Cholesterol overloading leads to hepatic L02 cell damage through activation of the unfolded protein response.
Reported data indicate that cholesterol loading in the liver can cause hepatic injury. To explore the possible mechanisms of cell damage resulting from cholesterol overloading in hepatocytes, cell apoptosis, the unfolded protein response (UPR) and the correlation between them were assessed in the cholesterol-overloaded normal human hepatic cell line L02. L02 cells were incubated with 200 microg/ ml of low density lipoprotein (LDL) for 24 h with or without 20 microg/ml 58035, an inhibitor of acyl-CoA:cholesterol acyltransferase (ACAT). In the LDL+58035 group, the intracellular cholesterol level was dramatically increased, which was measured by an enzymatic combined high performance liquid chromatography assay. Expression of immunoglobulin-binding protein, X-box binding protein 1, activating transcription factor 6, activating transcription factor 4, CCAAT/enhancer-binding protein homologous protein-10, markers of endoplasmic reticulum stress (ERS)/ UPR, were up-regulated as determined using reverse transcription-polymerase chain reaction (RT-PCR) or Western blot analysis. The rate of cell apoptic death increased 21.3+/-2.4%. Meanwhile, the active caspase-3 protein expression was increased 8.4-fold compared to the active caspase-3 protein expression in the controls. Furthermore, 4-phenylbutyric acid, an inhibitor of UPR, partly reduced cell apoptosis and activation of caspase-3. This study suggests that cholesterol overloading in hepatic L02 cells induces ERS and activates the UPR which, in part, leads to the apoptotic damage of cells.
Li
Qi
Q
Department of Biochemistry and Molecular Biology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, PR China.
Liu
Zhiguo
Z
Guo
Jianli
J
Chen
Jiangyuan
J
Yang
Pu
P
Tian
Jun
J
Sun
Jun
J
Zong
Yiqiang
Y
Qu
Shen
S
D051701:Activating Transcription Factor 4 / Q000235:genetics; D051702:Activating Transcription Factor 6 / Q000235:genetics / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D002460:Cell Line; D002784:Cholesterol / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics; D004791:Enzyme Inhibitors / Q000494:pharmacology; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism*; D006801:Humans; D008077:Lipoproteins, LDL / Q000494:pharmacology; D010654:Phenylbutyrates / Q000494:pharmacology; D000071701:Regulatory Factor X Transcription Factors; D020133:Reverse Transcriptase Polymerase Chain Reaction; D002785:Sterol O-Acyltransferase / Q000037:antagonists & inhibitors; D014157:Transcription Factors / Q000235:genetics; D056811:Unfolded Protein Response / Q000187:drug effects / Q000502:physiology*
D051702:Activating Transcription Factor 6; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; D008077:Lipoproteins, LDL; D010654:Phenylbutyrates; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors; D051701:Activating Transcription Factor 4; C075773:4-phenylbutyric acid; D002784:Cholesterol; D002785:Sterol O-Acyltransferase; D053148:Caspase 3