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Therapeutic potential of P irfenidone for treating equine corneal scarring
Author(s) -
Fink Michael K.,
Giuliano Elizabeth A.,
Tandon Ashish,
Mohan Rajiv R.
Publication year - 2015
Publication title -
veterinary ophthalmology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.594
H-Index - 50
eISSN - 1463-5224
pISSN - 1463-5216
DOI - 10.1111/vop.12194
Subject(s) - trypan blue , pirfenidone , viability assay , in vitro , fetal bovine serum , myofibroblast , cornea , in vivo , fibrosis , andrology , chemistry , pathology , medicine , biology , idiopathic pulmonary fibrosis , ophthalmology , biochemistry , microbiology and biotechnology , lung
Objective To evaluate the safety and efficacy of P irfenidone ( PFD ) in the treatment of equine corneal fibrosis using an in vitro model. Methods Healthy donor equine corneas were collected and used to generate primary equine corneal fibroblasts ( ECF s) by growing cultures in minimal essential medium supplemented with 10% fetal bovine serum. Equine corneal myofibroblasts ( ECM s), used as a model of equine corneal fibrosis, were produced by growing ECF cultures in serum‐free medium containing transforming growth factor β1 (1 ng/mL). Trypan blue viability assays and changes in ECF morphology were utilized to determine the optimal PFD dose for this in vitro model. Trypan blue viability, phase‐contrast microscopy, and TUNEL assays were used to evaluate the cytotoxicity of PFD . Scratch and MTT assays were used to evaluate the effect of PFD on cellular migration and proliferation. Real‐time PCR , immunoblot analysis, and immunocytochemistry were employed to determine the efficacy of PFD to inhibit ECM formation in vitro . Results Topical PFD application at 200 μg/mL successfully decreased α SMA expression when compared to the TGF β1 only treatment group ( P < 0.01). PFD application ≤200 μg/mL did not affect ECF phenotype or cellular viability and did not result in significant cytotoxicity. Conclusions Pirfenidone safely and effectively inhibits TGF β1‐induced equine corneal fibrosis in vitro . In vivo studies are warranted.