The Effect Of Polygonum Multiflorum And Its Preparation On Rat And Human Liver Cytochrome P450 Enzymes

Objective:This study investigated the changes of cytochrome P450 in vitro and in vivo levels to elucidate the metabolic mechamism of Polygonum multiflorum through the level phase I metabolism.Methods:In this work,the activities of five CYP450 enzymes[CYP1A2?CYP2E1?CYP3A4(CYP3A1)?CYP2D6 and CYP2C9]in rat liver were determined after SD rat were lavaged with water extract of Polygonum multiflorum(WE)at two dosage levels(1 g/kg BW/d,10 g/kg BW/d)and water extract of processed Polygonum multiflorum(PWE)at two dosage levels(2 g/kg BW/d,20 g/kg BW/d),respectively for 7 consecutive days,then the liver microsomes were prepared.By comparison with treated group(the different doses of water extract of Polygonum multiflorum and water extract of processed Polygonum multiflorum)and the group of distilled water,the group of the phenobarbital in the levels of liver coefficient and CYP450 level.Cocktail probe substrate and liver microsome in vitro temperature incubation method were adopted.Meanwhile,the metabolic elimination percentages of the five probe substrates were detected with HPLC,in order to evaluate the effect of each administration group on the enzymatic activity of rat liver microsome CYP450.The mRNA expression levels of CYP1A2,CYP2E1 and CYP3A1(CYP3A4)in rat liver were determined by real time quantitative reverse transcription polymerase chain reaction(qRT-PCR).The vitro experiment in cell:Normal human liver L02 cells and liver cancer HepG2 cells were treated by three dosage levels of water extract of Polygonum multiflorum(0.5 mg/mL,l mg/mL and 2 mg/mL),then when we using water extract of processed Polygonum multiflorum(0.45 mg/mL,0.9 mg/mL and 1.8 mg/mL),emodin(5 ug/mL,10 ug/mL and 20 ug/mL)and 2,3,5,4'-tetrahydroxy-stilbene-2-O-?-D-glucoside(75 ug/mL,150 ug/mL and 300 ug/mL)for 0?72h.And then cell growth inhibition of each group was determined by MTT method.The mRNA expression levels of CYP1A2 and CYP2E1 were determined by real-time quantitative PCR using specific target primers for CYP450 genes.Results:Compared with the control group,the liver coefficient of treated group had no obvious changes;the high dose of WE and PWE caused significant decrease in the activities of CYP1A2(P<0.01);meanwhile,the groups of WE caused a sigmificant decrease in CYP2E1 activity(P<0.01);whereas every treated group caused an obvious increase in the activities of CYP3A4(CYP3A1)(P<0.01).Compared with the control group,high dose of WE and PWE groups down-regulated CYP1A2 mRNA expression;both groups of WEdown-regulated CYP2E1 mRNA expression obviously.Whereas the CYP3A1(CYP3A4)mRNA expression of treated groups had no changes compared with the group of distilled water.The results of vitro experiment in cell revealed that compared with the blank control group,the level of CYP1A2 mRNA expression in L02 cells could be significantly inhibited by WE and PWE groups.Whereas the CYP1A2 and CYP2E1 mRNA expression in HepG2 cells of treated groups had no changes compared with the group of distilled water,it was the same with the CYP2E1 mRNA expression in L02 cells.Conclusion:WE and PWE can inhibit the rat liver microsome CYP1A2 and CYP2E1 expression significantly,this suggeststhat the inhibition mechanism of the enzymeactivities of CYP1A2 and CYP2E1 may be ascribed to its effect on the relative expression level of CYP450 enzymemRNA.WE 0.5 mg/mLand PWE 0.45 mg/mL can decrease the expressionlevels of CYP1A2 mRNA in L02 cells.