Secretome of stem cells from human exfoliated deciduous teeth (SHED) and its extracellular vesicles improves keratinocytes migration, viability, and attenuation of H2 O2 -induced cytotoxicity.

IF 3.8 3区 医学 Q2 CELL BIOLOGY
Wound Repair and Regeneration Pub Date : 2023-11-01 Epub Date: 2023-12-10 DOI:10.1111/wrr.13131
Juliana Girón Bastidas, Natasha Maurmann, Juliete Nathali Scholl, Augusto Ferreira Weber, Raíssa Padilha Silveira, Fabricio Figueiró, Marco Augusto Stimamiglio, Bruna Marcon, Alejandro Correa, Patricia Pranke
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Abstract

Therapies for wound healing using the secretome and extracellular vesicles (EVs) of mesenchymal stem/stromal cells have been shown to be successful in preclinical studies. This study aimed to characterise the protein content of the secretome from stem cells from human exfoliated deciduous teeth (SHED) and analyse the in vitro effects of SHED-conditioned medium (SHED-CM) and SHED extracellular vesicles (SHED-EVs) on keratinocytes. EVs were isolated and characterised. The keratinocyte viability and migration of cells treated with SHED-EVs and conditioned medium (CM) were evaluated. An HaCaT apoptosis model induced by H2 O2 in vitro was performed with H2 O2 followed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and live/dead assays. Finally, the expression of vascular endothelial growth factor (VEGF) in keratinocytes treated with secretome and EVs was evaluated by immunofluorescence staining and confirmed with RT-qPCR. SHED-EVs revealed a cup-shaped morphology with expression of the classical markers for exosomes CD9 and CD63, and a diameter of 181 ± 87 nm. The internalisation of EVs by HaCaT cells was confirmed by fluorescence microscopy. Proteomic analysis identified that SHED-CM is enriched with proteins related to stress response and development, including cytokines (CXCL8, IL-6, CSF1, CCL2) and growth factors (IGF2, MYDGF, PDGF). The results also indicated that 50% CM and 0.4-0.6 μg/mL EVs were similarly efficient for improving keratinocyte viability, migration, and attenuation of H2 O2 -induced cytotoxicity. Additionally, expression of VEGF on keratinocytes increased when treated with SHED secretome and EVs. Furthermore, VEGF gene expression in keratinocytes increased significantly when treated with SHED secretome and EVs. Both SHED-CM and SHED-EVs may therefore be promising therapeutic tools for accelerating re-epithelialization in wound healing.

人脱落乳牙(SHED)干细胞及其细胞外囊泡的分泌组改善了角质形成细胞的迁移、活力和H2诱导的细胞毒性的衰减。
在临床前研究中,利用间充质干细胞的分泌组和细胞外囊泡(EVs)治疗伤口愈合已被证明是成功的。本研究旨在表征人脱落乳牙(SHED)干细胞分泌组的蛋白质含量,并分析SHED条件培养基(SHED- cm)和SHED细胞外囊泡(SHED- ev)对角质形成细胞的体外影响。细胞外囊泡的分离和表征。对经shed - ev和条件培养基(CM)处理的细胞的角质形成细胞活力和迁移进行了评估。采用H2 O2诱导HaCaT体外凋亡模型,然后进行3-(4,5-二甲基噻唑-2-基)-2,5-二苯基溴化四唑(MTT)和活/死实验。最后,用免疫荧光染色法检测经分泌组和ev处理的角质形成细胞中血管内皮生长因子(VEGF)的表达,并采用RT-qPCR法证实。shed - ev呈杯状,表达外泌体CD9和CD63的经典标记,直径为181±87 nm。荧光显微镜下证实了HaCaT细胞对ev的内化。蛋白质组学分析发现,shedcm富含与应激反应和发育相关的蛋白质,包括细胞因子(CXCL8、IL-6、CSF1、CCL2)和生长因子(IGF2、MYDGF、PDGF)。结果还表明,50% CM和0.4-0.6 μg/mL ev在改善角质细胞活力、迁移和抑制H2 O2诱导的细胞毒性方面具有相似的效果。此外,当SHED分泌组和ev处理时,角化细胞上VEGF的表达增加。此外,当SHED分泌组和ev处理时,VEGF基因在角质形成细胞中的表达显著增加。因此,在伤口愈合过程中,SHED-CM和SHED-EVs可能都是很有前途的加速再上皮化的治疗工具。
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
Wound Repair and Regeneration
Wound Repair and Regeneration 医学-皮肤病学
CiteScore
5.90
自引率
3.40%
发文量
71
审稿时长
6-12 weeks
期刊介绍: Wound Repair and Regeneration provides extensive international coverage of cellular and molecular biology, connective tissue, and biological mediator studies in the field of tissue repair and regeneration and serves a diverse audience of surgeons, plastic surgeons, dermatologists, biochemists, cell biologists, and others. Wound Repair and Regeneration is the official journal of The Wound Healing Society, The European Tissue Repair Society, The Japanese Society for Wound Healing, and The Australian Wound Management Association.
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