Purchase this article with an account.
D. K. Augustin, S. R. Heimer, C. Tam, J. LeDue, D. J. Evans, S. M. J. Fleiszig; The Role of mBD-3 in Protecting the Intact Mouse Ocular Surface Against Colonization by Pseudomonas aeruginosa. Invest. Ophthalmol. Vis. Sci. 2010;51(13):6335.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Contact lens wear can predispose to P. aeruginosa keratitis, particularly during extended wear. We previously showed that 3 days of contact lens wear in vitro suppresses up-regulation of human -defensin-2 (hBD-2) by human corneal epithelial cells in response to P. aeruginosa antigens. Here, the role of mBD-3 (a murine homologue of hBD-2) in protecting the ocular surface against P. aeruginosa colonization was explored.
Age-matched wild-type and mBD-3 (-/-) C57BL/6 mice (6-7 weeks old) were anesthetized and 109 CFU of invasive P. aeruginosa strain PAO1 added to the intact ocular surface (i.e. corneas were not scarified). At 3 h or 6 h post-inoculation, viable bacteria remaining in the tear fluid were collected (wash with 5 µl PBS). In other experiments, corneas of mBD-3 (-/-) mice were compared to wild-type for susceptibility to bacterial binding/penetration. To ensure that the effective bacterial inocula were equivalent for mutant and wild-type, these experiments were done ex vivo. Enucleated eyes were washed twice with PBS, challenged with 109 CFU of GFP-expressing P. aeruginosa for 4.5 h or 7.5 h and bacterial distribution in the corneal epithelium quantified using confocal and 2-photon microscopy (enables bacteria and epithelial cells to be localized within unprocessed/unlabeled intact eyes). Animal experiments were conducted in compliance with ARVO policy, and with approval of the UC Berkeley ACUC.
The tear fluid of mBD-3 (-/-) mice contained significantly more bacteria at both 3 and 6 h post-inoculation as compared to wild-type (2.5 log and 1.5 log respectively (p < 0.03 in both cases, Mann-Whitney). Corneas of mBD-3 (-/-) mice were also more susceptible to colonization, however, colonizing bacteria did not traverse the full thickness of the corneal epithelium.
While mBD-3 protects both the tear fluid and corneal epithelium against colonization by invasive P. aeruginosa, it is not the only factor that prevents bacteria from penetrating to the basal lamina, which is the next barrier to bacteria. Defining the mechanisms that protect the corneal epithelium against bacterial traversal could lead to new strategies for preventing infection, and would provide a foundation for further research aimed at understanding corneal susceptibility to infection during contact lens wear.
This PDF is available to Subscribers Only