Probing biochemical/biophysical influences on endothelial-mesenchymal transition

描述（由申请人提供）：最近的几份报告表明，内皮细胞可以分化为间充质细胞类型[1-8]，这一过程被称为内皮-间充质转化（EndoMT或EndMT）。反过来，内皮细胞衍生的间充质细胞能够分化成多种细胞类型，并已被证明分别形成脂肪、软骨和骨。最近的研究表明，EndMT有助于发育后组织的疾病进展，并且深入的研究表明，EndMT在纤维化[1,3 -5]、癌症[2]和与进行性骨化性纤维发育不良（FOP）[8]相关的异位骨化中发挥作用。因此，影响EndMT的因素可能与理解和控制疾病进展特别相关。细胞外基质（ECM）提供了多种指导细胞功能的生化和生物物理线索，在组织发育过程中，几种基质成分与EndMT有关，包括蛋白聚糖[9]、玻璃体粘连蛋白、胶原蛋白、纤维连接蛋白和层粘连蛋白[10-11]。此外，TGF-b超家族生长因子可诱导EndMT[3,8]，可溶性TGF-b对内皮细胞行为的影响取决于培养bb0中存在的ECM成分。此外，生物物理因素也被认为是内皮细胞行为、间充质细胞分化、纤维化和骨病的重要决定因素[5,14 -24]。综上所述，这些先前的研究表明，ecm衍生的信号可能是EndMT的调节因子，进而影响疾病进展。然而，ECM在引导EndMT中的作用在很大程度上是一个未被探索的研究领域，这可能是由于相对较新的发现，该过程发生在成人组织中。在这里，我们提出了一种方法来识别影响EndMT的ecm模拟信号。实验方法是基于我们创造材料的经验来有效地控制细胞-材料的相互作用。该应用的指导假设是，模拟细胞外基质特性的生化和生物物理因素将各自显著影响EndMT的过程。拟议的研究具有重要意义，因为它们将为细胞外基质如何促进EndMT以及与之相关的疾病提供重要的新见解。已知影响成人组织中EndMT的少数因素（如BMP-4、TGF-b1、TGF-b2）以及由此导致的内皮细胞表型变化（如肌成纤维细胞活化、异位成骨）通常与一系列具有重大和广泛相关性的疾病有关。因此，更好地了解EndMT的作用可能对人类健康有实质性的好处。提出的研究是创新的，因为他们使用新颖的，化学定义的细胞培养阵列来有效地探测模拟ecm信号的影响。这种探测模拟ECM信号的方法在EndMT过程中是理想的，因为ECM在EndMT过程中起着鲜为人知但可能至关重要的作用。此外，了解合成生物材料如何影响EndMT最终可能与医疗植入物设计相关，特别是在限制纤维化或异位骨化的情况下。

DESCRIPTION (provided by applicant): Several recent reports have demonstrated that endothelial cells can differentiate into mesenchymal cell types [1-8], a process called endothelial-mesenchymal transition (EndoMT or EndMT). In turn, endothelial cell-derived mesenchymal cells are capable of differentiating into multiple cell types, and have been shown to form fat, cartilage, and bone, respectively[8]. Recent studies indicate that EndMT contributes to disease progression in post-developmental tissue, and thorough investigations have shown roles in fibrosis[1, 3-5], cancer[2], and heterotopic ossification associated with fibrodysplasia ossificans progressiva (FOP) [8]. Therefore, the factors that influence EndMT may be of particular relevance in understanding and manipulating disease progression. The extracellular matrix (ECM) provides a wide variety of biochemical and biophysical cues that guide cell function, and several matrix components have been implicated in EndMT during tissue development, including proteoglycans [9], vitronectin, collagen, fibronectin and laminin [10-11]. In addition, TGF-b superfamily growth factors can induce EndMT[3, 8] and the influence of soluble TGF-b on endothelial cell behavior is dependent on ECM components present in culture [12]. Further, biophysical factors are also known to be important determinants of endothelial cell behavior, mesenchymal cell differentiation [13], fibrosis, and bone disease [5, 14-24]. Taken together, these previous studies implicate ECM-derived signals as likely regulators of EndMT and, in turn, disease progression. However, the role for the ECM in guiding EndMT is a largely unexplored area of research, perhaps due to the relatively recent discovery that this process occurs in adult tissue. Here, we propose an approach to identify ECM-mimetic signals that influence EndMT. The experimental approach is based on our experience creating materials to efficiently control cell-material interactions. The guiding hypothesis of the application is that biochemical and biophysical factors that mimic extracellular matrix properties will each significantly influence the process of EndMT. The proposed studies are Significant, as they will provide critical new insights into how the extracellular matrix contributes to EndMT, and the diseases in which it is implicated. The few factors that are known to influence EndMT in adult tissue (e.g., BMP-4, TGF-b1, TGF-b2) and the changes in endothelial cell phenotype that result (e.g., myofibroblast activation, heterotopic osteogenesis) are generally implicated in a range of diseases that are of substantial and broad relevance. Therefore, a better understanding of the role of EndMT could have a substantial benefit for human health. The proposed studies are Innovative, as they use novel, chemically-defined cell culture arrays to efficiently probe the effects of ECM-mimetic signals. This approach for probing ECM-mimetic signals is ideal in the case of the EndMT process, as the ECM plays a poorly understood but likely critical role in EndMT. Further, understanding how synthetic biomaterials influence EndMT could ultimately have relevance to medical implant design, particularly in scenarios in which it is important to limit fibrosis or heterotopic ossification.