Regulation Of ApoM Expression And Secretion By Leptin In Vitro And In Vivo

Objective In the present study we further examined effects of leptin on apoM，apoAI，apoB and LDL-R expression in normal rats.Methods Healthy rats were infused with leptin (50ng/ml) through tail veins for6hours continuously, at the speed of2.5ml per hour by microinfusion pumps. The controlswere infused with saline as the same way. Serum apoM protein concentrations weredetermined by the dot-blotting method and hepatic apoM mRNA levels were determinedby real-time RT-PCR. The serum glucose, insulin, HDL, LDL, cholesterol and triglyceridewere measured by standard chemistry methods at the same time.Results After short time leptin administration, serum insulin levels were significantlydecreased by68.82%（P=0.0317）, at the meanwhile blood glucose levels were increased by14.68%（P=0.0079）in the leptin treated rats compared to the control rats. Moreover, leptincould significantly decrease hepatic mRNA levels of apoM, apoB and LDL-R by55.15%（P=0.0159）、48.60%（P=0.0079）and40.43%（P=0.0159）, respectively. whereas apoAImRNA level was increased by149%（P=0.0079）in the leptin treated rats. There were nosignificant change in the levels of HDL, LDL, cholesterol and triglyceride among groups.Conclusion Regulation of leptin on apoM expression may be associated with lowlevels of insulin and high blood glucose levels. Leptin could significantly promote theexpression of apoAI, but inhibited the expression of apoB and LDL. Objective In this part of works we further examined effects of leptin on apoM， expression in isolated rat’s liver perfusion system and in the human liver cell line, LO2cellsMethods Isolated rat’s liver were perfused with RPMI1640, RPMI1640containing10ng/ml leptin or100ng/ml leptin, respectively. After6hrs, mRNA levels of apoM, apoAI,apoB and LDL-R were detected by RT-PCR. The effect of leptin on apoM mRNAexpression in he human liver cell line, LO2, is measured by real-time PCR，after24hoursof treatment with various concentrations of leptins.Results In isolated liver perfusion model, physiological (10ng/ml) andsupraphysiological (100ng/ml) concentrations of leptin groups could increase expression ofapoM, apoB and LDLR, respectively, whereas leptin didn’t affect apoAI expression. InLO2cell line, supraphysiological concentrations of leptin could decrease apoM expressionsignificantly.Conclusion Comparing with the normal environment in vivo, the isolated liverperfusion model is lack of overall adjustment mechanism. The feedback regulationbetween leptin and insulin were blocked. This may involve in the different regulativeeffects of leptin in the liver perfusion model and in vivo. Leptin could also effect theexpression apoAI, suggesting leptin maybe involve in the regulation of cholesterolmetabolism. Objective This part of works is aimed to clarify the possible regulatory mechanism ofleptin on apoM expression in isolated rat’s liver perfusion system and LO2cells。Methods Isolated liver perfusion models are divided into four groups (each group had8perfued livers)： group1, culture medium (containing bovine serum albumin30g/L)+0ng/ml leptin; group2， culture medium (containing bovine serum albumin30g/L)+0ng/ml leptin+10μM LY294002; group3，culture medium (containing bovine serumalbumin30g/L)+100ng/ml leptin; and group4，culture medium (containing bovine serumalbumin30g/L)+100ng/ml leptin+10μM LY294002. All perfusate were pumped at5ml/min speed continued for6hrs. Then small piece of liver tissue were isolated andmRNA level were detected by RT-PCR. In another series experiments human liver cell line LO2were incubated with different incubates: group1, RPMI1640culture medium+500ng/ml leptin; group2, RPMI1640culture medium+500ng/ml leptin+1μM LY294002;group3, RPMI1640culture medium+500ng/ml leptin+5μM LY294002; group4,RPMI1640culture medium+500ng/ml leptin+10μM LY294002. After24hrs, apoMexpression level were detected by RT-PCR.Results After24hrs incubation with leptin and LY294002in LO2cell line, apoMmRNA expression level has no significant difference compared to the controls. Afterpriming with LY2940026hrs in isolated liver perfusion models, apoM expression levelsalso has no significantly change compared with the control group. By two-factor analysis,leptin and LY294002were no interaction.Conclusion In vivo, the regulation of leptin on apoM may not be the effect of a singleorganization or a single signal pathway. Leptin regulated apoM expression is not directlythrough the PI3K pathway.