Adrenergic receptor modulation of MSC exosome cargo to improve wound healing

慢性伤口，定义为那些在正常愈合过程中不会进展并仍然存在的伤口 在一个月的标准护理后仍未痊愈，在临床和经济上都造成了巨大的健康问题。 人们对直接从间充质干细胞中分离治疗药物的兴趣越来越大：特别是 富含外切体组分，现已被认为具有免疫调节和再生活性 各种因素。他们的miRNAs，Wnt配体，生长因子，细胞因子和信号脂质提供了 创面驻留细胞旁分泌刺激机制。然而，MEX的货物含量取决于 关于培养条件。MSCs表达完整的AR，包括β2-AR。我们之前的工作是 研究表明，β2-AR激动剂增加炎性细胞因子的分泌，反之，治疗 含有β2-AR拮抗剂的骨髓间充质干细胞显著增强其抗炎和伤口修复作用 属性。在这里，我们假设应激儿茶酚胺激动剂激活MSC儿茶酚胺受体2-AR改变其β2-AR。 相反，β2-AR拮抗剂阻断激动剂- 诱导改变，并将外体货物恢复为有利于修复的、抗炎的表型。我们的长期合作 目的是确定用β2-AR拮抗剂是否能产生骨髓间充质干细胞外体，从而 是改善伤口愈合的一种可行的治疗方案。我们的主要目标是评估效果 β2-AR的激活和拮抗作用对甲氧西林的货物含量和促愈合作用。具体而言 目标1a：我们将评估β2-AR激活对MEX货物的影响。确定应激儿茶酚胺 配体对货物进行修饰，使其变得更具促炎作用。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 细胞增殖。在目标1b中，我们将评估封锁β2-AR对MEX货物的影响。这个 将测定MSC内源性儿茶酚胺的产生，以及阻断其对其影响 目标1a将分析MSC对Exosome货物的激活。具体目标2：我们将确定 β-2-AR激动剂和拮抗剂是否交替导致创面缩小或增加 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作为一种创伤治疗药物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.