Flow Cytometric Analysis of Inflammatory Cells in Experimental Acute Pancreatitis
Archives of Medical Laboratory Sciences,
Vol. 1 No. 3 (2015),
20 Azar 2015
https://doi.org/10.22037/amls.v1i3.10857
Abstract
Background: Accumulating evidence indicates that inflammatory cells migrate into the pancreas tissue and play an important role in the pathogenesis of acute pancreatitis (AP). The aim of this study was to establish a flow cytometric method to enumerate these infiltrating cells in the pancreas of an experimental AP.
Materials and Methods: Twelve hours after inducing of AP, mice pancreatic tissues were cut into small fragments and single cells were prepared by mechanical dissociation. The isolated cells were stained with either anti-mouse CD45-PerCP or isotype antibody and analyzed by flow cytometry. Using side scatter (SSC)/CD45 gating we were able to identify inflammatory cells from non-inflammatory cells.
Results: The mean percentage of leukocytes was 5.9±1.6 in the control group whereas, it was 26.7±8.1 in the AP. Moreover, we found that the percentage of lymphocytes, monocytes and granulocytes were 1.1±0.2, 0.9±.04 and 2.9±1.8 of total pancreatic cells, respectively, in the control mice. In contrast to lymphocytes, the percentage of monocytes and granulocytes were significantly increased in the AP group and it was 3±1.3 and 18.2±3.2 for monocytes and granulocytes, respectively.
Conclusion: Quantitative flow cytometric analysis is feasible and provides a reliable and rapid assay to determine the number and percentage of inflammatory cells in experimental AP.
- Acute pancreatitis
- flow cytometry
- inflammatory cells
How to Cite
References
Sah RP, Garg P, Saluja AK. Pathogenic mechanisms of acute pancreatitis. Curr Opin Gastroenterol. 2012;28(5):507-15.
Bhatia M, Wong FL, Cao Y, Lau HY, Huang J, Puneet P, et al. Pathophysiology of acute pancreatitis. Pancreatology. 2005;5(2-3):132-44.
Dambrauskas Z, Giese N, Gulbinas A, Giese T, Berberat PO, Pundzius J, et al. Different profiles of cytokine expression during mild and severe acute pancreatitis. World journal of gastroenterology : WJG. 2010;16(15):1845-53.
Mayerle J, Dummer A, Sendler M, Malla SR, van den Brandt C, Teller S, et al. Differential roles of inflammatory cells in pancreatitis. J Gastroenterol Hepatol. 2012;2:47-51.
Panek J, Kusnierz-Cabala B, Dolecki M, Pietron J. Serum proinflammatory cytokine levels and white blood cell differential count in patients with different degrees of severity of acute alcoholic pancreatitis. Pol Przegl Chir. 2012;84(5):230-7.
Ohmoto K, Yamamoto S. Serum interleukin-6 and interleukin-10 in patients with acute pancreatitis: clinical implications. Hepatogastroenterology. 2005;52(64):990-4.
Ramudo L, De Dios I, Yubero S, Vicente S, Manso MA. ICAM-1 and CD11b/CD18 expression during acute pancreatitis induced by bile-pancreatic duct obstruction: effect of N-acetylcysteine. Exp Biol Med. 2007;232(6):737-43.
Abdulla A, Awla D, Thorlacius H, Regner S. Role of neutrophils in the activation of trypsinogen in severe acute pancreatitis. J Leukoc Biol. 2011;90(5):975-82.
Awla D, Abdulla A, Syk I, Jeppsson B, Regner S, Thorlacius H. Neutrophil-derived matrix metalloproteinase-9 is a potent activator of trypsinogen in acinar cells in acute pancreatitis. J Leukoc Biol. 2012;91(5):711-9.
Chooklin S, Pereyaslov A, Bihalskyy I. Pathogenic role of myeloperoxidase in acute pancreatitis. Hepatobiliary Pancreat Dis Int. 2009;8(6):627-31.
Rinderknecht H. Fatal pancreatitis, a consequence of excessive leukocyte stimulation? Int J Pancreatol. 1988;3(2-3):105-12.
Barie PS, Tahamont MV, Malik AB. Prevention of increased pulmonary vascular permeability after pancreatitis by granulocyte depletion in sheep. Am Rev Respir Dis. 1982;126(5):904-8.
