Cell-free regenerative approach in wound healing

伤口愈合中的无细胞再生方法

• 批准号: 10363209
• 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/10363209
• 关键词: Adipose tissue; Adult; Affect; Aging; Amputation; Biochemical; Biological; 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; Implementation readiness; In Vitro; Infection; Inflammation; Inflammatory; Injury; Intervention; Joints; Laceration; Lead; Learning; Lesion; Measures; Mechanics; Mediating; Molecular Target; Natural regeneration; Operative Surgical Procedures; Outcome; Pathway interactions; Patients; Pharmacodynamics; Physiological; Probability; Process; Psychological Impact; Quality of life; Recovery of Function; Regenerative pathway; Research; Research Design; Risk Factors; Rodent; SARS-CoV-2 infection; Safety; Severities; Site; Skin; Skin repair; Small Interfering RNA; Smoking; Strategic Planning; Structure; Surgical Wound Infection; Surgical incisions; System; Therapeutic; Time; Tissues; Topical application; Transcriptional Activation; Translating; United States Department of Veterans Affairs; Untranslated RNA; Veterans; Veterans Disability Claims; Wound models; base; 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 regeneration; skin wound; stem cell exosomes; stem cell therapy; stem cells; 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断裂，通过一个有规律的过程愈合 以组织动态平衡启动，随后是修复反应，涉及炎症、增殖和 改建。伤口可能是由于手术后的割伤和割伤或切伤造成的。有时， 这种愈合被延迟或减慢，导致慢性顽固性伤口。伤残人士的风险因素 伤口愈合包括手术部位感染和诸如衰老、吸烟、糖尿病等并发症。 引人注目的是，对19名COVID19患者进行的研究显示，创伤和 病变与正常患者比较(1-4例)。顽固的伤口本身并不会危及生命，但通常 导致慢性伤口相关后遗症。伤口愈合受损的长期后果可能会导致 会造成疤痕或截肢，并往往会对心理产生重大影响。治疗的方法 顽固性伤口和刺激愈合是一项迫切需要，并符合所述优先事项 退伍军人事务部(2018-2024年战略计划)。虽然创伤的多重进展 敷料促进愈合，但在现有的干预措施中仍然存在一个值得注意的缺口，即 大大改善了受伤的结局。为了实现这一目标，一种治疗方法是 获取人类脂肪干细胞来源的外切体的潜力(HASCexo) 调节组织修复和再生。因此，最重要的假设是hASCexo支持 创伤后通过调节基因组通路的创伤愈合和功能恢复。这 该项目将阐明局部应用HASC外切体修复皮肤的安全性和有效性 以使治疗更接近临床。为了实现这一目标，该提案将进行一次 对基因组靶点的体外评估与体内方法相补充以确定对 治疗。这些假设将使用多学科方法进行验证，包括 临床、生理、细胞和生化实验。具体目标1：确定分子 促进伤口愈合的HASC外切体lncRNA货物的靶点：使用RNAseq 检查HASC外切体的内容，鉴定出两个长的非编码RNA(LncRNAs)，它们 在hASCexo中高度丰富，在损伤后的恢复中起关键作用。人真皮成纤维细胞 (HDF)和人表皮角质形成细胞(HEK)将与siRNA介导的敲除或 这些LncRNA在hASCexo中过表达。细胞和生化结果将被测量 体外创伤模型及hASCexo对基因组变化的影响 治疗。这些将在人体皮肤外植体中得到验证。具体目标2：评估 在活体创伤模型中对HASC外切体治疗的反应：假设修复和 局部应用hASCexo的创面再生取决于其药效学和 功效。为了阐明这一点及其基因组影响，活体啮齿动物缺血损伤模型将 用于确定hASCexo治疗的生理反应，以改善愈合结果。它的 将评估在潜在感染情况下促进愈合的效果 HASCexo提供了一种新的、无细胞的再生方法来加速伤口闭合 并减轻伤口愈合受损的慢性影响。强劲的初步数据支持可行性 成功地实施了HASC exosome在伤口愈合中的局部治疗。这项研究 设计应通过“学习和确认”的方法，通过整合，大大提高可译性 体外实验结果与体内实验结果一致。HASCexo的治疗潜力将是彻底的 对安全性和可扩展性进行评估，以便它可以成功地转化为治疗 减少退伍军人的创伤，大大改善与慢性创伤相关的后遗症的结局。