Endotoxin-stimulated human macrophages produce a factor that induces polymorphonuclear leucocyte infiltration and is distinct from interleukin-1, tumour necrosis factor α and chemotactic factors

Previously we reported that rabbit macrophages (MΦ) in the presence of nanogram quantities of endotoxin (LPS) release factors that induce polymorphonuclear leucocyte (PMNL) infiltration into the skin of rabbits following i.d. injection. The predominant factor was a de novo synthesized protein of 45,000 MW on gel filtration that was distinguishable for IL-1 but not from TNFα. Here we examined human monocytes, in vitro monocyte-derived MΦ and peritoneal MΦ for the production of an analogous protein. Upon stimulation with LPS, they all rapidly (6 hr) produced a factor(s) that caused PMNL accumulation in the skin of rabbits when injected i.d. This activity, referred to as PMNL-recruiting activity (PRA), was heat labile and its production was blocked by cycloheximide. By Sephadex-G100 chromatography the major PRA of cultured MΦ or peritoneal MΦ had a molecular weight (MW) of 45,000-60,000. The active fractions were free of IL-1 (< 0.2 U/ml) and Superose-12 FPLC chromatography separated the peak of PRA, which eluted at 45,000 MW, from TNFα, eluting at 20,000 MW. The peak PRA was not neutralized by antisera to IL-1α, IL-1β, TNFα, IL-6 or GM-CSF, indicating that it was distinct from these cytokines. The major PRA did not induce the migration of PMNL in vitro in a filter chemotaxis assay. In contrast to the MΦ, the major PRA produced by LPS-stimulated monocytes eluted at 15,000-20,000 MW, contained IL-1 activity and was neutralized by antisera to IL-1α and IL-1β. Monocytes from a few donors also produced the 45,000-60,000 MW PRA simultaneously. We conclude that human peritoneal MΦ and in vitro monocyte-derived MΦ exposed to LPS secrete a protein of 45,000-60,000 MW, which is a potent inducer of PMNL infiltration but is distinct from IL-1, TNFα, IL-6, GM-CSF and PMNL chemotactic factors.