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The Protective Effect of Grape Procyanidins on the Mice Hepatocytes Injured by Ethanol

Author: LiJing
Tutor: ZhongJinYi
School: Qingdao University
Course: Health Toxicology
Keywords: mice grape procyanidins hepatocytes proliferative activity apoptosis
CLC: R285.5
Type: Master's thesis
Year: 2011
Downloads: 41
Quote: 0
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Abstract


Objective:In this study, we observed the changes of cell proliferation, serum ALT and TNF-a, the abnormal expression of Caspase-3 and Bax protein, pathological changes of liver tissue injured by ethanol. To study the protective effect of grape procyanidins on the liver cell injured by ethanol in vitro and vivoMethod:1 In vitro experiment:The mouse liver homogenate were made into cell suspension. They were randomly divided into 5 groups, low, medium and high dose of GPC groups、ethanol group and control group. Ethanol group and GPC groups were cultured with 100ml/l ethanol, at the same time, low, medium and high dose of GPC groups were given GPC in deferent density:10mg/L、50mg/L、100mg/L,The control group was given the same density PBS, the cells were cultured forl6h in 37℃、5%CO2. Then the proliferation activity of hepatocytes was determined by MTT assay.2 In vivo experiment:The mice were randomly divided into 5 groups, low, medium and high dose of GPC groups、ethanol group and control group.5 mice per group. Fifty,100 and 200mg/kg.bw GPC were orally given to the mice as low, medium and high dose of GPC group. Mice in ethanol and GPC group were orally given at 12ml/kg.bw ethanol. Mice in control group were orally given water,1 time per day. After 4 weeks, all the mice were sacrificed. The proliferation activity of the hepatocytes was determined by MTT assay. The levels of Caspase-3、Bax protein expression in the cell were measured by immunohistochemistry method. The serum levels of TNF-a were detected by ELISA method. The serum alanine aminotransferase (ALT) levels were measured by Automatic biochemical analyzer. The basic pathology change of the liver were detected by HE method.Result:1 In vitro experiment result: From the MTT assay result, The liver cells proliferation activity of the high dose GPC group were 0.412±0.025, higher than the ethanol group 0.304±0.075. The difference was significant (P<0.05).2 In vivo experiment result:(1) Proliferative activity:The liver cells proliferation activity of the high dose and medium dose GPC group were 0.356±0.009 and 0.336±0.020, higher than the ethanol group0.296±0.012. The difference was significant (P<0.05).(2) Apoptosis-related protein:The expression levels of Caspase-3 and Bax protein in high dose group were 19.08% and 14.06%, respectively, and those of the Ethanol-injured group were 51.78% and 58.08%, The differences were significant (P<0.05).(3) TNF-a:The TNF-a levels of the ethanol group were 12.89±1.06 (pg/ml),higher than that of the high and medium dose GPC group (P<0.05). There were no significant difference between control group and high dose GPC group (P>0.05).(4) ALT:The ALT levels of high and medium dose GPC group were 26.00±1.00 U/L、29.00±4.58U/L, lower than that of the ethanol group39.33±3.21 U/L (P<0.05).(5) The GSH content of liver tissue:The GSH levels of the ethanol group were 1.21±0.11, lower than that of normal group (P<0.05). The GSH levels of high and medium dose GPC group were higher than that of ethanol group(P<0.05). There were no significant difference between low dose GPC group and ethanol group (P>0.05).(6) The basic pathology change of the liver:Under light microscopy, the liver histology characteristic of the mice in the high dose GPC group was normal on the whole, but vacuolation, necrosis accompanied with inflammatory infiltration, steatosis were observed in the alcohol group.Conclusion:GPC can inhibit the hepatocyte induced by ethanol.

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