Evidence against the existence of a membrane form of murine IL-1α

Previous studies have demonstrated that paraformaldehyde-treated macrophages possess IL-1α activity in a variety of bioassay systems. However, no definitive biochemical data in support of the membrane IL-1α concept has been reported. The purpose of the present study was to determine if the biologic activity associated with treated cells is due to a membrane form of IL-1α or alternatively, to the leakage of IL-1α. If the former case was true, then the exposed membrane IL-1α should bind anti-IL-1α antibodies or be cleaved by mild trypsin treatment. In both instances, IL-1α activity should be lost when measured in a subsequent IL-1 bioassay. Our results indicate that pulsing paraformaldehyde-treated normal or cell line macrophages with anti-IL-1α antibodies or treating the cells with trypsin did not affect the ability of the treated cells to function in a murine thymocyte proliferation assay. Furthermore, the standard short term treatment of cells with paraformaldehyde (15 min) did not prevent the leakage of IL-1α from the cells or the processing of the precursor forms of the protein. When cells were treated with paraformaldehyde for 2 h, they no longer released IL-1α or possessed thymocyte stimulatory activity. We also found that short term glutaraldehyde treatment of macrophages completely blocked the release of IL-1α from cells as well as the appearance of cell-associated IL-1α activity. Our results support the conclusion that the stimulatory activity of paraformaldehyde-treated macrophages is not due to a membrane form of IL-1α but is, in fact, due to the continuous release of IL-1α from the cells.