Adrenergic receptor modulation of MSC exosome cargo to improve wound healing

慢性伤口，定义为那些不通过正常愈合过程进展， 经过一个月的标准治疗后仍未痊愈，在临床上和经济上都造成了巨大的健康问题。 直接从间充质干细胞中分离治疗剂的兴趣越来越大： 外泌体富集的级分，其现在被理解为携带活性免疫调节和再生活性多肽。 因素它们运载的miRNAs、Wnt配体、生长因子、细胞因子和信号脂质提供了 创伤驻留细胞的旁分泌刺激机制。然而，墨西哥商品的货物内容取决于 培养条件。MSC表达完整的AR补体，包括β2-AR。我们之前的工作 结果表明，β2-AR激动剂增加了MSC的炎性细胞因子分泌，相反， MSC与β2-AR拮抗剂噻吗洛尔，显著增强其抗炎和创伤修复作用， 特性.在此，我们假设，应激性儿茶酚胺激动剂激活MSC β2-AR，改变了MSC β2-AR的表达。 外泌体货物内容物，更促炎，相反，β2-AR拮抗剂阻断激动剂- 诱导的变化，并将外泌体货物恢复为促修复的抗炎表型。我们的长期 目的是确定用β2-AR拮抗剂引发MSC是否可以产生MSC外泌体（MEX）， 是改善伤口愈合的可行的治疗候选物。我们的主要目标是评估 β2-AR激活和拮抗对MEX货物含量和促进愈合功能的影响。在特定 目的1a：我们将评估β2-AR激活对MEX货物的影响。确定是否应激性儿茶酚胺 配体修饰所述货物以变得更促炎。MEX的蛋白质组学和转录组学分析 内容将被执行。它们的促炎潜力将通过细胞因子释放和体外T 细胞增殖在目标1b中，我们将评估阻断β2-AR对MEX货物的影响。的 将确定MSC内源性产生的儿茶酚胺，以及阻断它们的作用。 MSC在外泌体货物上的活化将如目的1a中分析。第2章：我们要确定 β2-AR激动剂和拮抗剂是否交替导致MEX减少或增加伤口 分别在体内受损愈合伤口模型（db/db小鼠）中的愈合特性。 这项工作将为MEX作为伤口治疗剂的未来发展提供基础信息。

Chronic wounds, defined as those that do not progress through the normal healing process and remain unhealed after one month of standard care, pose tremendous health problems- clinically and economically. There is growing interest in isolating ttherapeutic agents directly from mesenchymal stem cells: particularly the exosome enriched fractions that are now understood to carry the active immunomodulatory and regenerative factors. Their cargo of miRNAs, Wnt ligands, growth factors, cytokines and signaling lipids provide the mechanism for paracrine stimulation of wound resident cells. However, cargo contents of MEX are dependent on culture conditions. MSCs express a full complement of AR, including the β2-AR. Our prior work has demonstrated that β2-AR agonists increase inflammatory cytokine secretion MSCs and conversely, treating MSC with a β2-AR antagonist, timolol, substantially enhances their anti-inflammatory and wound reparative properties. Here we hypothesize that activation of MSC β2-AR by stress catecholamine agonists alters their exosomal cargo contents, to be more pro-inflammatory, and conversely, β2-AR antagonists block agonist- induced changes, and revert exosomal cargo to pro-reparative, anti-inflammatory phenotype. Our long-term goal is to determine whether priming MSC with a β2-AR antagonist can generate MSC exosomes (MEX) that are a viable therapeutic candidate for improving wound healing. Our major objective is to evaluate the effects of β2-AR activation and antagonism on MEX cargo contents and function in improving healing. In Specific Aim 1a: we will valuate the effects of activation of the β2-AR on MEX cargo. Determine if stress catecholamine ligands modify the cargo to become more pro-inflammatory. Proteomic, and transcriptomic analyses of MEX contents will be performed. Their pro-inflammatory potential will be evaluated by cytokine release and in vitro T cell proliferation. In Aim 1b we will evaluate the effects of blockade of the β2-AR on MEX cargo. The endogenous generation of catecholamines by MSC will be determined, and the effect of blockade of their activation of MSC on exosome cargo will be analyzed as in Aim 1a. In Specific Aim 2: we will determine whether β2-AR agonists and antagonists alternatively result in MEX with diminished or enhanced wound healing properties, respectively, in an in vivo impaired healing wound model (db/db mouse). This work will provide foundational information for the future development of MEX as a wound therapeutic.