Differential tumor necrosis factor alpha expression and release from peritoneal mouse macrophages in vitro in response to proliferating cram-positiveversus gram-negative bacteria

Viable Escherichia call and Staphylococcus aureus bacteria elicited markedl y different in vitro tumor necrosis factor alpha (TNF-alpha) responses when placed in coculture with peritoneal murine macrophages. These include quan titative differences in TNF-alpha mRNA expression and corresponding protein product secretion as well as kinetic differences in the profiles of the TN F-alpha responses. Further, lipopolysaccharide (from E. coli) is a major co ntributing factor to these differences, as revealed by comparative experime nts with endotoxin-responsive (C3Heb/FeJ) and endotoxin-hyporesponsive (C3H /HeJ) macrophages. Nevertheless, the eventual overall magnitude of the TNF- alpha secretion of macrophages in response to S. aureus was at least equiva lent to that observed with E. coli, while appearing at time periods hours l ater than the E. coli-elicited TNF-alpha response. Both the magnitude and k inetic profile of the TNF-alpha responses were found to be relatively indep endent of the rate of bacterial proliferation, at least to the extent that similar results were observed with both viable and paraformaldehyde-killed microbes. Nevertheless, S. aureus treated in culture with the carbapenem an tibiotic imipenem manifests markedly altered profiles of TNF-alpha response , with the appearance of an early TNF-alpha peak not seen with viable organ isms, a finding strikingly similar to that recently reported by our laborat ory from in vivo studies (R. Silverstein, J. G. Wood, Q. Xue, M. Norimatsu, D. L. Horn, and D. C. Morrison, Infect. Immun. 68:2301-2308, 2000). In con trast, imipenem treatment off. coli-cocultured macrophages does not signifi cantly alter the observed TNF-alpha response either in vitro or in vivo. In conclusion, our data support the concept that the host inflammatory respon se of cultured mouse macrophages in response to viable gram-positive versus gram-negative microbes exhibits distinctive characteristics and that these distinctions are, under some conditions, altered on subsequent bacterial k illing, depending on the mode of killing. Of potential importance, these di stinctive in vitro TNF-alpha profiles faithfully reflect circulating levels of TNF-alpha in infected mice. These results suggest that coculture of per itoneal macrophages with viable versus antibiotic-killed bacteria and subse quent assessment of cytokine response (TNF-alpha) may be of value in clarif ying, and ultimately controlling, related host inflammatory responses in se ptic patients.