Novel Mechanisms of Regenerative Tissue Repair

项目总结 出生后的皮肤损伤会留下疤痕，造成重大的健康、心理和经济负担。我们的 研究小组研究了胎儿再生组织修复的机制，最终目标是开发抗 纤维化疗法。在执行这项任务的过程中，在之前的R01支持下，我们的团队发现IL-10是 胎儿伤口修复和促进无疤痕愈合的关键调节因子，(Ii)复制了胎儿无疤痕表型 通过将IL-10应用于出生后皮肤伤口，(III)揭示了控制细胞外的新机制 基质(ECM)通过高分子量透明质酸(HMW-HA)信号的重塑和新生血管， (Iv)证实HMW-HA驱动T淋巴细胞介导的IL-10表达，(V)最近报道了 小鼠皮肤创面中以CD4T淋巴细胞为主的暂时性流入与 伤口愈合的增殖期和重建期，以及(Vi)获得的证据表明 过继转移功能性CD4T淋巴细胞对严重联合免疫缺陷小鼠的免疫作用 显著减少炎症，减少纤维化，并以更少的费用愈合受伤后的真皮伤口 伤痕累累。虽然我们的集体数据强烈表明HMW-HA介导的IL-10依赖的串扰 在成纤维细胞和动员的T淋巴细胞之间对损伤的反应，对其身份还知之甚少 产生IL-10的CD4T淋巴细胞亚群及其对皮肤创伤修复结果的影响。我们 假设：(A)CD4淋巴细胞亚群通过IL-10对出生后皮肤瘢痕形成有不同的调节作用。 依赖的信号和(B)HA和透明质粘附素转导信号以通过CD4驱动IL-10的产生 淋巴细胞，可降低结疤风险。以获得支持我们假设的实验证据 为设计有效的抗瘢痕疗法提供了令人信服的线索，我们提出了三个具体目标。在目标1中，我们 将确定产生IL-10的CD4T淋巴细胞亚群调节炎症的机制，ECM 可在体外和体内减少瘢痕的形成和血管生成。在这一目标和后续目标中，我们将 使用独特的10Bit4小鼠模型，该模型采用三重报告系统-IL-4(RFP)、IL-10(CD90.1)和 Foxp3(GFP)-明确评估不同T淋巴细胞亚群的需求 具有抗疤痕的功效。在目标2中，我们将研究HA如何调节CD4淋巴细胞的产生 IL-10通过CD44信号转导，并确定稳定透明质酸的透明质酸如何影响这一信号级联。 最后，在目标3中，我们将确定HA介导的内源性IL-10的产生是否可以引导成人成纤维细胞 向胎儿样再生表型转变，以及这种信号是否能影响人类的疤痕表型。 利用我们所获得的知识，我们将优化基于HMW-HA的平移型水凝胶处理系统 促进皮肤创面CD4淋巴细胞稳定产生IL-10及其对瘢痕形成的影响 临床前伤口愈合模型的结果。总体而言，拟议的目标将揭示新的CD4淋巴细胞 在伤口愈合的生理学，并导致一种新的转化式治疗范例，以减轻真皮疤痕。

PROJECT SUMMARY Postnatal dermal injuries heal with scars, resulting in major health, psychosocial and economic burdens. Our team has studied the mechanisms of fetal regenerative tissue repair with the ultimate goal to develop anti- fibrotic therapies. In pursuit of this mission and with previous R01 support, our team has (i) found that IL-10 is a key regulator of fetal wound repair and promotes scarless healing, (ii) replicated the fetal scarless phenotype by application of IL-10 on postnatal dermal wounds, (iii) revealed new mechanisms that control extracellular matrix (ECM) remodeling and neovascularization via high molecular weight hyaluronan (HMW-HA) signaling, (iv) demonstrated that HMW-HA drives T lymphocyte-mediated IL-10 expression, (v) recently reported a temporal influx of predominantly CD4+ T lymphocytes in murine skin wounds that correlated with the proliferative and remodeling phases of wound healing, and (vi) obtained evidence that reconstitution of immunity by adoptive transfer of functional CD4+ T lymphocytes in severe combined immune deficient mice significantly reduced inflammation, decreased fibrosis, and healed post-injury dermal wounds with less scarring. While our collective data strongly suggest a concerted HMW-HA-mediated IL-10-dependent crosstalk between fibroblast cells and mobilized T lymphocytes in response to injury, little is known about the identity of IL-10-producing CD4+ T lymphocyte subsets and how they contribute to dermal wound repair outcomes. We hypothesize that: (a) CD4+ lymphocyte subsets differentially regulate postnatal dermal scarring via IL-10- dependent signaling and (b) HA and the hyaladherins transduce signals to drive IL-10 production by CD4+ lymphocytes, which reduces scarring risks. To obtain experimental evidence that supports our hypothesis and provides compelling clues to design effective anti-scar therapies, we propose three specific aims. In aim 1, we will identify the mechanisms by which IL-10-producing CD4+ T lymphocyte subsets regulate inflammation, ECM formation and angiogenesis to reduce scar formation in vitro and in vivo. In this and subsequent aims, we will use unique 10Bit4 mouse models engineered with a triple reporter system - IL-4 (RFP), Il-10 (CD90.1), and Foxp3 (GFP) - to unambiguously assess the requirement of different T lymphocyte subsets presumed to transduce anti-scar functions. In aim 2, we will investigate how HA regulates CD4+ lymphocyte production of IL-10 via CD44 signaling, and determine how HA-stabilizing hyaladherins influence this signaling cascade. Finally in aim 3, we will determine if HA-mediated endogenous IL-10 production can direct adult fibroblasts toward a fetal-like regenerative phenotype and if this signaling can influence human scarring phenotypes. Using our gained knowledge, we will then optimize a translational HMW-HA-based hydrogel treatment system to promote stable IL-10 production by CD4+ lymphocytes in dermal wounds and test its effects on scarring outcomes in a preclinical wound healing model. Overall, the proposed aims will reveal new CD4+ lymphocyte physiology in wound healing and lead to a novel translational treatment paradigm to mitigate dermal scarring.