Anti-fibrosis potential of pirarubicin via inducing apoptotic and autophagic cell death in rabbit conjunctiva.
This study investigated the potential efficacy of pirarubicin (THP) in modulating rabbit conjunctival fibrosis both in vitro and in vivo and characterized the underlying mechanisms. Primary rabbit conjunctival fibroblasts (RCF) were cultured and treated with THP or mitomycin C (MMC) for 5 min, followed by assaying for cell viability, cell cycle distribution, apoptotic and autophagic pathways. The production of reactive oxygen species (ROS) and chemotaxis of macrophages by RCF were evaluated using 2',7'-dichlorofluorescein diacetate (DCFH-DA) labeling and transwell migration assay, respectively. Limbal stem cell excision in combination with alkali burn was performed on the rabbits to establish a model of limbal deficiency and conjunctival fibro-vascular invasion. After three months, the modeled fibro-vascular tissue was excised combined with topical subconjunctival 5-min exposure to THP compared with MMC intraoperatively. The recurrence of postoperative fibrosis and the expression of apoptosis, autophagy, and inflammation markers were evaluated by immunohistochemistry. All modeled rabbits developed conjunctival fibro-vascular lesions, which were similar to human recurrent pterygium (HRP). Both THP and MMC inhibited RCF proliferation and arrested cell cycle at the G0/G1 phase. In particular, 7.5 μmol/L THP remarkably promoted RCF autophagy by upregulating the levels of Beclin 1, Atg 5/12 conjugate, and LC3B, whereas, 15 μmol/L THP significantly triggered a cascade of mitochondrial-associated RCF apoptosis. THP induced the production of ROS and enhanced the chemoattraction of macrophages by RCF. Similar to 600 μmol/L MMC, both 7.5 μmol/L and 15 μmol/L THP attenuated postoperative conjunctival fibrosis in the models; 7.5 μmol/L THP preferentially enhanced autophagy while causing fewer side effects. THP exerted its antifibrotic action by modulating autophagy in RCF, inducing cell cycle arrest, and mitochondrial-mediated apoptosis. THP at the dose of 7.5 μmol/L prevented postoperative conjunctival fibrosis in an animal model.
['Xu|Li-Juan|LJ|', 'Rong|Shi-Song|SS|', 'Xu|Ye-Sheng|YS|', 'Zheng|Li-Bin|LB|', 'Qiu|Wen-Ya|WY|', 'Zhang|Xia|X|', 'Jiang|Lou-Jing|LJ|', 'Duan|Run-Ping|RP|', 'Tian|Tian|T|', 'Yao|Yu-Feng|YF|']
D000818:Animals D017209:Apoptosis D000080549:Autophagic Cell Death D002470:Cell Survival D004195:Disease Models, Animal D004317:Doxorubicin D005347:Fibroblasts D005355:Fibrosis D006801:Humans D011625:Pterygium D011817:Rabbits D017382:Reactive Oxygen Species
Pirarubicin Fibrosis Fibroblasts Apoptosis Autophagy