Understanding Biomaterials Stimulated Organ Regeneration

许多正在测试或临床使用的生物材料都是塑料基的，在动物和早期临床试验中具有良好的短期效果。然而，长期的结果可能是次优的，甚至是有害的。基于细胞外基质（ECM）的生物材料现在正在测试刺激再生，但它们实际上是否像人们所认为的那样更安全，因为已经有报道称死亡。我的长期目标是了解生物材料和细胞是如何相互作用的，以确定促进安全再生的“好”生物材料，并避免“坏”的导致问题。细胞外囊泡（EV）是细胞间通讯的关键组成部分。它们含有广泛的货物（mRNA，microRNA，蛋白质，信号分子和脂质），调节再生，并已被提议作为无细胞治疗剂。然而，没有关于生物材料诱导EV产生和再生的信息。我将检验EV是生物材料刺激再生的效应子的假设，并且它们的表达模式和货物将使我们能够在长期结果方面区分好的与坏的或中性的生物材料。我将使用角膜作为实验模型，因为由一系列生物材料制成的角膜植入物具有悠久的历史，记录了长期临床结果，可用于帮助生产EV“条形码”（大小，内容，表面标记）。我将开始以生物材料的形式，设计成促进再生的支架。我们将制造由甲基丙烯酸酯（再生不良）、pHEMA/胶原蛋白（再生中等）、重组人胶原蛋白（再生良好）和基于胶原蛋白模拟类似物的新材料制成的植入物。聚乙二醇（PEG）水凝胶由于其惰性已被证明是被动的和生物相容的。将制造一组基于PEG的支架，其中我们将添加增量的胶原蛋白和胶原蛋白样肽，直到这些支架促进角膜和神经细胞的最佳生长，但不激活抗原呈递树突状细胞，以检查EV曲线随细胞友好性增加的变化。我们将基于它们的亚型（微泡/微粒/外泌体;外泌体;或凋亡小体）和它们在体外细胞-材料相互作用期间的蛋白质和mRNA货物来定义EV表达，通过将细胞作为2D构建体接种到材料上，并将它们的相互作用作为我们已经非常好地表征的3D构建的角膜等同物中的“植入物”进行比较。这些相互作用将通过使用双光子共聚焦显微镜的共聚焦显微镜在真实的时间内观察到。体外结果将与体内植入相关。我们将是一个条形码的电动汽车和内容，将指向生物材料促进成功的再生与那些给穷人再生或不良反应。

Many biomaterials being tested or used clinically are plastic-based with promising short-term results in animals and early clinical trials. Longer term results, however, can be sub-optimal to harmful. Biomaterials based on the extracellular matrix (ECM) are now being tested for stimulating regeneration but are they actually safer as believed, as there have been reported fatalities. My long-term goal is to understand how biomaterials and cells interact, to identify the "good" biomaterials that promote safe regeneration and avoid "bad" ones leading to problems. Extracellular vesicles (EVs) are key components in intercellular communication. They contain a wide range of cargo (mRNA, microRNA, proteins, signaling molecules and lipids) that modulate regeneration and have been proposed as cell-free therapeutic agents. However, there is no information on biomaterials-induced EV production and regeneration. I will test the hypothesis that EV are effectors for biomaterials-stimulated regeneration and that their pattern of expression and cargo will allow us to distinguish the good versus bad, or neutral biomaterials in terms of long-term outcomes. I will use the cornea as an experimental model as there is a long history of corneal implants made from a range of biomaterials with documented long-term clinical outcomes that can be used to aid in producing the EV “barcodes” (size, content, surface markers). I will begin with biomaterials in the form of implants that are designed as pro-regeneration scaffolds. We will fabricate implants made from methacrylates (poor regeneration) to blends of pHEMA/collagen (mediocre regeneration) to recombinant human collagen (excellent regeneration) and new materials based on mimetic analogs of collagen. Polyethylene glycol (PEG) hydrogels have been shown to be passive, and biocompatible by virtue of their inertness. A set of PEG-based scaffolds will be fabricated where we will add incremental amounts of collagen and collagen-like peptides until these scaffolds promote optimal growth of corneal and nerve cells, but not activate antigen presenting dendritic cells, in order to examine the changes in EV profile with increasing cell-friendliness. We will define EV expression based on their sub-type (microvesicles/microparticles/ectosomes; exosomes; or apoptotic bodies) and their protein and mRNA cargoes during cell-material interactions in vitro, by seeding cells onto materials as 2D constructs and compare their interactions as “implants” in 3D constructed corneal equivalents that we have characterized very well. These interactions will be observed in real time by confocal microscopy using two-photon confocal microscopy. In vitro results will be corelated with in vivo implantation. Our will be a barcode of EVs and contents that will point to biomaterials promoting successful regeneration versus those that give poor regeneration or an adverse reaction.