Abstract
Purpose: :
In addition to antibiotic effects, azithromycin (AZM) has been noted to have anti-inflammatory effects, particularly in the context of bacterial infections. The goal of this study was to explore the suppressive effects of AZM on pro-inflammatory mediators stimulated by toll like receptor (TLR) ligands, in human corneal epithelial cells.
Methods: :
Primary human corneal epithelial cells were cultured from donor corneal limbal explants and grown to sub-confluence in SHEM. The cells were treated with extracted or synthetic microbial components, ligands for TLRs 2-6, respectively for 6-48 hours, with or without pre-incubation with azithromycin (1-50 µg/ml) or TLR antibodies. The cells were subjected to total RNA extraction, reverse transcription (RT) and real-time PCR using Taqman gene expression assays (Applied Biosystems). The medium supernatants of cells treated for 48 hours were collected for protein quantitation using Luminex immunobead assays.
Results: :
The expression of pro-inflammatory cytokines (TNF- and IL-1β), chemokines (IL-8, RANTES), and matrix metalloproteinases (MMP-1 and -9) by human corneal epithelial cells was dramatically stimulated by zymosan, poly I:C (dsRNA) and flagellin, ligands for TLRs 2, 3 and 5 respectively, with peak stimulation at 16 hours. AZM, added 1 hour pre-stimulation, suppressed expression of these pro-inflammatory mediators stimulated by zymosan, but not by flagellin or poly I:C. TLR2 antibody or AZM each partially blocked the expression of TNF-, IL-1β, IL-8, RANTES, MMP-1 and MMP-9 induced by zymosan in human corneal epithelial cells.
Conclusions: :
These findings indicate that AZM has a potential to suppress inflammatory responses stimulated by zymosan through a TLR2 pathway in human corneal epithelial cells.
Keywords: cornea: epithelium • inflammation • antibiotics/antifungals/antiparasitics