Inhibition of T Cell Apoptosis by IL–6 at Sites of Inflammation: Direct and Trans–Signalling Through the IL–6R Are Highly Regulated on Human T Cells

Abstract

Abstract: : Purpose: We have previously reported the inhibition of growth factor deprivation–induced T cell apoptosis by uveitic aqueous humor (AqH) through IL–6/soluble IL–6R trans–signalling. In this study we have investigated the role of direct IL–6 signalling by analyzing the expression of IL–6R and the signalling component gp130 during human T cell activation and differentiation. Methods: Peripheral blood mononuclear cells were isolated from normal healthy controls. Cells were stained with specific antibodies against IL–6R and gp130, in combination with antibodies to the surface markers CD4, CD8, CD45RA/RO and CCR7, and analysed by flow cytometry. Highly purified populations of CD4⁺RO⁺ T cells were isolated by magnetic bead depletion. Following antigenic stimulation or culture in IL–6, cells were analysed by real–time PCR and flow cytometry for the expression of IL–6R and gp130. The functionality of the receptor complexes was tested by culturing with cytokine and measuring the phosphorylation of Stat–1. Results: Peripheral blood CD4⁺CD45RO⁺ T cells, cultured in the presence of IL–6, down–regulated the surface expression of both IL–6R and gp130. In contrast, activation with anti–CD3 antibodies resulted in a reduction in the expression of IL–6R, but not gp130. Analysis of CD4+ naïve (CD45RA⁺) and memory (CD45RO⁺) lymphocytes in peripheral blood showed a reduction in gp130 as the cells differentiated, whereas IL–6R was maintained at a high level. The phosphorylation of Stat–1 correlated with the surface expression of gp130. Conclusions: The relative roles of direct and trans–signalling through the IL–6R were thought to be controlled only by the expression of IL–6R. However, our results suggest that expression and signalling through the gp130 chain, previously thought to be ubiquitously expressed, is highly regulated during T cell differentiation. This suggests that at sites of inflammation, including the uveitic eye, both direct and trans–signalling through the IL–6R may control T lymphocyte function.

Keywords: apoptosis/cell death • cytokines/chemokines • inflammation