Effects Of NF-ΚB On ABCA1 Expression And Cholesterol Efflux In THP-1 Derived-foam Cells Induced By AngII

PartⅠEffects of NF-κB on cholesterol content and efflux in THP-1 derived-foam cells induced byAngIIObjective To investigate whether NF-κB activation was involved in regulation of Cellular cholesterol content and cholesterol efflux in THP-1 derived-macrophage foam cells induced by AngIIMethods the THP-1 cells were induced into macrophages by PMA and transformed into foam cells by ox-LDL(50mg/L) for 48 hours.The foam cells were devided into three groups(1)treated with PBS, used for control group(,2)treated with AngII(10ng/ml) for 24h, used for AngII group,(3)treated with TPCK(10μmol/L) for 60min before AngII (10ng/ml) stimulation for 24h, used for TPCK group. Then the variance of cholesterol content was measured by zymochemistry via fluorospectrophotometer. To evaluate cholesterol efflux(liquid scintillation counting assay), [3H]cholesterol ( 0.37×106 Bq/ml ) and ox-LDL(50mg/L) were also added in this 48h period; Then the cells were divided into three groups as mentioned.Results Treatment of THP-1 derived foam cell with AngⅡresulted in an increase in cholesterol content by 140.5%( p<0.05) and a reduction in cholesterol efflux by 46.4%( p<0.05), as compared with control group. In contrast, addition of TPCK: inhibitor of NF-κB before AngⅡcaused a reduction in cholesterol content and an increase in cholesterol efflux by 24.1%(p<0.05) and 41.1%(p<0.05), respectively, as compared with AngⅡgroup.Conclusion AngⅡincreased cholerterol content and decrease cholesterol efflux in THP-1 derived foam cells markedly, which accelerated the development of atherosclerosis. Pretreatment with TPCK partially reversed the effect of AngⅡon cholesterol content and cholesterol efflux, which played a critical role in atherosclerosis acceleration by AngⅡ. PartⅡEffects of NF-κB on ABCA1 expression in THP-1 derived-foam cells induced by AngIIObjective To investigate whether NF-κB activation was involved in regulation of the expression of ABCA1 mRNA and protein in THP-1 derived-macrophage foam cells induced by AngII.Methods the THP-1 cells were induced into macrophages by PMA and transformed into foam cells by ox-LDL(50mg/L) for 48 hours.The foam cells were devided into three groups(1)treated with PBS, used for control group(,2)treated with AngII(10ng/ml) for 48h, used for AngII group,(3)treated with TPCK(10μmol/L) for 60min before AngII (10ng/ml) stimulation for 48h, used for TPCK group. Then the level of ABCA1 mRNA and protein were analyzed by RT-PCR and Westernblot, respectively. Sandwich ELISA was employed to evaluate the nuclear translocation of NF-κB subunit p65 at 0, 30min, 1h, 2h and 4h.Results The expression of ABCA1 mRNA and protein reduced 41% and 30.4%(p<0.05) by AngII, as compared with control group. Addition of TPCK before AngII partially reversed these responses, the ABCA1 mRNA and protein were increased by 30% and 19%(p<0.05), respectively, as compared with AngII group.AngII stimulation caused a rapid increase of NF-κBp65 nuclear translocation and the peak of NF-κB p65 was 30-60min in foam cells but Addition of TPCK to the cells for 2h before AngII stimulation attenuated the response of NF-κBp65 nuclear translocation induced by AngII and showed no peak in foam cells group.Conclusion AngII results in NF-κB activation which finally down-regulated ABCA1 gene expression and cholesterol efflux in THP-1 derived-foam cells, inhibition of NF-κB eliminates the responses of AngⅡpartially, which plays a critical role in atherosclerosis acceleration. Thus, AngII signals partially through NF-κB to repress ABCA1 expression in THP-1 derived-foam cells.