Understanding Biomaterials Stimulated Organ Regeneration

许多正在测试或临床使用的生物材料都是以塑料为基础的，在动物身上和早期临床试验中都有很好的短期结果。然而，较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接受刺激再生的测试，但它们实际上是否像人们所认为的那样更安全，因为有报道称有死亡病例。我的长期目标是了解生物材料和细胞是如何相互作用的，找出促进安全再生的“好”生物材料，避免“坏”生物材料导致问题。细胞外小泡(EVS)是细胞间通讯的关键成分。它们含有广泛的物质(信使核糖核酸、微RNA、蛋白质、信号分子和脂类)，可调节再生，并已被提议作为无细胞治疗剂。然而，目前还没有关于生物材料诱导的电动汽车生产和再生的信息。我将测试这样一种假设，即EV是生物材料刺激的再生的效应器，它们的表达模式和货物将允许我们从长期结果的角度区分生物材料的好坏或中性。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有着悠久的历史，这些材料具有有案可查的长期临床结果，可用于帮助产生EV“条形码”(大小、内容物、表面标记)。我将从植入物形式的生物材料开始，这些植入物被设计为促进再生的支架。我们将制造由甲基丙烯酸酯(再生较差)、PHEMA/胶原(再生一般)和重组人胶原(再生较好)的混合物制成的植入物，以及基于胶原蛋白模拟类似物的新材料。聚乙二醇水凝胶因其惰性而被证明是被动的和生物相容的。我们将制作一套基于聚乙二醇的支架，在其中我们将添加增量的胶原和胶原样多肽，直到这些支架促进角膜和神经细胞的最佳生长，但不激活抗原递呈的树突状细胞，以便随着细胞友好性的增加而检测EV图谱的变化。我们将通过将细胞作为2D结构种植到材料上，基于它们的亚型(微泡/微粒子/外胚体；外体；或凋亡小体)及其蛋白质和mRNA的运输来定义EV的表达，并将它们的相互作用作为我们已经很好地描述的3D构建的角膜等价物中的“植入”。这些相互作用将通过使用双光子共焦显微镜的共焦显微镜实时观察到。体外结果将与体内植入相关。我们将是电动汽车的条形码，内容将指向促进成功再生的生物材料，而不是那些再生能力差或不良反应的生物材料。

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.