microRNA regulated processes in keratinocytes upon exposure to sulfur mustard: modulation by mimics und anti-miRs

接触硫芥后角质形成细胞中的 microRNA 调节过程：模拟物和抗 miR 的调节

• 批准号: 424562951
• 负责人: Professor Dr. Christian Ries
• 金额: --
• 依托单位: Institut für Prophylaxe und Epidemiologie der Kreislaufkrankheiten
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2019
• 资助国家: 德国
• 起止时间: 2018-12-31 至 2022-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/424562951?language=en
• 关键词: microRNA; regulated; processes; keratinocytes; upon

Sulfur mustard (SM) is a highly toxic warfare agent that represents a continuous threat in various crisis regions, particularly through activities of terroristic groups. Exposure of the skin with SM evokes severe tissue damage and defects in wound healing leading to long-term hospitalization of patients. The underlying molecular and cellular pathomechanisms are poorly investigated. Up to now there is no pharmacological therapy available for the treatment of SM-induced defects in would healing. Own results from in vitro studies in skin cells provided evidence that SM affects the expression of microRNAs (miRNAs) and down-stream pathways and cell functions in keratinocytes. miRNAs are small ubiquitous RNA molecules that bind to specific mRNAs and inhibit subsequent translation thereby contributing to the regulation of important cellular processes. The aim of our studies in keratinocytes is to identify SM-evoked alterations in the expression of miRNAs with relevance in wound healing that may be counterbalanced by use of synthetic miRNAs (mimics) and miRNA inhibitors (anti-miRs) in order to rescue dysregulated cell functions. In the first part of our project, primary human keratinocytes are subjected to analysis by next-generation-sequencing technology aiming to compile a comprehensive signature of miRNAs and mRNAs regulated by SM. To this end, miRNA- and mRNA libraries are generated from the cells and subjected to RNA sequencing. Using specific computer software and various online databases, all miRNAs influenced by SM as well as their putative target mRNAs and potentially involved regulatory networks can be identified. A selection of the most promising miRNAs and target mRNAs are subjected to verification by experimental gene expression analysis. In the second part of our proposal, we use synthetic miRNAs (mimics) and miRNA inhibitors (anti-miRs) to investigate the role of the selected miRNAs in the control of signaling pathways important in wound healing as well as in growth, migration and differentiation of the cells. Finally, the third part of our studies is designed to validate our findings on mimic or anti-miR-based rescuing of SM-evoked cell defects obtained in cell culture by use of a complex three-dimensional epidermal skin model. The present studies allow for the first time the preparation of a comprehensive list of miRNAs and mRNAs that are regulated in keratinocytes upon exposure to SM. This may lead to the discovery and validation of novel molecular pathomechanisms, and provides experimental evidence whether mimics or anti-miRs may be basically useful as topically administrable agents for the treatment of SM-evoked wound healing disorders in the skin.

硫磺芥末（SM）是一种高毒的战争代理商，代表了各个危机地区的持续威胁，特别是通过恐怖组织的活动。用SM暴露于皮肤会引起严重的组织损伤和伤口愈合的缺陷，导致患者长期住院。基础分子和细胞病理机理的研究很少。到目前为止，还没有可用于治疗SM诱导的缺陷的药理疗法。在皮肤细胞中体外研究的结果提供了证据，表明SM会影响角质形成细胞中microRNA（miRNA）和下游途径以及细胞功能的表达。 miRNA是与特定mRNA结合并抑制随后的翻译的小无处不在RNA分子，从而有助于调节重要的细胞过程。我们在角质形成细胞中的研究的目的是鉴定miRNA表达中具有与伤口愈合相关的SM诱发的变化，这可能通过使用合成miRNA（MIMICS）和miRNA抑制剂（抗MIR）来抵消，以营救失调的细胞功能。在我们项目的第一部分中，主要人类角质形成细胞受到下一代测序技术的分析，旨在编译由SM调节的miRNA和mRNA的全面特征。为此，miRNA和mRNA库是从细胞产生的，并进行RNA测序。使用特定的计算机软件和各种在线数据库，可以识别所有受SM及其推定目标mRNA和潜在涉及的监管网络影响的miRNA。通过实验基因表达分析对最有希望的miRNA和靶mRNA进行选择。在提案的第二部分中，我们使用合成miRNA（模拟）和miRNA抑制剂（抗MIR）来研究所选miRNA在控制伤口愈合以及细胞生长，迁移和分化中重要的信号通路中的作用。最后，我们的研究的第三部分旨在验证我们对通过使用复杂的三维表皮皮肤模型在细胞培养中获得的模拟或基于抗MIR的拯救的发现。本研究首次允许在暴露于SM时在角质形成细胞中调节的miRNA和mRNA的全面列表。这可能会导致发现和验证新型分子病理机制，并提供实验证据，是否基本上可以用作局部管理的药物来治疗皮肤中SM诱发的伤口愈合障碍。