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Department of Anatomy and Cell Biology, Wayne State University School of Medicine, Detroit, MI 48201
Corneal infection with Pseudomonas aeruginosa results in corneal perforation in susceptible C57BL/6 (B6) mice, but not in resistant BALB/c mice. To explore the role of two important defensins, murine β-defensin-1 (mBD1) and mBD2, in the ocular immune defense system, their mRNA and protein expression levels were tested by real-time RT-PCR and Western blot, respectively. mRNA, protein, and immunostaining data demonstrated that both mBD1 and mBD2 were constitutively expressed in normal BALB/c and B6 corneas, and they were disparately up-regulated in BALB/c (more) vs B6 (less) corneas after infection. To determine whether either defensin played a role in host resistance, BALB/c mice were treated with either mBD1 or mBD2 small interfering RNA by subconjunctival injection together with topical application. Increased corneal opacity and worsened disease were displayed after knockdown of mBD2 but not of mBD1. mBD2 silencing also increased bacterial counts and polymorphonuclear neutrophil infiltration in BALB/c corneas. Real-time RT-PCR data further demonstrated that mBD2, not mBD1, differentially modulated mRNA expression of proinflammatory cytokines/molecules such as IFN-
, MIP-2, IL-1β, TNF-
, IL-6, and inducible NO synthase; TLR signaling molecules, including TLR2, TLR4, TLR9, and MyD88; and the transcription factor NF-
B. Additionally, in vivo studies indicated that mBD2 silencing enhanced corneal nitrite levels and NF-B activation. Collectively, the data provide evidence that mBD2, but not mBD1, is required for host resistance against P. aeruginosa-induced corneal infection.
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
1 This investigation was supported by Grants R01 EY016058, EY02986, and P30 EY04068 from the National Eye Institute, the National Institutes of Health.
2 Address correspondence and reprint requests to Dr. Linda D. Hazlett, Department of Anatomy and Cell Biology, Wayne State University School of Medicine, 540 E. Canfield Avenue, Detroit, MI 48201. E-mail address: lhazlett{at}med.wayne.edu
3 Abbreviations used in this paper: mBD, murine β-defensin; iNOS, inducible NO synthase; p.i., postinfection; siRNA, small interfering RNA; MPO, myeloperoxidase; PMN, polymorphonuclear neutrophil; DC, dendritic cell.
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