Figure 3 From: Tumor necrosis factor alpha and adalimumab differentially regulate CD36 expression in human monocytes Mechanisms involved in the regulation of CD36 expression by tumor necrosis factor (TNF)α. (a) TNFα inhibits both basal and rosiglitazone-induced peroxisome proliferator-activated receptor (PPAR)γ activation. Human monocytes were incubated with macrophage-serum-free medium (M-SFM) alone (control), or with M-SFM containing TNFα (10 ng/ml) for 5, 30 or 60 minutes, and then stimulated, or not, with a synthetic ligand of PPARγ, rosiglitazone (R; 5 μmol/l) for 45 minutes. Nuclear proteins were isolated and a [γ-32P]ATP labeled oligonucleotide expressing the PPAR DNA consensus binding sequence was added. PPARγ activation was analyzed by gel-shift assay. (b) TNFα inhibits both basal and rosiglitazone-induced CD36 mRNA expression. Human monocytes were incubated with M-SFM alone (control), or with M-SFM containing TNFα (10 ng/ml) for 1 h and then stimulated or not with rosiglitazone (R; 5 μmol/l) for 4 h. CD36 mRNA expression was quantified using RT-PCR and normalized to β-actin. Data represent the mean ± standard error (SE) of the relative quantification of CD36 mRNA expression measured in three experiments. *Significantly different from control (p < 0.05). (c) TNFα induces reactive oxygen species production. Monocytes were incubated with Hanks balanced salt solution (HBSS) alone (control), or with HBSS containing TNF (10 ng/ml) for 1 h. Reactive oxygen species production was measured by chemiluminescence in the presence of 5-amino-2,3-dihydro-1,4-phthalazinedione in a thermostatically controlled luminometer. Data represent total chemiluminescence emission (area under the curve) for 1 h, measured in three experiments. *Significantly different from the control (p < 0.05). (d) The decrease in CD36 membrane expression induced by TNFα is not inhibited by an anti-oxidant (Trolox). Monocytes were incubated with M-SFM alone (control), or with M-SFM containing either TNFα (10 ng/ml), Trolox® (1 μM), or TNFα combined with Trolox® for 24 h and the membrane expression of CD36 expression was quantified using flow cytometry. Data represent the geometric mean ± SE of the fluorescence measured in three experiments in duplicate. *Significantly different from the control (p < 0.05).Back to article page