Figure 4. Effect of Pla treatment on inflammatory parameters and survival rates during severe sepsis induced by CLP.

WT C57BL/6J mice (n = 4–8) were subjected to severe (18G needle) CLP and then treated with Pla (10 μg/mouse i.p.) 3 hours later. Cells present in the peritoneal cavity were harvested 12 hours after CLP. (A and C) The number of total cells, mononuclear cells, and neutrophils (A), and frequency of efferocytosis (C) were evaluated by counting cytospin slides treated with May–Grünwald–Giemsa stain. (B) The number of M1 (F4/80^low GR1⁺ CD11b^med) macrophages were determined by flow cytometry. (D and E) The levels of TNF, IL-10, IL-6, and CXCL1 were quantified in cell-free peritoneal lavages (D) and plasma (E), respectively, by ELISA. (F) Platelets counted from blood samples. (G and H)The peritoneal fluid (G) and blood (H) samples were plated in brain–heart infusion medium for the analysis of bacterial load. (I) ALT activity was measured from plasma. Results are shown as the mean ± SEM or median of 4–8 mice per group. The experiments were performed 3 times with similar results. *P < 0.05, **P < 0.01, or ***P < 0.001 when comparing the sham group with the CLP group by 1-way ANOVA with post hoc Newman-Keuls (multiple groups) or unpaired 2-tailed Student’s t test (when comparing 2 groups). P values are indicated in the graphs when comparing vehicle with Pla-treated mice. Outliers were removed from the graphs when detected. In the survival experiments, C57BL/6J mice (n = 7) were subjected to severe (18G needle) CLP and treated with Pla (10 μg/mouse, i.p.), Plg (10 μg/mouse, i.p.), imipenem (IMI; 30 mg/kg, i.p.), or a combination of both (Plg 10 μg/mouse i.p. + IMI 30 mg/k, i.p.) after 3 and 12 hours of sepsis induction. (J) The survival rates were monitored for 6 days. The experiment was performed 2 times with similar results. *P < 0.05 when comparing vehicle-treated mice with Pla-, Plg-, or IMI-treated mice. **P < 0.01 when comparing vehicle-treated mice with Plg- + IMI-treated mice by log-rank test.