TNF reduces LIF endocytosis despite increasing NFκB-mediated gp130 expression

To examine how the proinflammatory cytokine tumor necrosis factor alpha (TNF) modulates the response of cerebral microvessels to other cytokines, we used rat cerebral microvessel endothelial RBE4 cells to simulate the in vitro blood-brain barrier (BBB). The gp 130 receptor, which is shared by the interleukin (IL)-6 family of cytokines, showed specific upregulation by TNF. TNF treatment (5 ng/ml for 30 min to 24 h) increased gp 130 at both the levels of transcription and protein expression. The stability of gp 130 protein was mediated by NF kappa B activity, as the inhibitors quinazoline and MG 132 not only blocked the increase induced by 6 h of TNF treatment, but also reduced its basal level of expression. By contrast, the lysosomal inhibitor chloroquine and the extracellular regulated kinase inhibitor U0126 showed no effect. Despite the increase of gp 130, TNF caused a significant reduction in the cell binding and endocytosis of leukemia inhibitory factor (LIF), another proinflammatory cytokine that binds to the gp 130 co-receptor and its unique gp 190 receptor. This is consistent with our previous findings that TNF reduces gp 190 expression and Stat3 activation. Thus, TNF stimulation results in decreased responsiveness of RBE4 cells to LIF, indicating complex regulatory interactions of cytokines at the BBB.