Regenerative wound healing via inflammation-modulating biomaterials

描述(由申请人提供)：众所周知，长期的炎症阻碍了伤口修复的最佳目标，即以最小的疤痕迅速闭合伤口。然而，组织再生(与伤口修复相对)的目标是重建受损组织，并用丢失组织的功能生物复制品取代受损组织，而不形成疤痕。在后一种情况下，炎症的作用没有得到很好的确定，原因很简单，包括人类在内的大多数哺乳动物很少表现出任何再生能力(肝脏除外)，排除了实验方法。基于我们在小鼠附件损伤模型中的初步观察，这项建议将解决这样一种可能性，即增强至少某些方面的炎症将使伤口愈合转向再生。我们还将研究新型生物材料的使用，以局部控制炎症反应的关键方面，并促进组织再生。在兔子和近亲繁殖的MRL小鼠品系中看到的耳朵完全闭合被认为是哺乳动物再生的例子，类似于两栖动物的肢体再生。软骨置换后耳洞完全闭合，无疤痕形成。在这项提案中，我们提出了体外和原位证据，证明在这种再生反应中，伤口部位是促炎的，并且炎症的药理上调诱导了其他不能再生的小鼠的再生。炎症已被证明受转录因子HIF1a的调控，我们将重点研究HIF1a在C57BL/6小鼠中的上调，以进一步探讨其在再生反应中的作用。由于HIF1a是在蛋白质水平上被调控的，其降解是由脯氨酸羟基酶(PhDS)完成的羟化来启动的，所以我们将考察几种HIF1a降解的抑制剂，包括PHD抑制剂、直接HIF1a羟化抑制剂和泛素合成抑制剂。这些分子将在HIF-荧光素酶报告小鼠及其细胞的体外活性中进行筛选，1)局部用药或可溶性注射剂进入伤口部位或全身给药，2)偶联到生物材料上。鉴于HIF活性调节多达10%的所有细胞功能，特别是代谢状态和炎症，重要的是HIF激活药物仅存在于伤口部位并以受控的方式存在。新的原位形成的大分子生物材料的设计目标是在空间和时间上精细控制伤口处的HIF水平。我们设想，这种模拟愈合伤口复杂组织微环境的方法将允许生物材料降解并伴随组织再生，从而在未来产生新的组织再生临床模式。

DESCRIPTION (provided by applicant): It is well known that prolonged inflammation hinders the optimal goal of wound repair, namely prompt closure of the wound with minimal scarring. However, the objective of tissue regeneration (as opposed to wound repair) is to reconstitute and replace damaged tissue with a functional biological replica of the lost tissue without scarring. In this latter case, the role of inflammation is not well established for the simple reason that most mammals including humans have rarely demonstrated any capacity for regeneration (with the exception of the liver) precluding experimental approaches. Based upon our preliminary observations in a mouse model of appendage injury, this proposal will address the possibility that enhancing at least some aspects of inflammation will re-direct wound healing towards regeneration. We will also examine the use of novel biomaterials to locally harness key aspects of the inflammatory response and facilitate tissue regeneration. Ear whole closure, as seen in rabbits and in the inbred MRL mouse strain is considered to be a mammalian example of regeneration similar to limb regeneration in amphibians. Ear holes close completely with the replacement of cartilage and without scarring. In this proposal, we present in-vitro and in-situ evidence that the wound site during this regenerative response is pro-inflammatory and that pharmacological up- regulators of inflammation induce regeneration in otherwise non-regeneration competent mice. Inflammation has been shown to be regulated by the transcription factor HIF1a and we will focus on HIF1a up-regulation in C57BL/6 mice to further explore its role in the regenerative response. Since HIF1a is modulated at the protein level and its degradation is initiated by hydroxylation accomplished by prolyl hydroxylases (PHDs), we will examine several inhibitors of HIF1a degradation including a PHD inhibitor, a direct HIF1a hydroxylation inhibitor and an inhibitor of ubiquination. These molecules will be screened for activity in HIF-luciferase reporter mice and their cells in-vitro as 1) topical agents or soluble injectants into the wound site or administered systemically and 2) coupled to a biomaterial. Given that HIF activity regulates as much as 10% of all cellular functions and especially the metabolic state and inflammation, it is important that HIF-activating drugs be present only at the wound site and in a controlled manner. Novel in- situ forming macromolecular biomaterials will be designed with the goal of fine control, both spatial and temporal, of HIF levels at the wound site. We envision that this approach of mimicking the complex tissue microenvironment of a healing wound will allow for biomaterial degradation accompanied by tissue regeneration, leading in the future to new clinical modalities for tissue regeneration.