Lipid A-associated proteins provide an alternate "second signal" in the activation of recombinant interferon-gamma-primed, C3H/HeJ macrophages to a fully tumoricidal state

Abstract

Previous studies have shown that the activation of murine macrophages to a fully tumoricidal state requires that specific environmental signals be delivered to the macrophage in a stepwise manner: a "priming" signal first renders the macrophage responsive to a second or "trigger" signal. One potent "priming" signal has been identified as the T cell-derived lymphokine, interferon-gamma (IFN-gamma) and one often used "trigger" signal is lipopolysaccharide (LPS), the endotoxin derived from Gram-negative bacteria. In these studies, endotoxin-responsive C3H/OuJ (Lps(n)) and endotoxin-hyporesponsive C3H/HeJ (Lps(d)) macrophages were exposed in vitro to recombinant IFN-gamma (rIFN-gamma) and various preparations of endotoxin or purified lipid A-associated proteins (LAP). The resultant tumoricidal responses were evaluated to define the activation requirements of murine macrophages and to examine further the LPS defect exhibited by C3H/HeJ mice. The findings presented herein demonstrate that C3H/OuJ macrophages primed by rIFN-gamma respond to protein-free LPS (phenol-water extracted LPS), protein-rich LPS (butanol-extracted LPS), or purified LAP. In contrast, rIFN-gamma-primed C3H/HeJ macrophages failed to become cytolytic with phenol-water extracted LPS, but could be rendered fully tumoricidal if either butanol-extracted LPS or LAP were used as "second signals." These data indicate that C3H/HeJ macrophages are fully responsive to the priming effects of IFN-gamma, but remain restricted in their capacity to recognize protein-free LPS as a second signal. Alternate second signals, such as LAP, may provide a compensatory pathway by which these macrophages are rendered fully tumoricidal.