Modulation of the IL-1 cytokine network in keratinocytes by intracellular IL-1α and IL-1 receptor antagonist

Abstract

The IL‐1 cytokine network in epidermal cells was studied in vitro, using the spontaneously transformed HaCAT human keratinocyte line. Intracellular (ic) IL‐1α and IL‐1 receptor antagonist protein (IL‐1Ra) following cell lysis were readily identified assayed using a capture ELISA; whereas in culture supernatants IL‐1Ra was not detected, and IL‐1α was present at only very low levels. Confluent cultures of HaCAT cells were shown to provide optimal conditions for the study, since confluence increased the icIL‐1Ra: IL‐1α ratio to a level as seen in vivo, which was independent of Ca²⁺ concentration in the culture medium. The IL‐1Ra extracted from HaCAT cell lysates was functionally active, as demonstrated in the mouse thymocyte co‐proliferation assay which could be blocked using a rabbit anti‐IL‐1Ra antibody. Transforming growth factor‐beta (TGF‐α) stimulated a dose‐dependent increase in HaCAT cell 1L‐1α without changing IL‐1Ra concentration, with a resultant reduction in the iclL‐1Ra: IL‐1α ratio from 320:1 to 100:1. Similarly, TGF‐α, interferon‐gamma (IFN‐γ), IL‐6, and tumour necrosis factor‐alpha (TNF‐α) substantially increased HaCAT cell IL‐1α, but had no effect on the IL‐1Ra. with a concomitant reduction in the icIL‐1Ra: IL‐1α ratio. In contrast to their effects on monocytes, IL‐4and IL‐10 at biologically active levels had no effect on IL‐1α, IL‐1Ra or the icIL‐1Ra: IL‐1a ratio in confluent HaCAT cells. Hydrocortisone reduced IL‐1α to below the limit of sensitivity of the ELISA, and induced a small increase in IL‐1Ra of questionable biological significance. Thus, regulation of the IL‐1 cytokine network in keratinocytes involves modulation of icIL 1α rather than of icIL‐1Ra levels, and is markedly different from that noted in monocytes.