Effects Of Panax Notoginseng Saponins On Expression Of UCP2 MRNA In Steatotic Hepatocytes LO2

Objective:To investgate the effects of Panax Notoginseng Saponins(PNS) on expression of UCP2 mRNA in steatotic hepatocytes L02,so as to explore its Lipid metabolism and Molecular mechanism.Methods:①The establishment of model:To culture the hepatocytes with 50%calf serum after 24 hours,48 hours and 72 hours respectively,and establish a model of steatosis of hepatocytes L02.The content of TG in hepatocytes,liver enzymes(ALT,AST,ALP and LDH) in supernatant were measured by automatic biochemical analyzer,and the accumulations of lipid droplets in hepatocytes with oil red O staining were observed on optical microscope.To assesse and identify advantages of the steatosis model in different times.②Selecting the optiomized concentration:After establishing a model of steatosis of hepatocytes L02,it was divided into 9 groups:normal group,spontaneous recovery group,and 7 PNS treatment groups(1μg/mL,5μg/mL,10μg/mL,50μg/mL,100μg/mL,150μg/mL,2001μg/mL).Growth inhibition and avirulent concentrations of PNS on steatotic hepatocytes were performed using MTT assay.③Effeetion and observation:After establishing a model of steatosis of hepatocytes L02,sequently it was divided into 5 groups:control group,model group,spontaneous recovery group,PNS 10μg/mL group and PNS 50μg/mL group.Meantime,hepatocytes in model group were continuously cultured by 50%calf serum while others were cultured by 10%calf serum.After 24 hours and 48hours respectively,content of TG in each group was measured by automatic biochemical analyzer,as well as lipid droplets in hepatocytes were detected by oil red O staining.And the expressions of UCP2 mRNA in hepatocytes were detected by RT-PCR after 24 hours.Results:①The establishment of steatotie model in hepatocytes:Compared with control group of 24 hours,content of TG in model group were significantly increased(P＜0.01),and levels of ALT and ALP were not change obviously(P＞0.05),while levels of AST and LDH dramatically increased(P＜0.01) and lipid droplets in hepatocytes observed by oil red O staining were increase slightly.After 48 hours and 72 hours,levels of TG,ALT,AST,ALP and LDH were increased significantly(P＜0.05,P＜0.01),and lipid droplets in hepatocytes observed by oil red O staining were increase significantly,even forming the shapes as chains or rings.②Selection of optiomized concentration:Compared with spontaneous recovery group,there were no effect in growth inhibition and avirulent concentrations of each dose group of PNS(1μg/mL,5μg/mL,10μg/mL,50μg/mL) after 24 hours and 48 hours(P＞0.05),while the groups of PNS100μg/mL,150μg/mL and 200μg/mL have statistical significance(P＜0.05,P＜0.01).③Content of TG in hepatocytes:Compared with model group,content of TG in spontaneous recovery group and PNS groups(10μg/mL and 50μg/mL) was significantly declined for 24 hours and 48 hours(P＜0.01).Compared with spontaneous recovery group, content of TG in PNS treatment groups for 24 hours significant decreased,The accumulation of lipid droplets in each group observed by oil red O staining corresponded to its content of TG.④The expression of UCP2mRNA:Compared with normal group,expression of UCP2 mRNA in model group was significantly up-regulated(P＜0.01);Compared with spontaneous recovery group,expression of UCP2 mRNA in high concentration group of PNS has declined(P＜0.05),while the low concentration group of PNS has no statistical significance(P＞0.05).Conclusion:①A model of steatosis of hepatocytes L02 was established by induction with 50%calf seruin.②PNS can make the content of TG in steatosis of hepatocytes decline;and exposed to normal medium can reverse steatosis of hepatocytes,so they can promote each other.③Expression of UCP2 mRNA was close related to the steatosis of hepatocyte;PNS may play a role by downregulating the level of UCP2 mRNA in the hepatocytes of steatosis model.④Lipid accumulation in steatotic hepatocytes may be related to the Traditional Chinese Medicine Theory of "Wet mudd and Phlegm".Between downreguLating TG in steatosis of hepatocytes of PNS and Notoginseng for the thoery of"Huoxuehuayu" may have relevance.