Abstract
Purpose: :
To explore the role of murine β-defensin (mBD) 1 and 2 in Pseudomonas aeruginosa (P. aeruginosa)-induced corneal infection.
Methods: :
mRNA and protein expression levels of mBD1 and mBD2 in normal and infected corneas of C57BL/6 (B6) and BALB/c mice were tested by real-time RT-PCR and western blot, respectively. The distribution patterns of mBD1 and mBD2 were further determined by immunostaining. For in vivo study, BALB/c mice were treated with either mBD1 or mBD2 siRNA by subconjunctival injection together with topical application. Disease was monitored using slit lamp, bacterial plate counts, MPO assay, ELISA, Griess reaction, and NF-ΚB activation.
Results: :
Both mBD1 and mBD2 were constitutively expressed (mRNA and protein) in normal BALB/c and B6 corneas, and disparately up-regulated in BALB/c (more) vs B6 (less) corneas after infection. Increased corneal opacity and worsened disease were displayed after blocking mBD2, rather than mBD1. mBD2 silencing also increased bacterial counts and neutrophil (PMN) infiltration in BALB/c corneas. RT-PCR data further demonstrated that mBD2, not mBD1, differentially modulated the mRNA expression of several pro-inflammatory cytokines (IFN-γ, MIP-2, IL-1β , TNF-, IL-6), iNOS, and Toll-like receptor (TLR) signaling molecules (TLR2, TLR4, TLR9, MyD88), as well as transcription factor NF-ΚB. In addition, in vivo studies indicated that mBD2 silencing enhanced nitrite levels and NF-ΚB activation.
Conclusions: :
mBD2, rather than mBD1, is required for host resistance against P. aeruginosa-induced corneal infection.
Keywords: keratitis • RNAi • cytokines/chemokines