인간 섬유아세포에 대한 마이토마이신 C와 피르페니돈의 독성과 항섬유화효과
Cytotoxicities and Anti-Fibrotic Effects of Pirfenidone and Mitomycin C on Human Fibroblasts
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목적: 인간 진피 섬유아세포를 마이토마이신 C, 피르페니돈에 노출시켜 세포독성과 항섬유화 효과를 비교하기 위해 실험하였다. 대상과 방법: 섬유화 유도를 위해 transforming growth factor (TGF)-β1을 처리하였고 여러 농도의 마이토마이신 C, 피르페니돈을 첨가하여 24시간 배양하여 세포독성을 보았다. 마이토마이신 C 4, 20 μg/mL, 피르페니돈 250, 500 μg/mL에서 real time reversetranscription polymerase chain reaction (RT-PCR)과 western blot으로 fibronectin, collagen type I, α smooth muscle, β-actin의발현을 살펴보았다. 결과: TGF-β1만 처리한 군과 비교 시, 마이토마이신 C 4, 20, 40 μg/mL, 피르페니돈 500, 750, 1,000 μg/mL의 농도에서는 통계적으로 유의하게 세포독성이 증가되었다(p＜0.05, Mann Whitney U-test). Real time RT-PCR에서 fibronectin, collagen type I, αsmooth muscle 발현이 TGF-β1만 처리한 군보다 감소하였다. Western blot 실험에서 fibronectin, α smooth muscle 발현이 TGF-β1만 처리한 군보다 감소하였으나 collagen type I은 유의한 변화를 보이지 않았다. 결론: 진피 섬유아세포에 대해 약물 모두 세포 독성과 항섬유화 효과가 나타났는데 피르페니돈은 독성이 적었고, 마이토마이신 C는항섬유화 효과가 크게 나타났다.
Abstract
Purpose: The cytotoxicities and anti-fibrotic effects of mitomycin C and pirfenidone on human dermal fibroblast were evaluated. Methods: Initially, 24-hour cell cultures were exposed to transforming growth factor (TGF)-β1, different concentrations of mitomycinC, and pirfenidone solutions in order to evaluate cytotoxicity. Expressions of fibronectin, collagen type 1, α smooth muscle,and β-actin were evaluated by real-time reverse transcription-polymerase chain reaction (RT-PCR) and western blot in mitomycinC solutions at concentrations of 4 μg/mL and 20 μg/mL, and in pirfenidone solutions at 250 μg/mL and 500 μg/mL. Results: In comparison to cell cultures exposed to TGF-β1 solutions, cytotoxicities were increased in solutions of mitomycin C at 4μg/mL, 20 μg/mL, 40 μg/mL and pirfenidone at 500 μg/mL, 750 μg/mL, 1,000 μg/mL (p < 0.05, Mann Whitney U-test). The results ofreal-time RT-PCR show that expressions of fibronectin, collagen type 1, and α smooth muscle were significantly more decreased inall concentrations of mitomycin C and pirfenidone compared to those in TGF-β1 solution. In western blot analysis, expressions of fibronectinand α smooth muscle were decreased in all concentrations of mitomycin C and pirfenidone compared to TGF-β1 solution. Conclusions: Both drugs have cytotoxicities and anti-fibrotic effects, but pirfenidone was found to have less cytotoxicity and mitomycinC was found to have more anti-fibrotic effects when compared to each other.
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