Cell-free regenerative approach in wound healing

伤口愈合中的无细胞再生方法

• 批准号: 10616469
• 负责人: Niketa A. Patel
• 金额: --
• 依托单位: JAMES A. HALEY VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10616469
• 关键词: Acceleration; Adult; Affect; Aging; Amputation; Biochemical; Biological; COVID-19; COVID-19 patient; Cell Death; Cells; Chemicals; Chronic; Cicatrix; Clinic; Complement; Data; Dermal; Diabetes Mellitus; Economic Burden; Evaluation; Fibroblasts; Genomics; Goals; Harvest; Homeostasis; Human; Impaired wound healing; In Vitro; Infection; Inflammation; Inflammatory; Injury; Intervention; Ischemia; Joints; Laceration; Learning; Lesion; Measures; Mechanics; Mediating; Molecular Target; Natural regeneration; Operative Surgical Procedures; Outcome; Pathway interactions; Patients; Pharmacodynamics; Physiological; Probability; Process; Proliferating; Psychological Impact; Quality of life; Readiness; Recovery of Function; Regenerative pathway; Research; Research Design; Risk Factors; Rodent; Safety; Severities; Site; Skin; Skin repair; Small Interfering RNA; Smoking; Strategic Planning; Structure; Surgical Wound Infection; Surgical incisions; System; Therapeutic; Tissues; Topical application; Transcriptional Activation; Translating; United States Department of Veterans Affairs; Untranslated RNA; Veterans; Veterans Disability Claims; Wound models; adipose derived stem cell; chronic wound; comorbidity; exosome; experience; experimental study; genome-wide; healing; improved; improved outcome; in vivo; in vivo Model; infection risk; injury recovery; interdisciplinary approach; keratinocyte; knock-down; novel; overexpression; pre-clinical; regenerative approach; repaired; response; skin wound; stem cell exosomes; tissue regeneration; tissue repair; transcriptome sequencing; treatment response; wound; wound closure; wound dressing; wound healing; wound treatment

Wounds, which are breaks in structure and function of skin, are healed through a regulated process initiated with tissue homeostasis followed by repair response involving inflammation, proliferation and remodeling. Wounds may be due to cuts and lacerations or incisional wounds post surgeries. At times, this healing is delayed or slowed resulting in chronic recalcitrant wounds. Risk factors for impaired wound healing include surgical site infection and comorbidities such as aging, smoking, diabetes. Strikingly, ongoing research with COVID19 patients showcased increased severity in wounds and lesions compared to normal patients(1-4). Recalcitrant wounds per se are not life-threatening but often lead to chronic wound related sequelae. The long-term outcome of impaired wound healing may lead to scarring or amputations, and often has a substantial psychological impact. The treatment of recalcitrant wounds and stimulation of healing is a critical need and consistent with priorities outlined by Department of Veterans Affairs (2018-2024 Strategic plan). While multiple advances in wound dressing materials promote healing, a noteworthy gap remains in the interventions available which considerably improve the outcome of injury. Towards this goal, a therapeutic approach was undertaken to harvest the potential of exosomes derived from human adipose stem cells (hASCexo) mediating tissue repair and regeneration. Thus, the overarching hypothesis is that hASCexo enable wound healing and recovery of function by modulating genomic pathways following injury. This project will elucidate the safety and efficacy of hASC exosomes applied topically to repair dermal wounds to move the therapy closer to the clinic. Towards this goal, the proposal will undertake an in vitro evaluation of genomic targets complemented with an in vivo approach to determine response to treatment. These hypotheses will be validated using a multi-disciplinary approach including pre- clinical, physiological, cellular and biochemical experiments. Specific Aim 1: Determine the molecular targets of the lncRNA cargo of hASC exosomes that promote wound healing: Using RNAseq to examine the contents of hASC exosomes, two long noncoding RNAs (lncRNAs) were identified which are highly enriched in the hASCexo and are pivotal in recovery post injury. Human dermal fibroblasts (HDF) and human epidermal keratinocytes (HEK) will be used with siRNA mediated knockdown or over-expression of these lncRNA in hASCexo. Cellular and biochemical outcomes will be measured in in vitro wound models along with elucidation of genomic changes in response to hASCexo treatment. These will be validated ex vivo in human skin explants. Specific Aim 2: Evaluation of response to hASC exosomes’ treatment in an in vivo wound model: The hypothesis is that repair and regeneration of wounds by topical application of hASCexo depends on its pharmacodynamics and efficacy. To elucidate this along with its genomic impact, an in vivo rodent ischemic wound model will be used to determine physiological response of hASCexo treatment to improve healing outcomes. Its efficacy to promote healing in conditions of underlying infection will be evaluated The hASCexo provide a novel, cell-free regenerative approach to accelerate wound closure and to alleviate chronic effects of impaired wound healing. Robust preliminary data supports feasibility and successful implementation of hASC exosomes’ topical treatment in wound healing. The research design is expected to greatly enhance translatability by the “learn and confirm” approach by integrating the in vitro results in the in vivo model. The therapeutic potential of hASCexo will be thoroughly evaluated for safety and scalability such that it may be translated successfully to the treatment of Veterans’ wounds and substantially improve outcome of chronic wound related sequelae.

