Date of Award

Fall 12-1-2004

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Life Sciences

First Advisor

Mary T. Johnson

Second Advisor

Veena Antony

Third Advisor

Roy W. Geib

Abstract

Asthma is characterized by infiltration and shedding of the bronchial epithelium. The Th2 cytokines IL-4 and IL-13 are involved in the cellular recruitment and infiltration seen in asthma. The effects ofIL-4 and IL-13 on cell-matrix interactions, gap formation, increased monolayer permeability and epithelial shedding are unknown. Experiments described in this report test the hypothesis that bronchial airway epithelial cells (BAEC) express paxillin, a structural focal adhesion protein, and down-regulation of paxillin by Th2 cytokines lead to BAEC hyperpermeability. Furthermore, the hypothesis that the Th2 cytokines IL-4 and IL-13 could down-regulate the adherens junction proteins Pcatenin and E-cadherin in BAEC, and that this decrease in P-catenin and E-cadherin also may play a role in BAEC hyperpermeability was tested. The presence of paxillin in BAEC was shown by confocal microscopy which also demonstrated gap formation in IL- 4 and IL-13-stimulated BAEC confluent monolayers that stained positive for P-catenin and E-cadherin. W estem blot analysis demonstrated that IL-4 and IL-13 stimulation results in down-regulation of paxillin, P-catenin and E-cadherin production. IL-4 and IL- 13 stimulation decreased epithelial cell-matrix attachment as measured by electrical cellsubstrate impedance sensing system. IL-4 and IL-13 stimulation increased BAEC monolayer permeability as measured by Evans blue dye/BSA protein leak assay. These results suggest that Th2 cytokines IL-4 and IL-13 down-regulate paxillin production by BAEC, thereby disrupting the cell-matrix interactions. These results also suggest that IL- 4 and IL-13 down-regulate P-catenin and E-cadherin production by BAEC, induce gap formation and promote monolayer hyperpermeability. In conclusion, the Th2 cytokines IL-4 and IL-13 appear to disrupt cell-cell and cell-matrix interactions within the bronchial epithelium. This may help explain the epithelial shedding and epithelial membrane hyperpermeability that occurs in asthma.

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