Fraxinellone-mediated targeting of cathepsin B leakage from lysosomes induces ferroptosis in fibroblasts to inhibit hypertrophic scar formation.

IF 5.7 2区 生物学 Q1 BIOLOGY
Wei Xu, Hao Lv, Yaxin Xue, Xiaofeng Shi, Shaotian Fu, Xiaojun Li, Chuandong Wang, Danyang Zhao, Dong Han
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引用次数: 0

Abstract

Background: Hypertrophic scar (HS) is a common fibrotic skin disorder characterized by the excessive deposition of extracellular matrix (ECM). Fibroblasts are the most important effector cells involved in HS formation. Currently no satisfactory treatment has been developed.

Methods: The impact of fraxinellone (FRA) on the proliferation and migration capacity of human hypertrophic scar-derived fibroblasts (HSFs) was assessed by EdU proliferation, wound healing and transwell assays. Quantitative real-time PCR (qRT‒PCR), Western blot (WB), immunofluorescence staining and collagen gel contraction assays were performed to evaluate the collagen production and activation capacity of HSFs. Oxford Nanopore Technologies long-read RNA sequencing (ONT long-read RNA-seq) revealed the occurrence of ferroptosis in HSF and ferroptosis executioner-cathepsin B (CTSB). The mechanisms underlying FRA-induced HSF ferroptosis were examined through fluorescence staining, qRT‒PCR, WB and molecular docking study. The therapeutic efficacy of FRA was further validated in vivo using a rabbit ear scar model.

Results: FRA treatment significantly suppressed the proliferation, migration, collagen production and activation capacity of HSFs. ONT long-read RNA-seq discovered that FRA modulated the expression of transcripts related to ferroptosis and lysosomes. Mechanistically, FRA treatment reduced the protein expression level of glutathione peroxidase 4 (GPX4) and induced the release of CTSB from lysosomes into the cytoplasm. CTSB further induced ferroptosis via spermidine/spermine-N1-acetyltransferase (SAT1)-mediated lipid peroxidation, mitochondrial damage and mitogen-activated protein kinase (MAPK) signalling pathway activation, eventually affecting the function of HSFs. Moreover, FRA treatment attenuated the formation of HS in rabbit ears via CTSB-mediated ferroptosis. The antifibrotic effects of FRA were abrogated by pretreatment with a CTSB inhibitor (CA-074-me).

Conclusions: This study reveals that FRA ameliorates HS by inducing CTSB leakage from lysosomes, causing SAT1-mediated lipid peroxidation, mitochondrial damage and MAPK signalling pathway activation, thus mediating HSF ferroptosis. Therefore, FRA could be a promising therapeutic agent for treating HS.

曲辛内酮介导的溶酶体组织蛋白酶B渗漏诱导成纤维细胞铁下垂以抑制增生性瘢痕形成。
背景:增生性瘢痕(HS)是一种常见的皮肤纤维化疾病,其特征是细胞外基质(ECM)过度沉积。成纤维细胞是参与HS形成的最重要的效应细胞。目前还没有令人满意的治疗方法。方法:采用EdU增殖、创面愈合和transwell实验,观察曲霉内酮(FRA)对人增生性瘢痕源性成纤维细胞(hsf)增殖和迁移能力的影响。采用实时荧光定量PCR (qRT-PCR)、Western blot (WB)、免疫荧光染色和胶原凝胶收缩等方法评价hsf的胶原生成和活化能力。Oxford Nanopore Technologies的长读RNA测序(ONT long-read RNA-seq)揭示了HSF和铁下垂执行分子-组织蛋白酶B (CTSB)的发生。通过荧光染色、qRT-PCR、WB和分子对接研究,探讨fra诱导HSF铁下垂的机制。通过兔耳瘢痕模型进一步验证了FRA的治疗效果。结果:FRA处理显著抑制hsf的增殖、迁移、胶原生成和活化能力。ONT长读RNA-seq发现FRA调节与铁下垂和溶酶体相关的转录本的表达。机制上,FRA处理降低了谷胱甘肽过氧化物酶4 (GPX4)的蛋白表达水平,诱导溶酶体释放CTSB到细胞质中。CTSB进一步通过亚精胺/精胺- n1 -乙酰转移酶(SAT1)介导的脂质过氧化、线粒体损伤和丝裂原活化蛋白激酶(MAPK)信号通路激活诱导铁凋亡,最终影响hsf的功能。此外,FRA治疗通过ctsb介导的铁下垂减少了兔耳HS的形成。用CTSB抑制剂(CA-074-me)预处理FRA的抗纤维化作用被取消。结论:本研究表明,FRA通过诱导溶酶体漏出CTSB,引起sat1介导的脂质过氧化、线粒体损伤和MAPK信号通路激活,从而介导HSF铁凋亡,从而改善HS。因此,FRA是一种很有前景的治疗HS的药物。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
Biology Direct
Biology Direct 生物-生物学
CiteScore
6.40
自引率
10.90%
发文量
32
审稿时长
7 months
期刊介绍: Biology Direct serves the life science research community as an open access, peer-reviewed online journal, providing authors and readers with an alternative to the traditional model of peer review. Biology Direct considers original research articles, hypotheses, comments, discovery notes and reviews in subject areas currently identified as those most conducive to the open review approach, primarily those with a significant non-experimental component.
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