伤口是皮肤结构和功能的破坏，通过一个受调节的过程愈合 起始于组织稳态，随后是涉及炎症、增殖和炎症的修复反应。 重塑伤口可能是由于手术后的割伤和撕裂伤或切口伤口。有时候， 这种愈合被延迟或减慢，导致慢性痉挛性伤口。受损的风险因素 伤口愈合包括手术部位感染和合并症，例如衰老、吸烟、糖尿病。 引人注目的是，正在进行的COVID 19患者研究显示，伤口严重程度增加， 与正常患者相比（1-4）。顽固性伤口本身并不危及生命，但往往 导致慢性创伤相关后遗症。伤口愈合受损的长期结果可能导致 到疤痕或截肢，并且往往会产生重大的心理影响。治疗 愈合伤口和刺激愈合是一个关键的需要，并与优先事项概述一致 退伍军人事务部（2018-2024年战略计划）。虽然伤口上有多处伤口 敷料材料促进愈合，但在可用的干预措施中仍存在值得注意的差距， 大大改善了受伤的后果。为了实现这一目标， 进行收获来自人脂肪干细胞（hASCexo）的外泌体的潜力 介导组织修复和再生。因此，总体假设是hASCexo使 通过调节损伤后的基因组途径来促进伤口愈合和功能恢复。这 该项目将阐明局部应用hASC外泌体修复皮肤的安全性和有效性。 让治疗更接近诊所。为了实现这一目标，该提案将在 基因组靶点的体外评价与体内方法互补，以确定对 治疗这些假设将使用多学科方法进行验证，包括预 临床、生理、细胞和生化实验。具体目标1：确定分子量 促进伤口愈合的hASC外泌体的lncRNA货物的靶点：使用RNAseq 检查hASC外泌体的内容物，鉴定了两种长的非编码RNA（lncRNA）， 在hASCexo中高度富集，并且在损伤后的恢复中是关键的。人真皮成纤维细胞 (HDF)和人表皮角质形成细胞（HEK）将与siRNA介导的敲低一起使用， 这些lncRNA在hASCexo中的过表达。将测量细胞和生化结果 在体外创伤模型中沿着阐明响应于hASCexo的基因组变化 治疗这些将在人体皮肤外植体中进行离体验证。具体目标2：评估 在体内伤口模型中对hASC外泌体治疗的反应：假设修复和 通过局部应用hASCexo的伤口再生取决于其药效学， 功效为了阐明这沿着其基因组影响，体内啮齿动物缺血性伤口模型将 用于确定hASCexo治疗的生理反应，以改善愈合结果。其 将评价在潜在感染条件下促进愈合的功效 hASCexo提供了一种新的无细胞再生方法来加速伤口闭合 并减轻受损伤口愈合的慢性效应。强大的初步数据支持可行性 以及在伤口愈合中成功实施hASC外来体的局部治疗。研究 设计通过“学习和确认”的方法， 体外结果是体内模型。hASCexo的治疗潜力将被彻底 评估安全性和可扩展性，以便可以成功地转化为治疗 退伍军人的伤口，并大大改善慢性伤口相关后遗症的结果。