TGF-beta1 prevents up-regulation of the P2X7 receptor by IFN-gamma and LPS in leukemic THP-1 monocytes.

摘要

The P2X7 receptor is an extracellular ATP-gated cation channel critical in inflammation and immunity, and can be up-regulated by IFN-gamma and LPS. This study aimed to examine the effect of TGF-beta1 on the up-regulation of P2X7 function and expression in leukemic THP-1 monocytes differentiated with IFN-gamma and LPS. Cell-surface molecules including P2X7 were examined by immunofluorescence staining. Total P2X7 protein and mRNA was assessed by immunoblotting and RT-PCR respectively. P2X7 function was evaluated by ATP-induced cation dye uptake measurements. Cell-surface P2X7 was present on THP-1 cells differentiated for 3days with IFN-gamma and LPS but not on undifferentiated THP-1 cells. ATP induced ethidium(+) uptake into differentiated but not undifferentiated THP-1 cells, and the P2X7 antagonist, KN-62, impaired ATP-induced ethidium(+) uptake. Co-incubation of cells with TGF-beta1 plus IFN-gamma and LPS prevented the up-regulation of P2X7 expression and ATP-induced ethidium(+) uptake in a concentration-dependent fashion with a maximum effect at 5ng/ml and with an IC(50) of ~0.4ng/ml. Moreover, ATP-induced YO-PRO-1(2+) uptake and IL-1beta release were abrogated in cells co-incubated with TGF-beta1. TGF-beta1 also abrogated the amount of total P2X7 protein and mRNA induced by IFN-gamma and LPS. Finally, TGF-beta1 prevented the up-regulation of cell-surface CD86, but not CD14 and MHC class II, by IFN-gamma and LPS. These results indicate that TGF-beta1 prevents the up-regulation of P2X7 function and expression by IFN-gamma and LPS in THP-1 monocytes. This suggests that TGF-beta1 may limit P2X7-mediated processes in inflammation and immunity.