Functional characterization of cytokine autoantibodies in chronic renal failure patients

Abstract

We have recently reported on an increase of IL-1 alpha and IL-6 autoantibodies in patients maintained on chronic hemodialysis treatment. The aim of the present study was to evaluate functional properties of these autoantibodies. Serum samples of more then 500 chronic renal failure patients with and without replacement therapy were screened for the presence of IL-1 alpha and IL-6 autoantibodies by second antibody precipitation. The neutralizing capacity of IL-1 alpha autoantibody serum was studied by immunofluorescence flow cytometry analysis of IL-1 alpha induced expression of E-selectin (ELAM-1, CD62e) on HUVEC (human umbilical vein endothelial cells). Results of these inhibition studies were confirmed with IgG preparations from antibody positive sera, purified by affinity chromatography. Functional studies on IL-6-dependent B9 cell proliferation were performed with IL-6 autoantibody positive sera, and quantitated with the colorimetric MTT assay. IL-1 alpha induced expression of E-selectin on HUVEC (considered 100% positive cells) was inhibited by each IL-1 alpha autoantibody positive serum sample (N = 13; anti-IL-1 alpha activity: 7.62 to 57.52% binding). Inhibition of E-selectin expression by IL-1 alpha autoantibodies ranged from 0.11 to 80.22% positive cells (0.15 to 92.31% mean fluorescence intensity). A strong correlation of E-selectin expression with IL-1 alpha autoantibody concentration was observed (P < 0.005). Furthermore, IgG eluates from autoantibody positive patients inhibited E-selectin expression to 41.0 +/- 23.1% positive cells if compared with 83.7 +/- 5.7% positive cells of the IgG depleted serum samples.(ABSTRACT TRUNCATED AT 250 WORDS)