Using axolotls to define innate mechanisms for combatting fibrosis

软组织纤维化和皮肤疤痕每年给1亿患者带来巨大的临床负担，以及 目前，治疗选择相当有限。抗击纤维化和疤痕形成的新方法是 这是必要的。有效的伤口闭合是伤口愈合的关键。如果没有它，受伤的人会多得多 容易失水和感染。有人假设，疤痕进化成了一种解决方案，可以最大限度地 治愈的速度。然而，这一假设并不能解释为什么再生动物，可以说是最多的 非凡的愈合能力，能够无疤痕愈合。在这里，我建议理解Axolotl是如何 火蜥蜴无疤痕地愈合伤口并在再生过程中拮抗纤维化将提供关键 治疗方法。为了克服这些动物对抗纤维化的令人印象深刻的能力，我将创造 轴突的遗传性和化学性纤维化模型。我已经成功地开发了 一种使用药物博莱霉素的化学模型，这种药物限制了动物的再生能力，以及一种基因- 基于TSP-1功能缺失突变体的模型，该突变体降低了再生速度和胚泡大小。在AIM 1，我将进行假设驱动的研究，以及基于发现的研究，以询问两者之间的平衡 纤维化和再生。我将使用基因组编辑来创建TSP-1/TSP-2和TSP-1/TSP-4双突变体 据预测，Axolotls会加剧纤维化表型。同时，我将使用established 凝血酶敏感蛋白抑制肽(可促进培养的人皮肤纤维化表型 成纤维细胞)。对于这两种模型，我将确定对抗轴突纤维化的候选分子靶点。 使用RNAseq和差异基因表达分析。在与组织工程师的合作下，我将使用 这些发现开发了以丝绸为基础的微图案水凝胶，其中负载了旨在抑制 后来可能发展成人类治疗的纤维化。在目标2中，我将确定各种机制， Axolotls可能会在硅胶植入物存在的情况下保持无疤痕组织，这些硅胶植入物通常会导致 人类患者，需要他们的替换。这些机制代表了未来的新方法 可以预防性应用于需要医疗植入的人类患者的治疗方法。他们可能会 在修复植入物周围现有的纤维化方面也要积极。同时，我将测试Axolotl的细胞外 MATRIX重建移植的人类纤维组织的能力。这些方法加在一起，是非常重要的 小说。它们既利用了现在这些非凡动物中可用的分子遗传工具的爆炸性增长， 他们利用针对纤维性侮辱的自然解决方案，而这些解决方案尚未应用于人类。这些 这些策略可以为改善人类患者的纤维化结果提供强有力的新方法。

Soft tissue fibrosis and cutaneous scarring represent huge clinical burdens to 100 million patients per year, and therapeutic options are currently quite limited. Novel approaches to combat fibrosis and scarring are necessary. Efficient wound closure is a crucial part of wound healing. Without it, injuries would be far more susceptible to fluid loss and infection. It has been hypothesized that scarring evolved as a solution to maximize healing speed. This hypothesis, however, does not explain why regenerative animals, with arguably the most remarkable healing abilities, are capable of scar-free healing. Here, I propose that understanding how axolotl salamanders heal wounds scarlessly and antagonize fibrosis during regeneration will provide critical therapeutic approaches. To overcome the impressive ability of these animals to combat fibrosis, I will create both genetic- and chemical-based fibrotic models in axolotls. I have already successfully managed to develop a chemical-based model using the drug bleomycin, which limits the animals' regenerative ability, and a genetic- based model using tsp-1 loss-of-function mutants, which reduces regenerative rate and blastema size. In Aim 1, I will perform hypothesis-driven, as well as discovery-based, studies to interrogate the balance between fibrosis and regeneration. I will use genome editing to create tsp-1/tsp-2 and tsp-1/tsp-4 double-mutant axolotls, which are predicted to have exacerbated fibrotic phenotypes. In parallel, I will use established Thrombospondin-inhibiting peptides (shown to promote fibrotic phenotypes in cultured human dermal fibroblasts). For both models, I will identify candidate molecular targets that antagonize fibrosis in axolotls using RNAseq and differential gene expression analysis. In collaboration with tissue engineers, I will then use these findings to develop micro-patterned, silk-based hydrogels loaded with biologicals designed to inhibit fibrosis that could be later developed toward human treatments. In Aim 2, I will identify mechanisms whereby axolotls might maintain scar-free tissue in the presence of silicone implants that routinely lead to fibrosis in human patients, necessitating their replacement. These mechanisms represent novel approaches for future therapies that might be preventively employed in human patients necessitating medical implants. They might also be active in remediating existing fibrosis around implants. In parallel, I will test the axolotl extracellular matrix's capability to remodel transplanted human fibrotic tissues. Together, these approaches are extremely novel. They leverage both the explosion of molecular genetic tools now available in these remarkable animals, and they capitalize on natural solutions to fibrotic insults that have yet to be applied to humans. These strategies could provide powerful new approaches to improving fibrosis outcomes in human patients.