Engineering Immunomodulatory Scaffolds for Dental Pulp Regeneration

项目总结 这个K01奖项描述了一个为期3年的培训计划，以支持Franca博士向科学 独立。弗兰卡博士是一名牙医兼科学家，她的长期目标是领导一个研究项目，以(I) 开发用于牙科和颅面再生的免疫调节生物材料和(Ii)创建组织 了解和治疗口腔疾病的芯片。这款K01的科学目标是开发一种 用于牙髓再生的免疫调节支架。尽管发展取得了不可否认的进步 对于牙髓、坏死牙的再生策略提出了具有挑战性的再生条件。 即使在仔细消毒后，以前感染的根管的微环境也会残留 脂多糖(LPS)，激活免疫细胞，使其处于与组织不相容的促炎状态 再生。最近，免疫反应的早期事件被证明对组织有很大的影响。 工程战略的成功。特别是，巨噬细胞对生物材料的反应已被确认为 支架重塑的基本介质和免疫介导再生的潜在靶点 战略。这些细胞可以通过将其表型从促炎改变为 促进再生，有许多中间状态，在一个称为M1/M2极化的过程中。这两种状态 具有高度的可塑性，具有协同作用，并可通过支架的特性进行调节。当一个脚手架 由于牙髓再生是与脂多糖污染的牙本质接触的，可以预期M1 如果不提供组织形成所需的促再生环境，激活将持续存在。 已经在革兰氏阴性感染和靶标中使用了小抗体或纳米抗体来中和内毒素 难以接触的组织微环境中的病原体。不幸的是，这种治疗工具并没有 用于牙齿再生。我们的主要假设是一种免疫调节支架 这中和了内毒素并增加了支架-组织界面中的M2/M1比率，将提供一个可持续的 促进再生的微环境更有利于牙髓组织的形成。因此，我们的目标是 设计了一种新型的载纳米体光交联水凝胶，具有微调的硬度和 中和牙本质脂多糖。这种支架将能够可控地引导M1巨噬细胞的激活， 尽量减少它们在根管内的存在，以便进入的单核细胞可以极化成M2细胞， 渗透水凝胶，分泌抗炎细胞因子环境，促进牙髓组织形成。 我们提出了以下目标：(目标1)确定支架刚性对M1/M2巨噬细胞的影响 极化，(目标2)中和牙本质内毒素和减少M1极化，以及(目标3)评估 纳米载体支架在体内的免疫调节能力。这些目标的成功实现将 生成支架属性和巨噬细胞功能之间相互作用的数据，形成以下基础 R01意见书和其他新型免疫调节材料的开发。

PROJECT SUMMARY This K01 award describes a 3-year training program to support Dr. Franca's transition to scientific independence. Dr. Franca is a dentist-scientist, and her long-term goal is to lead a research program to (i) develop immunomodulatory biomaterials for dental and craniofacial regeneration and (ii) to create tissue chips to understand and treat oral diseases. The scientific goal of this K01 is to develop an immunomodulatory scaffold for dental pulp regeneration. Despite undeniable progress to develop regenerative strategies for the dental pulp, necrotic teeth present challenging conditions for regeneration. Even after careful disinfection, a previously infected root canal's microenvironment has remnant lipopolysaccharide (LPS) that activates immune cells into a pro-inflammatory state incompatible with tissue regeneration. Recently, early events of the immune response were shown to highly influence tissue engineering strategies' success. Especially, macrophage responses to biomaterials have been identified as essential mediators of scaffold remodeling and promising targets for immune-mediated regenerative strategies. These cells can respond to external stimuli by changing their phenotype from pro-inflammatory to pro-regenerative, with many intermediate states, in a process known as M1/M2 polarization. These two states are highly plastic, function synergistically, and can be modulated by the scaffold's properties. When a scaffold for dental pulp regeneration is placed in contact with LPS-contaminated dentin, it can be expected that M1 activation will persist without providing the pro-regenerative environment necessary for tissue formation. Small antibodies or nanobodies have been used to neutralize LPS in Gram-negative infections and target pathogens in scarcely accessible tissue microenvironments. Unfortunately, this therapeutic tool has not been used for dental regeneration purposes. Our overarching hypothesis is that an immunomodulatory scaffold that neutralizes LPS and increases the M2/M1 ratio in the scaffold-tissue interface will provide a sustainable pro-regenerative microenvironment more favorable to dental pulp tissue formation. Hence, our goal is to engineer a novel nanobody-laden photocrosslinkable hydrogel with fine-tuned stiffness and the ability to neutralize dentin LPS. This scaffold will enable controllable guidance of M1 macrophages activation, minimizing their presence within the root canal so that incoming monocytes can polarize into M2 cells, infiltrate the hydrogel, and secret an anti-inflammatory cytokine milieu to foster dental pulp tissue formation. We propose the following aims: (Aim 1) to determine the effect of scaffold's stiffness on M1/M2 macrophage polarization, (Aim 2) to neutralize dentin LPS and reduce M1 polarization, and (Aim 3) to evaluate the nanobody-laden scaffold's immunomodulatory capacity in vivo. Successful completion of these aims will generate data on the interplay between scaffold's properties and macrophage function, forming the basis for an R01 submission and the development of additional novel immunomodulatory materials.