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Human corneal epithelial extracellular matrix perlecan serves as a site for Pseudomonas aeruginosa binding
1Department of Anatomy/Cell Biology, Wayne State University School of Medicine, Detroit, Michigan 48201, USA.
Purpose:
Previous data has shown that basement membrane associated perlecan serves as a binding site for Pseudomonas aeruginosa in the wounded mouse cornea. The current study determined whether it also provides a binding site for Pseudomonas aeruginosa in transformed human corneal epithelium.
Methods:
Bacterial adherence to transformed human corneal epithelial cells grown in normal or in media containing various inhibitors of glycosaminoglycan synthesis was tested. Bacterial binding was similarly tested in wild-type and in mutant Chinese hamster ovary cell lines naturally deficient in glycosaminoglycan synthesis. Transformed human corneal epithelial extracellular matrix also was tested before and after treatment with anti-proteoglycan monoclonal antibodies or heparinase III before bacterial inoculation. Scanning electron microscopy was used to quantitate adherent bacteria. Intact transformed human corneal epithelial cells or extracellular matrix, the latter either treated or not treated with heparinase III or chondroitin ABC lyase were stained to localize perlecan.
Results:
Examination of the binding of bacteria to transformed human corneal epithelial cells (normal media vs with inhibitors) and Chinese hamster ovary cell lines suggested that bacterial binding was not associated with the surface of either cell type. In contrast, anti-perlecan antibody, as well as heparinase III decreased the binding of bacteria to corneal extracellular matrix. Fluorescence staining localized perlecan to the extracellular matrix beneath the corneal epithelial cells.
Conclusions:
Perlecan localized to the extracellular matrix but not the apical surface of transformed human corneal epithelial cells, provides a binding site for Pseudomonas aeruginosa.
Insights
Perlecan in the extracellular matrix, not on the cell surface, binds Pseudomonas aeruginosa to human corneal epithelium. This finding is crucial for understanding bacterial adhesion in corneal infections.
Area of Science:
- Ophthalmology
- Microbiology
- Cell Biology
Background:
- Basement membrane perlecan binds Pseudomonas aeruginosa in mouse corneas.
- Understanding bacterial binding in human corneal epithelium is essential.
Purpose of the Study:
- To determine if perlecan in transformed human corneal epithelium serves as a binding site for Pseudomonas aeruginosa.
- To investigate the role of extracellular matrix components in bacterial adhesion.
Main Methods:
- Assessed bacterial adherence to human corneal epithelial cells and Chinese hamster ovary cell lines with varying glycosaminoglycan synthesis.
- Tested bacterial binding to extracellular matrix treated with anti-perlecan antibodies or heparinase III.
- Quantified adherent bacteria using scanning electron microscopy and localized perlecan via fluorescence staining.
Main Results:
- Bacterial binding was not observed on the surface of transformed human corneal epithelial cells or Chinese hamster ovary cells.
- Anti-perlecan antibodies and heparinase III significantly reduced bacterial binding to the corneal extracellular matrix.
- Perlecan was localized to the extracellular matrix underlying the corneal epithelial cells.
Conclusions:
- Perlecan in the extracellular matrix, not on the apical surface of transformed human corneal epithelial cells, acts as a binding site for Pseudomonas aeruginosa.
- These findings highlight the importance of the extracellular matrix in Pseudomonas aeruginosa corneal infections.