress blocks hepatocyte nuclear factor 4αand leads to hepatic steatosis in mice[J].J Cell Physiol,2018,233(6):4770-4782.
[9]HOMMA T,KURAHASHI T,LEE J,et al.Double knockout of peroxiredoxin 4(Prdx4)and superoxide dismutase 1(Sod1)in mice results in severe liver failure[J].Oxid Med Cell Longev,2018(6):1-12.
[10]RONG S,CORTéS V A,RASHID S,et al.Expression of SREBP-1c requires SREBP-2-mediated generation of a sterol ligand for LXR in livers of mice[J].eLife,2017(6):e25015.
[11]CALKIN A C,TONTONOZ P.Liver x receptor signaling pathways and atherosclerosis[J].Arterioscler Thromb Vasc Biol,2010,30(8):1513-1518.
[12]孙建光.茵杞消脂饮治疗脂肪肝30例临床观察[J].中医临床研究,2011,3(7):9-11.
[13]吕宝伟,冯春青,孙建光.基于“肝主疏泄”理论探讨脂肪肝分期治疗[J].吉林中医药,2018,38(3):266-269.
[14]LIM D W,KIM Y T,JANG Y J,et al.Anti-obesity effect of Artemisia capillaris extracts in high-fat diet-induced obese rats[J].Molecules,2013,18(8):9241-9252.
[15]王永辉,高丽,周文静,等.决明子乙醇提取物对高脂血症模型大鼠糖脂代谢及相关炎性细胞因子的影响[J].中国实验方剂学杂志,2014,20(7):178-181.
[16]冯孔龙,朱晓艾,陈彤,等.川陈皮素对高脂膳食诱导大鼠的降脂减肥及预防脂肪肝形成作用[J].食品科学,2018,39(1):213-220.
[17]高丽,周文静,马艳苗,等.决明子乙醇提取物对高脂血症模型大鼠瘦素及神经肽Y的影响[J].中国实验方剂学杂志,2013,19(8):235-238.
[18]夏建萍,应豪,胡爱荣,等.非酒精性脂肪性肝病肝脏脂肪转运环节的变化及白术多糖的干预作用[J].中国卫生检验杂志,2017,27(4):461-463.
[19]胡慧明,邵峰,朱彦陈,等.南山楂提取物对高脂血症模型小鼠胆固醇生物合成的影响[J].中药材,2015,38(5):1034-1036.
[20]王鹏,高敏艳,高岚,等.山楂提取物对脂肪酸合酶的体外抑制作用[J].天津中医药大学学报,2012,31(4):229-232.
[21]余轶群,陈希,李军祥.绞股蓝、生山楂水提物对非酒精性脂肪性肝炎大鼠脂质代谢相关因子的调控作用研究[J].辽宁中医药大学学报,2012,14(7):168-170.
[22]杨华,张知贵,李小慧.山楂叶总黄酮对高脂血症大鼠血脂和血液流变性的影响[J].中国实验方剂学杂志,2012,18(12):257-259.
[23]刘新迎,周联,梁瑞燕,等.通过对3T3-L1细胞的作用探讨山楂叶总黄酮调脂机制[J].中华中医药学刊,2009,27(5):1066-1068.
[24]李金梅,丁丽丽,宋保亮,等.大黄酚对Huh-7细胞SREBP表达及脂质代谢的影响[J].药学学报,2015,50(2):174-179.
[25]岑柏春,张谈,袁建芬,等.大黄酸对高脂饮食诱导的大鼠非酒精性脂肪性肝病的防治作用[J].中华中医药学刊,2013,31(3):545-547.
[26]何百川,薛超,韩一益,等.佩兰对2型糖尿病合并脂代谢紊乱大鼠肝脏DGAT2表达的影响[J].辽宁中医杂志,2017,44(3):607-610.
[27]卜晓芬,李骏,朱虹.甘草甜素对体外诱导的酒精性脂肪肝细胞的影响及机制研究[J].重庆医学,2018,47(4):436-438.
[2]李生鹏,王全楚.非酒精性脂肪性肝病的流行病学进展[J].胃肠病学和肝病学杂志,2017,26(10):1085-1087.
[3]KWAK M S,KIM D.Non-alcoholic fatty liver disease and lifestyle modifications,focusing on physical activity[J].Korean J Intern Med,2018,33(1):64-74.
[4]王鹏,严妍,于文君,等.他汀类药物不良反应的研究进展[J].中国医院药学杂志,2016,36(22):2035-2039.
