Pirfenidone potential for treating corneal fibrosis

Abstract

Purpose: Pirfenidone is clinically approved in Europe and Asia for the treatment of idiopathic pulmonary fibrosis. Pirfenidone is shown to reduce the expression of several profibrogenic cytokines including TGFβ, CTGF etc. implicated in the pathogenesis of corneal fibrosis. We tested the effects of pirfenidone on human corneal fibroblast (HCF) viability, proliferation, migration, and death (apoptosis/necrosis), and corneal myofibroblast inhibition using an in vitro model.

Methods: Human donor corneas were used to generate HCF primary cultures. HCFs were exposed to TGFβ1 (5ng/ml) to produce myofibroblasts/fibrosis for 7-9 days under serum-free conditions. Cultures were treated with pirfenidone (0-300µg/ml) in +/- of TGFβ1 to study its effects on HCF function and myofibroblast inhibition. Real-time PCR, western blotting, immunofluorescence, TUNEL, scratch, trypan blue and MTT assays were used to study pirfenidone effects on HCF viability, migration, proliferation and differentiation to myofibroblasts.

Results: Pirfenidone 200µg/ml or less demonstrated no significant loss of HCF viability at 12h, 24h and 48h. Pirfenidone 200µg/ml showed remarkable decrease in HCF proliferation (7-22%), migration (47-73%), and differentiation (up to 93%) without significant toxicity/apoptosis. Further, this Pirfenidone dose significantly reduced myofibroblast formation measured by alpha smooth muscle actin mRNA (84-93%; p<0.001) and protein (68-76%; p<0.01) levels.

Conclusions: Pirfenidone has the potential to treat corneal scarring effectively without compromising normal HCF function. In vivo studies are warranted.

Keywords: 480 cornea: basic science • 484 cornea: stroma and keratocytes • 765 wound healing