Antifibrotic effect of Pirfenidone on orbital fibroblasts of patients with thyroid-associated ophthalmopathy by decreasing TIMP-1 and collagen levels.
OBJECTIVE
The aim of this study was to determine the antifibrotic effects of pirfenidone in orbital fibroblasts of patients with thyroid-associated ophthalmopathy (TAO).


METHODS
The effects of interleukin (IL)-1beta and of fibroblast growth factor (FGF), platelet-derived growth factor (PDGF), and transforming growth factor (TGF)-beta on the induction of tissue inhibitors of metalloproteinases (TIMP)-1 were assessed in orbital fibroblasts of TAO patients. TIMP-1 protein levels were measured by ELISA and Western blot analyses, and TIMP-1 activity was assessed by reverse zymography. The effect of pirfenidone on TIMP-1 induction in orbital fibroblasts was evaluated with the same methods using dexamethasone as a reference agent. A hydroxyproline assay was used to determine the effect of pirfenidone and dexamethasone on collagen production in orbital fibroblasts, and the tetrazolium-based MTT assay was used to assess pirfenidone cytotoxicity.


RESULTS
IL-1beta strongly and dose dependently increased the level of active TIMP-1 protein, whereas FGF, PDGF, and TGF-beta did not significantly induce TIMP-1 protein. Pirfenidone was more effective than dexamethasone in inhibiting IL-1beta-induced increases in TIMP-1, reducing TIMP-1 levels to less than those in untreated controls at a minimal concentration (5 mM). Moreover, pirfenidone effectively decreased hydroxyproline levels in orbital fibroblasts, whereas dexamethasone had no effect on hydroxyproline levels. Pirfenidone exhibited no toxicity in orbital fibroblasts at the concentrations used.


CONCLUSIONS
These results indicate that nontoxic concentrations of pirfenidone have significant antifibrotic effects on orbital fibroblasts from patients with TAO.
Kim
Hyunmi
H
Department of Pharmacology, Ajou University School of Medicine, Suwon, Korea.
Choi
Youn-Hee
YH
Park
Soo Jung
SJ
Lee
Sang Yeul
SY
Kim
Sung Joo
SJ
Jou
Ilo
I
Kook
Koung Hoon
KH
D000328:Adult; D000368:Aged; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D015153:Blotting, Western; D018929:Cell Culture Techniques; D002470:Cell Survival; D003094:Collagen / Q000378:metabolism*; D003907:Dexamethasone / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism; D049970:Graves Ophthalmopathy / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D006801:Humans; D006909:Hydroxyproline / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology; D053583:Interleukin-1beta / Q000494:pharmacology; D008297:Male; D008875:Middle Aged; D009915:Orbit / Q000187:drug effects* / Q000378:metabolism; D011728:Pyridones / Q000494:pharmacology*; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors; D055815:Young Adult
D000894:Anti-Inflammatory Agents, Non-Steroidal; D036341:Intercellular Signaling Peptides and Proteins; D053583:Interleukin-1beta; D011728:Pyridones; D019715:Tissue Inhibitor of Metalloproteinase-1; D014409:Tumor Necrosis Factor-alpha; D003907:Dexamethasone; D003094:Collagen; C093844:pirfenidone; D006909:Hydroxyproline