Therapeutic efficacy of CXCR3 blockade in an experimental model of severe sepsis

Abstract

Introduction: In our previous studies we demonstrated that CXC chemokine receptor 3 (CXCR3) participates in the regulation of lymphocyte trafficking during cecal ligation and puncture (CLP)-induced sepsis. In this study, we evaluated the effects of treatment with anti-CXCR3 immunoglobulin (IgG) and antibiotics on outcome during septic shock caused by CLP.

Methods: C57BL/6J mice were treated with neutralizing IgG against CXCR3 plus Primaxin either 24 hours prior to, 2 hours after or 6 hours after CLP. Control mice received nonspecific IgG plus Primaxin in the same regimen. Survival, core body temperature, bacterial clearance and systemic cytokine production were evaluated.

Results: Our results show that treatment with anti-CXCR3 IgG plus Primaxin significantly improved survival when administered 24 hours prior to CLP (50% vs. 10%), 2 hours after CLP (55% vs. 10%) or 6 hours after CLP (55% vs. 25%) compared with mice receiving nonspecific IgG plus Primaxin. Treatment with anti-CXCR3 plus Primaxin 24 hours prior to CLP attenuated hypothermia and IL-6 and macrophage inflammatory protein 2 (MIP-2) production but did not alter bacterial clearance. Treatment with anti-CXCR3 IgG and Primaxin 2 hours after CLP did not improve bacterial clearance and systemic cytokine production compared with mice treated with IgG and Primaxin, whereas 6 hours after CLP the bacterial clearance and IL-6 and MIP-2 concentrations, both in plasma and peritoneal lavage fluid, were significantly improved in mice receiving anti-CXCR3 IgG and Primaxin compared with mice that only received nonspecific IgG and Primaxin.

Conclusion: The results from this study indicate that neutralization of CXCR3 prior to, 2 hours after or 6 hours after the initiation of CLP-induced septic shock improves survival and attenuates CLP-induced inflammation and physiologic dysfunction.

Figure 1

Treatment with anti-CXCR IgG improves survival during cecal ligation and puncture-induced sepsis. Mice were treated with anti-CXC chemokine receptor 3 (anti-CXCR3) IgG (100 μg) or nonspecific IgG (100 μg) 24 hours prior to (n = 10 mice per group), 2 hours after (n = 20 mice per group) or 6 hours after (n = 20 mice per group) cecal ligation and puncture (CLP). Mice were followed for survival over a 2-week period. *P <0.05 compared with nonspecific IgG.

Figure 2

Effect of anti-CXCR3 IgG treatment on core temperature. Mice were treated with anti-CXC chemokine receptor 3 (anti-CXCR3) IgG (100 μg) or nonspecific IgG (100 μg) 24 hours prior to (n = 10 mice per group), 2 hours after (n = 10 mice per group) or 6 hours after (n = 13 mice per group) cecal ligation and puncture (CLP). Rectal temperature was measured 18 hours after CLP. Mice designated as control represent nonseptic mice. *P <0.05 compared with nonspecific IgG, ⁺P <0.05 compared with control.

Figure 3

Effect of anti-CXCR3 IgG treatment on bacterial burden. Mice were treated with anti-CXC chemokine receptor 3 (anti-CXCR3) IgG (100 μg) or nonspecific IgG (100 μg) 24 hours prior to, 2 hours after or 6 hours after cecal ligation and puncture (CLP). Blood and peritoneal lavage fluid were collected 18 hours after CLP and bacterial colony-forming units (CFU) were measured. Mice designated as control represent septic mice that did not receive antibiotic treatment. n = 10 to 13 mice per group. *P <0.05 compared with nonspecific IgG, ⁺P <0.05 compared with control.

Figure 4

Effect of anti-CXCR3 IgG treatment on cytokine production. Mice were treated with anti-CXC chemokine receptor 3 (anti-CXCR3) IgG (100 µg) or nonspecific IgG (100 µg) 24 hours prior to (n = 10 mice per group), 2 hours after (n = 10 mice per group) or 6 hours after (n = 13 mice per group) cecal ligation and puncture (CLP). Plasma and peritoneal lavage fluid were harvested 18 hours after CLP for measurement of IL-6 and macrophage inflammatory protein 2 (MIP-2) concentrations by ELISA. *P <0.05 compared with nonspecific IgG.