The In Vitro Study Of The Correlation Between Proteasome Inhibition And Endoplasmic Reticulum Stress In Alcoholic Liver Disease

Objective To study the potential correlation between inhibited proteasome activity and the occurrence and development of endoplasmic reticulum stress(ERS) in alcoholic liver disease(ALD) by building the in vitro model of ALD and making an assay of proteasome activity and mRNA expression of ERS marker glucose regulation protein 78 (GRP78).We choose the measurement of cellular triglycercide(TG) content in different groups to show the pathologic effects of ERS.Finally,we make correlation analysis between proteasome activity and ERS marker mRNA expression.Methods HepG2 cells were treated with ethanol(150mmol/L) or betulinic acid(20μmol/L)or both of them or DMEM,marked as ethanol group,betulinic acid group.ethanol plus betulinic acid group and control group respectively.We detected the mRNA expression of ERS maker GRP78 by PCR.In addition,we adopted fluorescent substrate assay(S-LLVY-AMC) method to detect the activity of proteasome.The measurement of intracellular triglyceride content was done by GPO-PODase method.Finally,we made correlation analysis between proteasome activity and GRP78 mRNA expression.Results Ethanol inhibited the activity of proteasome,with the increase of GRP78 mRNA expression and intracellular TG content.Betulinic acid promoted the proteasome activity with the reduction of GRP78 mRNA level and intracellular TG content.Correlation analysis revealed negative relativity between proteasome activity and the expression of the GRP78 mRNA level.While the GRP78 mRNA level and the intracellular triglyceride content had positive relativity.Conclusions Ethanol can inhibit proteasome activity thus induce ERS while betulinic acid promote proteasome activity and reduce ERS.The proteasome activity and ERS have some negative relativity and the latter will aggravate the pathogenesis of ALD.