| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
1From the Department of Cellular Biology and Anatomy, Medical College of Georgia, Augusta, Georgia; and the 2Department of Ophthalmology, University of Michigan, Ann Arbor, Michigan.
PURPOSE. To determine the role of epidermal growth factor (EGF) receptor (EGFR)–mediated signaling pathways in preventing infection-induced apoptosis in human corneal epithelial cells (HCECs).
METHODS. Epithelial monolayers of a telomerase-immortalized HCEC line, HUCL, and primary culture of HCECs were infected with Pseudomonas aeruginosa in the presence of the EGFR inhibitor tyrphostin AG1478, the extracellular signal-regulated kinase (ERK) inhibitor U0126, the phosphoinositide 3-kinase (PI3K) inhibitor LY294002, the heparin-binding EGF-like growth factor (HB-EGF) antagonist CRM197, the HB-EGF neutralizing antibody, or the matrix metalloproteinase inhibitor GM6001. The activation of EGFR was analyzed by immunoprecipitation using EGFR antibodies, followed by Western blot analysis with phosphotyrosine antibody. Phosphorylation of ERK and Akt, a major substrate of PI3K, and generation of cleaved caspase-3 and poly (ADP-ribose) polymerase (PARP) were determined by Western blot analysis. Apoptotic cells were characterized by positive staining of active caspase-3, loss of mitochondrial cytochrome c, and condensation of chromosomes. Apoptosis was also confirmed by measuring caspase-3 activity and assessing the generation of cleaved caspase-3 and PARP.
RESULTS. P. aeruginosa infection of HUCL cells resulted in EGFR activation and EGFR-dependent ERK1/2 and PI3K phosphorylation. Inhibition of EGFR, ERK1/2, and PI3K activities with kinase-specific inhibitors (AG1478, U0126, and LY294002, respectively) resulted in an increase in the number of apoptotic cells, in elevated cellular caspase-3 activity, and/or in increased cleaved PARP in P. aeruginosa–infected HUCL cells or primary culture of HCECs. Blocking HB-EGF ectodomain shedding by inhibition of matrix metalloproteinase–mediated proteolysis, downregulation of HB-EGF, or neutralization of its activity retarded infection-induced EGFR transactivation and, as a consequence, increased infection-induced HUCL apoptosis.
CONCLUSIONS. Bacterial infection of HCECs induces EGFR transactivation through HB-EGF ectodomain shedding. EGFR and its downstream ERK and PI3K signaling pathways play a role in preventing epithelial apoptosis in the early stage of bacterial infection.
This article has been cited by other articles:
|
|
 |
|
  M. A. Oberhardt, J. Puchalka, K. E. Fryer, V. A. P. Martins dos Santos, and J. A. Papin Genome-Scale Metabolic Network Analysis of the Opportunistic Pathogen Pseudomonas aeruginosa PAO1 J. Bacteriol., April 15, 2008; 190(8): 2790 - 2803. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Ashare, M. M. Monick, A. B. Nymon, J. M. Morrison, M. Noble, L. S. Powers, T. O. Yarovinsky, T. L. Yahr, and G. W. Hunninghake Pseudomonas aeruginosa Delays Kupffer Cell Death via Stabilization of the X-Chromosome-Linked Inhibitor of Apoptosis Protein J. Immunol., July 1, 2007; 179(1): 505 - 513. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. L. Roxas, A. Koutsouris, and V. K. Viswanathan Enteropathogenic Escherichia coli-Induced Epidermal Growth Factor Receptor Activation Contributes to Physiological Alterations in Intestinal Epithelial Cells Infect. Immun., May 1, 2007; 75(5): 2316 - 2324. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 K.-P. Xu, J. Yin, and F.-S. X. Yu SRC-family tyrosine kinases in wound- and ligand-induced epidermal growth factor receptor activation in human corneal epithelial cells. Invest. Ophthalmol. Vis. Sci., July 1, 2006; 47(7): 2832 - 2839. [Abstract] [Full Text] [PDF]
|
|
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
