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. 2007 Mar 2;9(2):R22. doi: 10.1186/ar2133

Figure 3.

Figure 3

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).