Figure S1 - Global Analysis of Neutrophil Responses to Neisseria gonorrhoeae Reveals a Self-Propagating Inflammatory Program

Phagocytosis and production of reactive oxygen species are not essential for inflammatory cytokine production. (A) Cytochalasin D treatment prevents bacterial uptake by CEABAC PMNs. WT and CEABAC PMNs were left untreated or pre-incubated with cytochalasin D (10 mg/ml) for 10 min prior to infection with either Opa− or Opa+ N. gonorrhoeae (MOI 10). Extracellular and total bacteria were differentially stained, and quantified via immunofluorescence microscopy. (B) Human CEACAM1-expressing PMNs do not induce inflammatory cytokine production in response to Opa+ N. gonorrhoeae. Mouse (WT, CEABAC, and TG418) neutrophils were infected with either Opa− or Opa+ N. gonorrhoeae (MOI 10). MIP-1α and MIP-2 was measured 3 h post infection. Data shown as fold induction by Opa+ infection over Opa− infection. N = 3. (C) DPI treatment abolishes oxidative burst response in CEABAC PMNs infected with Opa+ N. gonorrhoeae. WT and CEABAC PMNs were left untreated or pre-incubated with DPI (10 µM) for 30 min prior to infection with either Opa− or Opa+ N. gonorrhoeae (MOI 10). Oxidative burst was measured with as described in Materials and Methods. (D) Inhibition of Erk1/2 kinases does not affect inflammatory cytokine production. Erk1/2 inhibition does not affect cytokine production by CEABAC PMNs. WT and CEABAC PMNs were left untreated or pre-incubated with Erk1/2 inhibitor (UT0126, 10 µM) for 30 min prior to infection with Opa+ N. gonorrhoeae (MOI 10). MIP-1α production was measured 3 h post infection, N = 3. (EPS)