Effects Of AMPK On Synthesis Of Cholesterol In Cultured Hepatocytes Of Laying Hen

Two experiments were conducted to determine whether AMP-activated protein kinase (AMPK) might involve in regulation of cholesterol metabolism in the liver of laying hen. In the study, cultured hepatocytes of laying hen were used to study the effect of AMPK on the synthesis and transference of cholesterol. Experiment 1 was conducted to study the optimal liver perfusion time to get enough live hepatocytes from laying hen. The hepatocytes were harvested by the way of the simplified two-step collagenase perfusion from the entrance of liver portal vein with syringe for 20min. 30min and 40min. The results showed that with the perfusing time increased from 20min to 40min. the total cells were increased but with the decreased of cell survival rate. The optimal perfusion time was 30min with cell number 1.1 X 109 and surivival rate 88.6%. In experiment 2. hepatocytes of laying hens were cultured without or with 0.5mM 5-amino-imidazolecarboxamide riboside (AICAR) or 0.5mM ~8Br-AMP for 60. and 100min. The results were: There were no effects on ALT. LDH and UA level of the cultured suspension among the treatments. Cultured with 0.5mM AICAR. the hepatocytes AMPK activity was increased by 53.0% (p<0.05) for 60min and 64.5%(p<0.01) for 100min. The HMGR activity was decreased by 82.3% (p<0.01) for 60min and 84.8% (p<0.01) for 100min. The Total Cholesterol (TC) concentration and Total Triglycerides(TG) of the culture were decreased by 44.4% (p<0.05) and 16.7% (p>0.05) respectively for 60min, while TC and TG were decreased by 44.4% (p<0.05) and 37.5%(p>0.05) respectively for 100min. Cultured with 0.5mM ~8Br-AMP. the hepatocytes AMPK activity was decreased by 28.1% (p>0.05) for 60min and 17.1% (p>0.05) for 100min. The HMGR activity was not influenced significantly with decreased by 9.7%(p>0.05) for 60min and 7.5% (p>0.05) for 100min. The TC and TG of the culture were increased by 11.1% (p<0.05) and 83.3% (p<0.01) respectively for 60min. while TC and TG were decreased by 33.3% (p>0.05) and 12.5% (p>0.05) respectively for 100min.that AMPK could regulate the cholesterol synthesis of hepatocytes of laying hen by regulating HMGR.