Ratio of bacteria to polymorphonuclear neutrophils (PMNs) determines PMN fate

This study was designed to investigate how live Escherichia coli influence the fate of polymorphonuclear neutrophils (PMNs) in vitro. PMNs from 10 healthy volunteers were cocultured with or without live E. coli at different ratios. Heat-killed E. coli (Hk) were also added to PMNs at a ratio of 1:10. The PMNs were then analyzed by flow cytometry for cell death, reactive oxygen intermediates (ROI) production, and CD16 expression. Morphologic features were also assessed. PMN apoptosis was confirmed by DNA gel electrophoresis. Low doses of E. coli (PMN:E. coli ratios of 1:0.01 and 1:0.1) inhibited PMN apoptosis. In contrast, a high dose of E. coli (PMN:E. coli ratio of 1:10) increased PMN necrosis. ROI production was significantly greater at PMN:E. coli ratios of 1:10 and 1:10 (Hk) than at ratios of 1:0.01 and 1:0.1, or in PMNs cultured alone after a 15 or 30 minute coculture. CD16 expressions were significantly lower in PMNs cocultured with E. coli than in those cultured alone after a 4 or 12-h coculture. Tumor necrosis factor-alpha, interleukin (IL)-1beta and IL-6 levels in cell-free supernatants were also measured. The mean percentages of apoptosis at PMN:E. coli ratios of 1:0.01 and 1:10 (Hk), and in PMNs cultured alone after a 12-h coculture showed significant inverse correlations with these cytokine levels in cell-free supernatants at 12 h. Our results demonstrate that low doses of live E. coli inhibits predominantly PMN apoptosis, whereas a high dose of E. coli increases necrosis. Augmented PMN bactericidal function, via inhibition of PMN cell death, may be beneficial for host defense against bacterial infection and/or sepsis.