Anti-fibrotic action of pirfenidone in Dupuytren’s disease-derived fibroblasts

Abstract

BackgroundDupuytren’s disease (DD) is a complex fibro-proliferative disorder of the hand that is often progressive and eventually can cause contractures of the affected fingers. Transforming growth factor beta (TGF-β1) has been implicated as a key stimulator of myofibroblast activity and fascial contraction in DD. Pirfenidone (PFD) is an active small molecule shown to inhibit TGF-β1-mediated action in other fibrotic disorders. This study investigates the efficacy of PFD in vitro in inhibiting TGF-β1-mediated cellular functions leading to Dupuytren’s fibrosis.MethodsFibroblasts harvested from (DD) and carpal tunnel (CT)- tissues were treated with or without TGF-β1 and/or PFD and were subjected to cell migration, cell proliferation and cell contraction assays. ELISA; western blots and real time RT-PCR assays were performed to determine the levels of fibronectin; p-Smad2/Smad3; alpha-smooth muscle actin (α-SMA), α2 chain of type I collagen and α1 chain of type III collagen respectively.ResultsOur results show that PFD effectively inhibits TGF-β1-induced cell migration, proliferation and cell contractile properties of both CT- and DD-derived fibroblasts. TGF-β1−induced α-SMA mRNA and protein levels were inhibited at the higher concentration of PFD (800 μg/ml). Interestingly, TGF-β1 induction of type I and type III collagens and fibronectin was inhibited by PFD in both CT- and DD- derived fibroblasts, but the effect was more prominent in DD cells. PFD down-regulated TGF-β1-induced phosphorylation of Smad2/Smad3, a key factor in the TGF-β1 signaling pathway.ConclusionTaken together these results suggest the PFD can potentially prevent TGF-β1−induced fibroblast to myofibroblast transformation and inhibit ECM production mainly Type I- and Type III- collagen and fibronectin in DD-derived fibroblasts. Further in-vivo studies with PFD may lead to a novel therapeutic application in preventing the progression or recurrence of Dupuytren’s disease.

Pirfenidone reduces profibrotic responses in human dermal myofibroblasts, in vitro

“…Our data clearly showed an upregulation of phosphorylated SMAD2/3 in response to TGF-β1 stimulation, although PFD did not affect this response at any dose or time tested (0.1, 0.5, or 1.0 mg/mL from 5 to 60 min after TGF-β1 stimulation). This contrasts previous findings of PFD inhibition of SMAD phosphorylation in TGF-β1-stimulated human lung fibroblasts and Dupuytren's disease-derived fibroblasts [27,52]. This difference in PFD action may be due to differences in fibroblasts sourced from different tissues, disease states, and in vitro culture systems [56][57][58].…”

“…While the exact mechanism(s) of PFD remain unknown, previous studies demonstrated its effectiveness at mitigating myofibroblast differentiation, proliferation, and profibrotic cytokine production in both in vitro and in vivo models of lung, liver, and renal fibrosis [22,23]. The potential of PFD as a topical therapeutic agent for use during wound healing [24,25] or following HTS development [26] has been suggested, however the mechanism by which PFD exerts its antifibrotic properties on dermal cells has not been fully described [24][25][26][27][28].…”

A Review of Pirfenidone as an Anti-Fibrotic in Idiopathic Pulmonary Fibrosis and Its Probable Role in Other Diseases

“…In a study done on PFD in malignant mesothelioma, it was shown that PFD reduced the TGF-β-induced extracellular-signal-regulated kinase (ERK) and serine/threonine-specific protein kinase (AKT) pathways [ 19 ]. Not only did PFD reduce the proliferation of cells in mesothelioma, but also it reduced the proliferation of fibroblasts derived from Dupuytren's disease in another study done in vitro [ 19 , 20 ]. The Smad2 phosphorylation induced by TGF-β1 was down-regulated in Dupuytren's disease derived-fibroblasts (DD-fibroblasts); hence PFD acted through the canonical Smad signaling pathway at least partially [ 20 ].…”

“…Not only did PFD reduce the proliferation of cells in mesothelioma, but also it reduced the proliferation of fibroblasts derived from Dupuytren's disease in another study done in vitro [ 19 , 20 ]. The Smad2 phosphorylation induced by TGF-β1 was down-regulated in Dupuytren's disease derived-fibroblasts (DD-fibroblasts); hence PFD acted through the canonical Smad signaling pathway at least partially [ 20 ]. While in the study of malignant mesothelioma, the canonical Smad pathway was not affected [ 19 ].…”

Prevention of Recurrences in Dupuytren’s Contracture: Are We in the Right Side?

“…It has already been described that therapies preventing the contraction of myofibroblasts could prevent the contraction and subsequent continuous remodeling of ECM [31]. Pirfenidone [32] acts at this level, inhibiting the functions mediated by TGFß-1 in the cells. 5-fluorouracil has also been tested in DC at this level, reducing the fibroblast and myofibroblast differentiation ratios and their contractility [33].…”