Effects And Mechanisms Of Tumor Necrosis Factor-α On The Expression Of ABCA1 In THP-1 Macrophage Foam Cells?

Part I Effects of tumor necrosis factor-αon intracellular cholesterol content and the expression of ABCA1 in THP-1 macrophage foam cellsObjective To observe the influence of Tumor Necrosis Factor-α(TNF-α) on the expression of ATP binding cassette transporter A1(ABCA1) and ABCA1-mediated cellular cholesterol efflux in THP-1 macrophage foam cells.Methods THP-1 cells were be transformed into foam cells. Then the content of cellular cholesterol in foam cells was estimated as cells were stimulated with TNF-αat different concentrations and times. Subsequently foam cells were treated with TNF-αat appropriate concentration(10ng/ml)or without TNF-αstimulation. Gene expression of ABCA1 was analyzed using RT-PCR and ABCA1 protein was detected by Western-blot.Results TNF-αdecreased intracellular cholesterol content in concentration and time dependent manners. 10ng/ml TNF-αdown regulated the levels of ABCA1 mRNA and ABCA1 protein in time dependent manner.Conclusions TNF-αmight decrease cellular cholesterol efflux mainly via the down-regulation of ABCA1,which finally leads to the accumulation of intracellular cholesterol. Part II Relationship between activation of nuclear factor-κB and ABCA1 expression in THP-1 derived-macrophage foam cellsObjective To investigate whether nuclear factor (NF-κB) activation participate in the regulation and control of ABCA1 gene expression in THP-1 derived-macrophage foam cells.Methods First, THP-1 cells were be transformed into foam cells. Then, foam cells were treated with TNF-α(10ng/ml), or with TPCK(10μM) at 60min before TNF-αstimulation or at different time points after TNF-αstimulation. Gene expression of ABCA1 was analyzed using RT-PCR and its protein was detected by Western-blot. Sandwich ELISA method was employed to evaluate the nuclear translocation of NF-κB subunit p65.Results Stimulation of foam cells by TNF-αled to a rapid increase of NF-κBp65 nuclear translocation and the peak of NF-κB p65 was 30-60min. TNF-αcould down regulate both ABCA1 mRNA expression and ABCA1 protein level. Addition of TPCK to the cells at 60 min before TNF-αattenuated the increase of nuclear factor( NF-κB) translocated into the nucleus and the decrease of ABCA1 gene expression induced by TNF-α. Contrastively, there was no difference in the level of ABCA1 gene expression between cells incubated with TNF-αand cells added TPCK 4h after stimulation with TNF-α.Conclusions TNF-αcould swiftly result in NF-κB activation. Activation of NF-κB signal transduction may finally inhibit ABCA1 gene expression, which may play a critical role in cellular cholesterol efflux.