On study day 0, mice are given intraperitoneal injections of zymosan (1 mg) in sterile saline to induce peritonitis.
Disease Parameters:
Inflammation is the response of vascularized tissues to injury, irritation, and infection. Acute inflammation is most often associated with a neutrophil-rich cellular infiltrate and is generally resolved in a period of days, whereas chronic inflammation is characterized by a cellular infiltrate containing many more mononuclear cells (monocyte/macrophages and lymphocytes).1,2,3 The ability to investigate the role of individual inflammatory mediators in acute inflammation, such as complement proteins, prostaglandins, leukotrienes, cytokines, and chemokines, has been aided by the availability of simple models of sterile peritonitis, which allow the accurate quantitation of leukocyte recruitment into an easily accessible serosal cavity.4-10 Recently, sterile peritonitis models have proved useful for the analysis of animals in which specific inflammatory mediators or their receptors have been ablated genetically or targeted by neutralizing antibodies.11,12
Dosing Paradigms:
Begin dosing 30 minutes prior to zymosan injection or just before injection (t = 0) and continue until necropsy at t = 4, 8, 12, 16, or 24 hours.
Route of administration: SC, PO, IP, IV
Clinical Assessment:
The peritoneal cavity is lavaged, the lavage fluid is transferred to a 15 ml tube and the peritoneal exudate cells are pelleted by centrifugation. The pellets are re-suspended and single cell suspensions are prepared by thoroughly washing each pellet. The cell suspensions are spun down in a Cytospin® collection funnel with an attached slide. The slides are dried overnight followed by staining with a Diff-Quick stain. The cell differentiation counts (granulocyte, monocyte/macrophage, lymphocyte, etc.) are quantified under a light microscope. Cell counts are performed by counting squares on a Bright-Line Hemocytometer.
For examples of positive controls, please contact us.
Sample Data (Click on image to enlarge):
Notes:
Experimental peritonitis models have proved to be an indispensable tool for the explanation of acute inflammatory processes, the mechanisms of action of anti-inflammatory drugs, and are increasingly being used to test novel anti-inflammatory therapies, including anti-chemokine and anti-chemokine receptor strategies.13,14
Optional Endpoint
PK/PD blood collections
Cytokine/chemokine analysis via Luminex(R)
Other sandwich ELISAs
CBC/clinical chemistry analysis
Soft tissue collection
Histopathologic analysis
Immunohistochemistry analysis
References:
Medzhitov, R. (2008). Origin and physiological roles of inflammation. Nature 454, 428–435.
Nathan, C. (2002). Points of control in inflammation. Nature 420, 846–852.
Majno, G., and Joris, I. (1996). ‘‘Cells, Tissues, and Disease’’ Blackwell Science, Oxford.
Firestein, G. S. (2003). Evolving concepts of rheumatoid arthritis. Nature 423, 356–361.
Fantuzzi, G., Ku, G., Harding, M. W., Livingston, D. J., Sipe, J. D., Kuida, K., Flavell, R. A., and Dinarello, C. A. (1997). Response to local inflammation of IL-1 beta-converting enzyme- deficient mice. J. Immunol. 158, 1818–1824.
Forrest, M. J., Jose, P. J., and Williams, T. J. (1986). Kinetics of the generation and action of chemical mediators in zymosan-induced inflammation of the rabbit peritoneal cavity. Br. J. Pharmacol. 89, 719–730.
Ajuebor, M. N., Das, A. M., Virag, L., Flower, R. J., Szabo, C., and Perretti, M. (1999a). Role of resident peritoneal macrophages and mast cells in chemokine production and neutrophil migration in acute inflammation: Evidence for an inhibitory loop involving endogenous IL-10. J. Immunol. 162, 1685–1691.
Ajuebor, M. N., Das, A. M., Virag, L., Szabo, C., and Perretti, M. (1999b). Regulation of macrophage inflammatory protein-1 alpha expression and function by endogenous interleukin-10 in a model of acute inflammation. Biochem. Biophys. Res. Commun. 255, 279–282.
Ajuebor, M. N., Flower, R. J., Hannon, R., Christie, M., Bowers, K., Verity, A., and Perretti, M. (1998). Endogenous monocyte chemoattractant protein-1 recruits mono- cytes in the zymosan peritonitis model. J. Leukoc. Biol. 63, 108–116.
Lefkowith, J. B. (1988). Essential fatty acid deficiency inhibits thein vivogeneration of leukotriene B4 and suppresses levels of resident and elicited leukocytes in acute inflammation. J. Immunol. 140, 228–233.
Mack, M., Cihak, J., Simonis, C., Luckow, B., Proudfoot, A. E. I., Bruhl, H., Frink, M., Anders, H.-J., Vielhauer, V., Pfirstinger, J., Stangassinger, M., and Schlondorff, D. 396 Jenna L. Cash et al. (2001). Expression and Characterization of the Chemokine Receptors CCR2 and CCR5 in Mice. J. Immunol. 166, 4697–4704.
Wengner, A. M., Pitchford, S. C., Furze, R. C., and Rankin, S. M. (2008). The coordinated action of G-CSF and ELR þ CXC chemokines in neutrophil mobilization during acute inflammation. Blood 111, 42–49.
Bursill, C. A., Cai, S., Channon, K. M., and Greaves, D. R. (2003). Adenoviral-mediated delivery of a viral chemokine binding protein blocks CC-chemokine activity in vitro andin vivo. Immunobiology 207, 187–196.
Van Wanrooij, E. J. A., de Jager, S. C. A., van Es, T., de Vos, P., Birch, H. L., Owen, D. A., Watson, R. J., Biessen, E. A. L., Chapman, G. A., van Berkel, T. J. C., and Kuiper, J. (2008). CXCR3 Antagonist NBI-74330 attenuates atherosclerotic plaque formation in LDL receptor-deficient mice. Arterioscler. Thromb. Vasc. Biol. 28, 251–257.
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