Carrigan, Svetlana O.2014-10-2120072007AAINR27175http://hdl.handle.net/10222/54879Neutrophil migration into and across epithelium is indicative of ongoing intestinal inflammation. Moreover, transmigration across the intestinal epithelium appears to be a critical step for neutrophil activation and tissue damage. Using normal human neutrophils migrating across inverted colonic epithelial monolayers we previously discovered that a percentage of neutrophils do not require the engagement of beta2 integrin (CD11b/CD18, Mac-1) when migrating in response to the chemoattractants C5a, CXCL8 and LTB 4. In contrast, neutrophil transepithelial migration in response to the chemoattractant n-formyl methionyl leucyl phenylalanine (fMLP) was Mac-1-dependent.In the present work we describe a modification using promyelocytic cell line HL-60 differentiated with dibutyryl cAMP (dbcAMP) which is devoid of Mac-1. This allowed us to establish that neutrophils do not require engagement of Mac-1 to migrate across intestinal epithelium in response to C5a. Attempting to define the mechanisms that distinguish neutrophil beta2 integrin-dependent migration in response to fMLP versus beta2 integrin-independent migration in response to C5a we tested a number of adhesion molecules using function blocking mAbs and pharmacological inhibitors of intracellular signaling molecules. We found that migration in response to both chemoattractants was dependent on protein kinase C (PKC), Extracellular Regulated Kinases (ERK) and Myosin Light Chain Kinase (MLCK). In contrast, dependence on phospholipase D (PLD), which controls Mac-1 expression, is what distinguished migration in response to fMLP versus C5a. Overall, this work extends the knowledge of mechanisms involved in neutrophil transepithelial migration and might be important when developing anti-inflammatory strategies for the intestinal inflammation.Thesis (Ph.D.)--Dalhousie University (Canada), 2007.Biology, Microbiology.text