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Author

Mehri Zareie

Date of Award

2000

Degree Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

Department

Medical Sciences

Supervisor

Mary H. Perdue

Abstract

Crohn's disease is characterized by altered epithelial physiology including ion secretion and disrupted epithelial barrier. It has been suggested that the pathophysiology and epithelial damage in Crohn's disease may be due to inappropriate immune reactions. The purpose of my studies was to examine the role of cells of the monocyte/macrophage lineage in mediating epithelial physiology. Confluent monolayers of the T84 colonic epithelial cell line were co-cultured with human monocyte/macrophages (peripheral blood monocytes [PBM], or lamina propria mononuclear cells [LPMC]) from controls or patients with crohn's disease ± the activating agents, bacterial lipopolysaccharide (LPS) and/or the bacterial tripeptide, f-met-leu-phe (fMLP). Monolayers were then removed and epithelial secretory (baseline short circuit current [Isc] and ΔIsc to 10-5 forskolin) and barrier transepithelial resistance [TER]) were measured 48h later in the Ussing chambers. Epithelial functional parameters were unchanged after co-culture with non-activated PBM. Addition of activating agent (LPS or LPS/fMLP) significantly increased Isc and reduced TER with PBM from both control and Crohn's disease. No pathophysiology occurred after co-culture with control LPMC ± LPS. However, non-activated LPMC from patients with Crohn's disease spontaneously induced changes in T84 physiological parameters similar to those produced by activated PBM. A non-pathogenic control strain of E. coli , added to the luminal compartment of the culture well, induced comparable changes in epithelial physiology in the presence of PBM. The role of Tumor Necrosis Factor α (TNFα) in the regulation of epithelial secretory and barrier functions was examined. TNFα production was significantly increased by activated PBM and non-activated LPMC from Crohn's disease. The epithelial abnormalities could be successfully abrogated by inclusion of a neutralizing antibody to TNFα. These studies show that monocyte/macrophages are potent immune cells which may mediate some of the pathophysiology in inflammatory disorders of the gut. TNFα is a key factor mediating the monocyte/macrophage induced epithelial pathophysiology.

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