The Mechanism Studies About Activity Of Uridine Diphosphate Glucuronyl-transferase In Liver Induced By S-adenosyl-l-methionine

Neonatal hyperbilirubinemia is a common disease of the neonatalperiod, it can cause multiple organ system damage. The key of the treatmentof neonatal hyperbilirubinemia is to reduce the serum bilirubin levelquickly. Three aspects of clinical treatment are light therapy、bloodtransfusion therapy and medication in commonly，these aspects haveadvantages and disadvantages. Therefore，looking for a safe、economicaland effective drugs to reduce serum bilirubin level have outstandingsignificance in neonatal hyperbilirubinemia treatment. Uridinediphosphate glucuronyltransferase is a key enzyme of bilirubincombination, it could transfer the fat-soluble conjugated bilirubin intowater-soluble conjugated bilirubin and excrete it.S-adenosyl-L-methionine（SAMe) is the endogenous substances in thebody, participate many important biochemical reactions in our body. Itparticipate the synthesis of cysteine, taurine, glutathione and coenzymeA as substrate material. It as the gene donor or enzyme inducers play amethyl transfer, sulpho-transfer and C-ammonification(polyamine synth-esis)role in the three major metabolic process in human tissue, which usedclinically for many years, the indications is jaundice such asintrahepatic cholestasis、hepatitis and hyperbilirubinemia. It can reduce serum unconjugated bilirubin levels effectively, recent years, there havebeen reported that SAMe as adjuvant treat the neonatal hyperbilirubinemiahave a certain effect. Therefore, we from the point of view of molecularbiology, analyze the effect about SAMe on mRNA expression、proteinsynthesis and activity of uridine diphosphate glucuronyltransferase onthe human fetal liver cells(LO2cells).Part oneThe study of mRNA expression in S-adenosyl-L-methionine induced liver uridine diphosphateglucuronosyltransferaseObjectivesWe used SAMe to treat the LO2cells with different concentrations andtime， compared the change in mRNA levels of uridine diphosphateglucuronosyltransferase in LO2cells，and explored the UGT RNA levelchange and intervention conditions under the treatment of SAMe in LO2cells.MethodsWe cultured the LO2cells in vitro, until the cells70%covered thebottom of the plate and grouped it:（1）SAMe treatment group：accordingto the pre-experimental results， added SAMe and maked the finalconcentration were1mmol/L,0.5mmol/L and0.1mmol/L;（2）Positive controlgroup：the final concentration were2mmol/L benzene barbiturates and5mmol/L nikethamide;（3）The negative control group：without any drugtreatment; Collected the cells after cultured24h,48h,72h,92h.To detectUGT1A1mRNA levels of each group by quantitative-PCR.ResultsThe UGT1A1mRNA was increase after SAMe induced in LO2cells, the most obvious concentration is0.5mmol/L, the UGT1A1mRNA of this group beganto increase after SAMe induced24h（49.18±10.16，P<0.05）,peaked after48h（130.69±9.23，P<0.05），decreased significantly after72h（28.25±9.42，P<0.05），since then there was no significant change; Compared withthe liver enzymes inducer group，the0.5mmol/L SAMe group generated moreUGT1A1mRNA，and had longer induced time from LO2cells（P<0.05）.ConclusionsThe SAMe could induce UGT1A1mRNA expression in the LO2cells inappropriate concentration，the expression and aging were better than theliver enzyme inducer. Part twoThe study of protein expression in S-adenosyl-L-methionine induced liver uridine diphosphateglucuronosyltransferaseObjectivesWe used SAMe to treat the LO2cells with different concentrations andtime，compared the change in protein expression and excretion levels ofuridine diphosphate glucuronosyltransferase in LO2cells，and exploredthe UGT protein expression and excretion level change and interventionconditions under the treatment of SAMe in LO2cells.MethodsWe cultured the LO2cells in vitro, until the cells70%covered thebottom of the plate and grouped it:（1）SAMe treatment group：accordingto the pre-experimental results， added SAMe and maked the final concentration were1mmol/L,0.5mmol/L and0.1mmol/L;（2）Positive controlgroup：the final concentration were2mmol/L benzene barbiturates and5mmol/L nikethamide;（3）The negative control group：without any drugtreatment; Collected the cells after cultured24h,48h,72h,92h.To detectUGT1A1protein expression and excretion levels of each group by westernblot and ELISA.ResultsThe UGT1A1protein was increase after SAMe induced in LO2cells, themost obvious concentration is0.5mmol/L, the UGT1A1protein expressionand excretion level of this group began to increase after SAMe induced24h（49.18±10.16或526.18±5.21，P<0.05）,peaked after48h（130.69±9.23或1278.22±9.38，P<0.05），decreased significantly after72h（28.25±9.42或296.62±6.42，P<0.05），since then there was no significant change;Compared with the liver enzymes inducer group，the0.5mmol/L SAMe groupexpressed and excreted more UGT1A1protein，and had longer induced timefrom LO2cells（P<0.05）.ConclusionsThe SAMe could induce UGT1A1protein expression in the LO2cells inappropriate concentration，the expression and aging were better than theliver enzyme inducer.