[5]吕宝伟,冯春青,孙建光.补肾降浊饮对非酒精性脂肪肝大鼠胰岛素抵抗的影响[J].中国实验方剂学杂志,2018,24(12):107-113.
[6]丁婧,王辉,余诗灏,等.肥胖大鼠模型的建立及其脂代谢相关分子机制研究[J].中国实验动物学报,2012,20(5):20-24.
[7]闫蓉,牛春燕,于璐,等.非酒精性脂肪性肝病细胞模型中自噬与脂质代谢的相互调节[J].临床肝胆病杂志,2017,33(10):1981-1986.
[8]YU D,CHEN G,PAN M,et al.High fat diet-induced oxidative stress blocks hepatocyte nuclear factor 4αand leads to hepatic steatosis in mice[J].J Cell Physiol,2018,233(6):4770-4782.
[9]HOMMA T,KURAHASHI T,LEE J,et al.Double knockout of peroxiredoxin 4(Prdx4)and superoxide dismutase 1(Sod1)in mice results in severe liver failure[J].Oxid Med Cell Longev,2018(6):1-12.
[10]RONG S,CORTéS V A,RASHID S,et al.Expression of SREBP-1c requires SREBP-2-mediated generation of a sterol ligand for LXR in livers of mice[J].eLife,2017(6):e25015.
[11]CALKIN A C,TONTONOZ P.Liver x receptor signaling pathways and atherosclerosis[J].Arterioscler Thromb Vasc Biol,2010,30(8):1513-1518.
[12]孙建光.茵杞消脂饮治疗脂肪肝30例临床观察[J].中医临床研究,2011,3(7):9-11.
[13]吕宝伟,冯春青,孙建光.基于“肝主疏泄”理论探讨脂肪肝分期治疗[J].吉林中医药,2018,38(3):266-269.
[14]LIM D W,KIM Y T,JANG Y J,et al.Anti-obesity effect of Artemisia capillaris extracts in high-fat diet-induced obese rats[J].Molecules,2013,18(8):9241-9252.
[15]王永辉,高丽,周文静,等.决明子乙醇提取物对高脂血症模型大鼠糖脂代谢及相关炎性细胞因子的影响[J].中国实验方剂学杂志,2014,20(7):178-181.
[16]冯孔龙,朱晓艾,陈彤,等.川陈皮素对高脂膳食诱导大鼠的降脂减肥及预防脂肪肝形成作用[J].食品科学,2018,39(1):213-220.
[17]高丽,周文静,马艳苗,等.决明子乙醇提取物对高脂血症模型大鼠瘦素及神经肽Y的影响[J].中国实验方剂学杂志,2013,19(8):235-238.
[18]夏建萍,应豪,胡爱荣,等.非酒精性脂肪性肝病肝脏脂肪转运环节的变化及白术多糖的干预作用[J].中国卫生检验杂志,2017,27(4):461-463.
[19]胡慧明,邵峰,朱彦陈,等.南山楂提取物对高脂血症模型小鼠胆固醇生物合成的影响[J].中药材,2015,38(5):1034-1036.
[20]王鹏,高敏艳,高岚,等.山楂提取物对脂肪酸合酶的体外抑制作用[J].天津中医药大学学报,2012,31(4):229-232.
[21]余轶群,陈希,李军祥.绞股蓝、生山楂水提物对非酒精性脂肪性肝炎大鼠脂质代谢相关因子的调控作用研究[J].辽宁中医药大学学报,2012,14(7):168-170.
[22]杨华,张知贵,李小慧.山楂叶总黄酮对高脂血症大鼠血脂和血液流变性的影响[J].中国实验方剂学杂志,2012,18(12):257-259.
[23]刘新迎,周联,梁瑞燕,等.通过对3T3-L1细胞的作用探讨山楂叶总黄酮调脂机制[J].中华中医药学刊,2009,27(5):1066-1068.
[24]李金梅,丁丽丽,宋保亮,等.大黄酚对Huh-7细胞SREBP表达及脂质代谢的影响[J].药学学报,2015,50(2):174-179.
[25]岑柏春,张谈,袁建芬,等.大黄酸对高脂饮食诱导的大鼠非酒精性脂肪性肝病的防治作用[J].中华中医药学刊,2013,31(3):545-547.
[26]何百川,薛超,韩一益,等.佩兰对2型糖尿病合并脂代谢紊乱大鼠肝脏DGAT2表达的影响[J].辽宁中医杂志,2017,44(3):607-610.
[27]卜晓芬,李骏,朱虹.甘草甜素对体外诱导的酒精性脂肪肝细胞的影响及机制研究[J].重庆医学,2018,47(4):436-438.