Fig 1.
Yersinia pestis type III injectisome activates IL-1β and IL-18 cytokines early during lung infection.
A. IL-1β Western blot analysis performed on total lung homogenate harvested at 6, 12, and 24 hpi with LD100 infection of Y. pestis strain CO92, compared to uninfected mice. B. IL-1β ELISA performed on total lung homogenates harvested at 6 and 24 hpi. C. Bacterial burdens from lungs of mice 24 hpi with 104 CFU/ mouse with the strains mentioned (denoted by T3SS) or parent stain (unlabeled). D. IL-1β ELISA performed on total lung homogenates harvested at 24 hpi with with the strains mentioned. All infections were performed in triplicate, with representative analysis shown. * p< 0.05, ** p<0.001.
Fig 2.
IL-1β/IL-18 cytokine activation contributes to pathology of pneumonic plague.
A. A survival analysis was performed comparing wild-type (n = 8), Caspase 1-/- (n = 15) and Il-1β/IL-18-/-(n = 12) mice (on C57BL/6 background) after LD100 infection (104 CFU/mouse) of Y. pestis strain CO92. B. Total lung burden (CFU/lung) at 48 hpi after LD100 infection with Y. pestis strain CO92. C. fixed H&E stained lung sections from mice at 48 hpi. D. Inflammation from histopathology sections was quantified as described in Methods. * p<0.05, ** p<0.001.
Fig 3.
TLR4- and NLRC4-dependent IL-1β and IL-18 activation by Y. pestis contributes to respiratory pathology.
A. IL-1β ELISA performed on lung homogenates obtained from wild-type and TLR4-/-mice, 24 hpi with LD100 dose of Y. pestis. B. Inflammation from histopathology sections from wild-type and TLR4-/- mice 48 hpi with Y. pestis was quantified as described in Methods. C. IL-1β ELISA performed on lung homogenates of wild-type, Caspase 1-/-, NLRP3-/- and NLRC4-/- mice, 24 hpi with LD100 dose of Y. pestis. D. Inflammation from histopathology sections from wild-type, NLRP3-/- and NLRC4-/- mice 48 hpi with Y. pestis. (N = 5–8 mice/group). E. Histology of fixed H&E stained lung sections from mice at 48 hpi. F. Bacterial burdens from lungs of age-matched female wild-type, NLRP3-/-, and NLRC4-/- mice infected intranasally with LD100 dose of Y. pestis strain CO92 for 48 hrs. * p<0.05, ** p<0.001.
Fig 4.
IL-1RA is induced in the lung early during pulmonary infection with Y. pestis, but not K. pneumoniae.
Real time qRT-PCR analysis for (A) IL-1RA and (B) IL-18BP activation from lungs of mice inoculated with either an LD100 dose of Y. pestis (1x104 CFU) or K. pneumoniae (2x105 CFU), at 12, 24 and 36 hpi (N = 3, per experiment). Fold difference calculated as ΔΔ Ct.
Fig 5.
IL-1RA contributes to control of Y. pestis replication and pathology in the lung.
A. Histology of fixed H&E stained lung sections from mice at 24 hpi with Y. pestis, +/- IL-1RA nAb. Small foci of inflammation indicated by arrows. B. Inflammation from histopathology sections was quantified as described in Methods. C. Bacterial burdens were assessed from mice 24 hpi with Y. pestis, +/- IL-1RA nAb. D. Cytokine analysis from lysates of homogenized lungs of mice infected with Y. pestis +/- IL-1RA nAb for 24 hrs was perfomed by ELISA (BD Pharmingen). * p<0.05, ** p<0.001.