Immunopathologic alterations in murine models of sepsis of increasing severity - PubMed (original) (raw)

Immunopathologic alterations in murine models of sepsis of increasing severity

S Ebong et al. Infect Immun. 1999 Dec.

Abstract

We investigated inflammatory and physiologic parameters in sepsis models of increasing lethality induced by cecal ligation and puncture (CLP). Mice received imipenem for antibiotic therapy, and groups were sacrificed at 2, 4, 8, 12, 16, 20, and 24 h after CLP. The severity of sepsis increased with needle puncture size (lethality with 18-gauge puncture [18G], 100%; 21G, 50%; 25G, 5%; sham treatment, 0%). While the temperature (at 12 h) and the activity and diurnal rhythm (at day 4) of the 25G-treated CLP group recovered to normal, the 21G and 18G treatment groups exhibited severe hypothermia along with decreased activities. A direct correlation was also observed between the severity of sepsis and cytokine (interleukin 1beta [IL-1beta], tumor necrosis factor [TNF], IL-6, and IL-10) concentrations in both the peritoneum and the plasma. There were substantially higher cytokine levels in the more severe CLP models than in the sham-treated one. Peritoneal and plasma TNF levels were always less than 40 pg/ml in all models. None of the cytokines in the septic mice peaked within the first hour, which is in contrast to the results of most endotoxin models. Chemokine (KC and macrophage inflammatory protein 2) profiles also correlated with the severity of sepsis. Except for the chemokines, levels of inflammatory mediators were always higher at the site of inflammation (peritoneum) than in the circulation. Our study demonstrated that sepsis of increasing severity induced increased cytokine levels both within the local environment (peritoneum) and systemically (plasma), which in turn correlated with morbidity and mortality.

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Figures

FIG. 1

FIG. 1

CLP lethality. Mortality coincided with the severity (needle gauge) of CLP.

FIG. 2

FIG. 2

Temperature profile. The temperature of sham-treated mice (n = 6) returned to normal faster than that of the CLP-treated mice. The 25G-treated mice (n = 19) recovered by day 3, while the 21G-treated mice (n = 18) never attained the temperature of the sham-treated mice. The 18G-treated mice (n = 6) were severely hypothermic and died by 48 h. ∗, P < 0.05. Error bars indicate +SEM.

FIG. 3

FIG. 3

Gross motor activity or diurnal rhythm. (A) Recovery coincided with the severity of insult. (B) The nonlethal treated (25G-treated) mice recovered to the activity level of the sham-treated mice by day 3. (C) The 21G-treated CLP group did not fully recover to the activity level of the sham-treated mice throughout the study period. The bar along the x axis representing the dark-light cycle indicates that mice were maintained in a temperature-controlled room with a 12-h lights-on (solid bar) and 12-h lights-off (open bar) cycle. The SEM may be smaller than the symbol. Each value is the mean ± SEM for 6 to 19 mice.

FIG. 4

FIG. 4

TNF. There were higher TNF levels in the mice of the more severe treatment models. There were no detectable levels of biologically active TNF in the circulation at any time point (data not shown). ∗∗∗, P < 0.05, 18G- versus sham-treated mice (n = 6). The SEM may be smaller than the symbol.

FIG. 5

FIG. 5

IL-1β. (A) All mice that underwent CLP had elevated levels of IL-1β in the peritoneum regardless of puncture size. (B) The 18G-treated CLP model showed increasing circulating levels. Each value is the mean ± SEM for 6 to 10 mice. ∗, ∗∗, and ∗∗∗, P < 0.05 for 25G, 21G, and 18G treatment groups (versus sham treatment group). The SEM may be smaller than the symbol.

FIG. 6

FIG. 6

IL-6. There were elevated levels of IL-6 within the peritoneum (A) and in the circulation (B) (see also Table 1) in the 21G- and 18G-treated CLP models. ∗∗∗, P < 0.05 for 18G- sham-treated groups. The SEM may be smaller than the symbol. Values are expressed as means ± SEM for 6 to 10 mice.

FIG. 7

FIG. 7

IL-10. There were elevated levels of IL-10 in mice of the more severe treatment models locally and systemically. The SEM may be smaller than the symbol. Values are expressed as means ± SEM for 6 to 10 mice.

FIG. 8

FIG. 8

KC. (A) There were elevated KC levels within the peritoneum in the 18G-treated CLP model. (B) All mice that underwent CLP had elevations in the circulation regardless of puncture size. KC levels in plasma are substantially higher than those in the peritoneum. Values are expressed as means ± SEM for 6 to 10 mice. ∗∗, P < 0.05 for 21G versus sham treatment groups; ∗∗∗, P < 0.05 for 18G versus sham treatment groups. The SEM may be smaller than the symbol.

FIG. 9

FIG. 9

MIP-2. Concentrations were higher in mice in the more severe treatment models in both the peritoneum (A) and the plasma (B). Overall, MIP-2 concentrations were lower than KC levels. ∗∗∗,P < 0.05 for 18G versus sham treatment groups (n = 6); ∗∗, P < 0.05 for 21G versus sham treatment groups. The SEM may be smaller than the symbol. Values are expressed as means ± SEM for 6 to 10 mice.

FIG. 10

FIG. 10

Peritoneal cell counts after CLP or sham surgery. There was neutrophil influx into the local site of inflammation in all mice that underwent CLP. Each value is the mean ± SEM for 6 to 10 mice. The SEM may be smaller than the symbol.

FIG. 11

FIG. 11

Relation between CLP treatments and inflammatory scores. There is a clear relationship between mortality and severity of sepsis. Scores were obtained from the individual physiologic responses—lethality, temperature, and activity (A) — and from the cytokines (IL-1β, TNF, IL-6) and chemokines (KC and MIP-2) (B). Each bar represents the mean + SEM. ∗, P < 0.05 versus sham treatment group.

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