The Combined CXCR1/CXCR2 Antagonist IP10 Release Inflammation In C57BL/6 Mouse Infected By Klebsiella Pneumoniae

Objective: During the process of acute inflammation, the leucocytes permeate the blood vessels to the infected location. Neutrophil responses are a critical element in host defense during bacterial infections, but neutrophil responses can also be overtly pathogenic. Thus, during many inflammatory diseases, neutrophils contribute more to the pathology than do the microbes themselves. A proper regulation to inflammation can relieve the local damage of tissue. Neutrophil recruitment is a complex process involving activation of local structural cells (e.g., epithelial cells, macrophages), such that the structural cells express inflammatory mediators (e.g., IL-1, CXCL8). In the early phase of inflammation, the neutorphils recruitment to the infected sites and play phagocytosis by the action of ELR+CXC chemokines. iP10 is an antagonist of ELR+CXC chemokine receptor, which inhibited CXCL8-induced chemotactic responses and responses of neutrophils to CXCR1/2-exclusive ligands. iP10 can relief the local damage caused by inflammation for it blocks the chemotaxis induced by ELR+CXC chemokines. Kp is a common opportunistic pathogen in hospitals, and with the increase in resistant strains, Kp has become a difficult miscellaneous. In this paper, we will research on the effect of iP10 to pneumonia induced by Kp of C57BL/6 mice. Methods: Pathogenic strains: Klebsiella pneumoniae, sputum culture from clinial patients. The bateria is cultured on the plate of LB and choose single colony. According to the relation between the growth of bacteria and OD values, the bacteria is most active when it has been cultured for 7h to 11h. Animal model: Diluted a single Klebsiella pneumoniae colony which is most active with sterile saline and adjust the concentration at 107 cfu and inject 20μl into the mouse trachea to induce pneumonia. Animal treatment: Forty C57BL/6 mice were divided into 5 groups randomly, the saline control group, iP10 treated group, DXM treated group, Ceftazidime treated group and bacterial challenged group. Mice were anesthetized by pentobarbital. Briefly, for challenge the animals (n=8) were given a 20μl bolus of Kp (107 cfu) in sterile saline into the trachea. At 30 min before challenge, 500mg/kg iP10, 0.2g/kg ceftazidime, 0.8mg/kg dexamethasone or an equivalent volume of saline was given s.c., and at 24h postchallenge the animals were euthanized with halothane. BALF was collected using 0.6ml vol of sterile saline. BAL total WBC were enumerated by direct counting, and the results were expressed as the mean number of cells per BAL sample (±SEM). BAL cell differentials were determined using Wright's solution-stained BAL cell cytospin preparations, and the total numbers of neutrophils were calculated using these data. All of the data are expressed as means(±SEM). All BAL fluids were assessed independently immediately for myeloperoxidase (MPO). This experiment was repeated three times. Lung was removed to use for MPO, HE stain, RT-PCR(e.g. IL-1β, IL-6, IL-10, TNF-α, IFN-γ).Results: Compared among 5 groups, treatment of mice with iP10 displayed lower account of the neutrophils in BALF in response to other treated groups mice. Pathology and MPO analysis showed that the extent of inflammation and cellular infiltration in the lungs were reduced after treatment with iP10. iP10 interrupted IL-1β, IL-6, IL-10, TNF-α, IFN-γto express in lung tissue. The level of IL-1β, IL-6, IL-10, TNF-α, IFN-γexpression were inhibited by iP10.