Willemer S, Feddersen CO, Karges W, Adler G. Lung injury in acute experimental pancreatitis in rats. I. Morphological studies. Int J Pancreatol. 1991;8(4):305-21.
Aho HJ, Nevalainen TJ. Experimental pancreatitis in the rat. Light and electron microscopical observations on early pancreatic lesions induced by intraductal injection of trypsin, phospholipase A2, lysolecithin and non-ionic detergent. Virchows Arch B Cell Pathol Incl Mol Pathol. 1982;40(3):347-56.
Shrivastava P, Bhatia M. Essential role of monocytes and macrophages in the progression of acute pancreatitis: World J Gastroenterol. 2010 Aug 28;16(32):3995-4002.
Carlson SL, Parrish ME, Springer JE, Doty K, Dossett L. Acute inflammatory response in spinal cord following impact injury. Exp Neurol. 1998;151(1):77-88.
Malleo G, Mazzon E, Genovese T, Di Paola R, Muia C, Crisafulli C, et al. Effects of thalidomide in a mouse model of cerulein-induced acute pancreatitis. Shock. 2008;29(1):89-97.
Campanella M, Sciorati C, Tarozzo G, Beltramo M. Flow cytometric analysis of inflammatory cells in ischemic rat brain. Stroke. 2002;33(2):586-92.
Kerfoot SM, Kubes P. Overlapping roles of P-selectin and alpha4 integrin to recruit leukocytes to the central nervous system in experimental autoimmune encephalomyelitis. J Immunol. 2002;169(2):1000-6.
Tjoa T, Strausbaugh HJ, Maida N, Dazin PF, Rosen SD, Noble-Haeusslein LJ. The use of flow cytometry to assess neutrophil infiltration in the injured murine spinal cord. J Neurosci Methods. 2003;129(1):49-59.
Li K, Feng JY, Li YY, Yuece B, Lin XH, Yu LY, et al. Anti-inflammatory role of cannabidiol and O-1602 in cerulein-induced acute pancreatitis in mice. Pancreas. 2013;42(1):123-9.
Perides G, Weiss ER, Michael ES, Laukkarinen JM, Duffield JS, Steer ML. TNF-alpha-dependent regulation of acute pancreatitis severity by Ly-6C(hi) monocytes in mice. J Biol Chem. 2011;286(15):13327-35.
Ogawa M. Acute pancreatitis and cytokines: "second attack" by septic complication leads to organ failure. Pancreas. 1998;16(3):312-5.
Barone FC, Hillegass LM, Tzimas MN, Schmidt DB, Foley JJ, White RF, et al. Time-related changes in myeloperoxidase activity and leukotriene B4 receptor binding reflect leukocyte influx in cerebral focal stroke. Mol Chem Neuropathol. 1995;24(1):13-30.
Sandoval D, Gukovskaya A, Reavey P, Gukovsky S, Sisk A, Braquet P, et al. The role of neutrophils and platelet-activating factor in mediating experimental pancreatitis. Gastroenterology. 1996;111(4):1081-91.
Schober A, Weber C. Mechanisms of monocyte recruitment in vascular repair after injury. Antioxid Redox Signal. 2005;7(9-10):1249-57.
Geissmann F, Manz MG, Jung S, Sieweke MH, Merad M, Ley K. Development of monocytes, macrophages, and dendritic cells. Science. 2010;327(5966):656-61.
Park P, Haas M, Cunningham PN, Bao L, Alexander JJ, Quigg RJ. Injury in renal ischemia-reperfusion is independent from immunoglobulins and T lymphocytes. Am J Physiol Renal Physiol. 2002;282(2):F352-7.
Azab B, Jaglall N, Atallah JP, Lamet A, Raja-Surya V, Farah B, et al. Neutrophil-lymphocyte ratio as a predictor of adverse outcomes of acute pancreatitis. Pancreatology. 2011;11(4):445-52.
Qin Y, Pinhu L, You Y, Sooranna S, Huang Z, Zhou X, et al. The role of fas expression on the occurrence of immunosuppression in severe acute pancreatitis. Dig Dis Sci. 2013;58(11):3300-7.
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