Down-regulation of TNF-alpha receptors by conophylline in human T-cell leukemia cells

In the course of our screening for tumor necrosis factor-alpha (TNF-alpha) function inhibitors, conophylline, a vinca alkaloid isolated from the plant Ervatamia microphylla, was found to inhibit TNF-alpha-induced NF-kappaB activation. We studied the effect of conophylline on TNF-alpha-induced NF-kappaB and JNK activations in human T-cell leukemia Jurkat cells. Conophylline inhibited both of these TNF-alpha-induced activations. It also inhibited phosphorylation and degradation of I-kappaB-alpha. Moreover, a receptor binding assay using [125I]-TNF-alpha showed that this inhibitory effect was due to a decrease in the binding of TNF-alpha to the cells. Scatchard analysis of the binding data indicated that conophylline induced only a small change in the affinity of the receptors but a significant change in the receptor number. FACS analysis showed that conophylline reduced the expression of CD120a/TNFR1, the high-affinity receptor for TNF-alpha, on the cell surface. On the other hand, conophylline did not affect the kinetics of internalization and degradation of TNF-alpha/receptor complexes or the half-life of TNF-alpha binding sites. These results indicate that conophylline down-regulates the expression of the TNF-alpha receptors on the cell surface.