ENDOTOXIN-INDUCED EARLY GENE-EXPRESSION IN C3H HEJ (LPS(D)) MACROPHAGES/

C3H/HeJ (Lps(d)) macrophages have been shown to respond to certain LPS s, especially from rough mutant bacteria. C3H/OuJ (Lps(n)) macrophages are induced by wild-type LPS, rough LPS, or lipid A to express many g enes, including TNF-alpha, TNFR-2, IL-1 beta, IP-10, D3, and D8. C3H/H eJ macrophages failed to induce any of these genes when cultured with wild-type LPS or synthetic lipid A, even when pretreated with IFN-gamm a. However, rough mutant Salmonella minnesota Ra, Rc, and Rd LPS, and Escherichia coli D31 m3 Rd LPS induced Lps(d) macrophages to express a subset of genes within the gene panel. Because bioactive preparations contained trace quantities of endotoxin protein(s), a deoxycholate-mo dified, phenol-water method was used to repurify rough LPS into an aqu eous phase, and extract endotoxin proteins into a phenol phase. Repuri fied LPS failed to stimulate Lps(d) macrophages; however, phenol fract ions were similar to 10% as potent in Lps(d) macrophages as crude roug h LPS. Full potency was restored in C3H/HeJ macrophages when aqueous p hase LPS and phenol-phase proteins were coprecipitated, suggesting tha t LPS and endotoxin proteins interact synergistically. Endotoxin prote ins alone induced TNF-alpha, TNFR-2, and IL-1 beta, but not IP-10, D3, and D8 genes in both Lps(d) and Lps(n) macrophages. Tyrosine phosphor ylation of three 41- to 47-kDa proteins was induced by endotoxin prote ins, but not by LPS, in Lps(d) macrophages. Thus, endotoxin proteins s eem to activate a signaling pathway(s) that converges (distal to the L ps gene product) with a subset of LPS-signaling pathways.