Defining the mechanisms of neutrophil transintestinal epithelial migration: Identifying a role for beta2 integrin.
Date
2007
Authors
Carrigan, Svetlana O.
Journal Title
Journal ISSN
Volume Title
Publisher
Dalhousie University
Abstract
Description
Neutrophil 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.
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.
Keywords
Biology, Microbiology.