EAGER: Exploring Cell-Cell Gap as a Critical Parameter in Biological Phase Changes

当细胞数量足够大时，生物系统有时会迅速改变行为。 这个早期概念的探索性研究资助（EAGER）项目是基于初步数据，这些数据表明，对于一种类型的哺乳动物细胞，是细胞之间的距离触发了不同的行为。 这是在一种特殊类型的肌肉细胞中观察到的。当细胞相距100微米时，自动形成肌肉“纤维”。 当它们相距超过100微米时，它们就不会。 将在成纤维细胞中进一步检查该观察结果。 成纤维细胞对细胞外基质的压缩是伤口愈合和器官原始形态发生的基础。 该项目将确定100微米的阈值是否也适用于小鼠，猴子和人类的成纤维细胞。 作为观察宽度的测试，还将使用在某些条件下具有聚集成较大结构的性质的原生生物网骨藻来测试100微米阈值。 如果实验结果在不同细胞类型之间具有可比性，则该项目将为成纤维细胞和网骨藻聚集中组织“相变”激活的阈值距离的存在提供重要支持。这可以提高我们对各种条件的理解，在这些条件下，组织显然表现出行为的相变，包括胚胎发生、癌症肿瘤发生、体外组织形成、细菌生物膜形成和微生物世界中的各种集体现象。识别临界距离条件将有助于预测具有细胞和细胞外基质的3D打印组织的形态发生路径。 这项研究将由一名研究生和两名本科生进行。其中一名UG学生将来自生物系，其他人来自工程系。因此，学生将在多学科领域接受培训。该项目将特别努力招募来自少数民族和代表性不足的群体的学生进行研究。该项目的灵感来自最近的实验发现，即在3D培养中，彼此相距100微米以内的肌肉细胞会压缩ECM并形成肌管（PI的实验室），内皮细胞在100微米内形成脉管系统，胶原中的成纤维细胞彼此在100微米内压缩胶原。当细胞-细胞间隙超过100微米时，这些过程都不会发生。 将使用来自多个物种（小鼠、猴和人）的成纤维细胞在与荧光珠混合的胶原基质中测试临界距离的假设，用于光学跟踪黏菌细胞（网柄菌）的压实和簇形成。 为了深入了解阈值细胞-细胞间隙的实验观察，将创建数学模型来模拟实验观察。 每一个细胞通过粘附在基质上和当细胞丝状伪足产生收缩力时拉动纤维来重塑它们周围的基质。每个细胞形成在张力下具有更高刚度的重塑基质区。如果细胞相距很远，它们的区域保持相互独立。在具有许多细胞的系统中，这种非相互作用区域保持全局对称性。当细胞彼此靠近时，它们的区域重叠，对称性被破坏。细胞之间的刚度变得更高，并且细胞在它们之间形成更硬的纤维桥。与其他方向相比，更多的丝状伪足倾向于沿着桥沿着延伸，细胞在保持收缩的同时变得极化和伸长。这导致细胞之间的吸引力相互作用，这将使基质紧凑。将使用来自小鼠（胚胎成纤维细胞，NIH/3 T3）、猴（肾成纤维细胞）和人（肺）的三种类型的成纤维细胞检验基质压实的假设，并将使用黏菌细胞（盘基网柄菌）检验一般性。所有这些细胞类型具有相当的大小，直径约10微米。 因此，我们预测所有细胞类型的临界距离相似。

Biological systems sometimes rapidly change behavior when the number of cells becomes large enough. This EArly-concept Grant for Exploratory Research (EAGER) project is based on preliminary data showing that for one type of mammalian cell, that it is the distance between the cells that triggers the different behavior. This was observed for a particular type of muscle cell. When the cells are 100 micrometers apart, the form into muscle "fibers" automatically. When they are further than 100 micrometers apart, they do not. This observation will be further examined in fibroblasts. The compression of extracellular matrix by fibroblasts is fundamental to the healing of wounds and also to the original morphogenesis of organs. The project will determine whether this threshold of 100 micrometers is also true for fibroblasts from mice, monkeys and humans. As a test of the breadth of the observation, the 100 micrometer threshold will also be tested using the Protista Dictyostelium that has the property of aggregating into larger structures under some conditions. If the results of the experiments are comparable among the different cell types, the project will have developed significant support for the existence of a threshold distance for the activation of a tissue "phase change" in the aggregation of fibroblasts and of Dictyostelium. This could improve our understanding of a variety of conditions where tissues apparently show a phase change in behavior including embryogenesis, cancer tumorigenesis, in vitro tissue formation, bacterial biofilm formation and a variety of collective phenomena in the microbial world. Identification of critical distance conditions will help with predicting the path of morphogenesis of 3D printed tissues with cells and extracellular matrix. The research will be carried out by a graduate and two undergraduate students. One of the UG students will be from the Department of Biology, the others from Engineering. Thus, the students will be trained in multidisciplinary fields. Special effort will be made to recruit students from minorities and under-represented groups to conduct the research.The project is inspired by recent experimental findings suggesting that muscle cells within 100 micrometers of each other in 3D culture compact the ECM and form myotubes (PI's lab), endothelial cells within 100 micrometers of each other form vasculature, fibroblasts in collagen within 100 micrometers of each other compact the collagen. When the cell-cell gap exceeds 100 micrometers, none of these processes occur. The hypothesis of a critical distance will be tested using fibroblasts from multiple species (mouse, monkey and human) in a collagen matrix mixed with fluorescent beads for optically tracking compaction and cluster formation by slime mold cells (Dictyostelium). In order to gain insight on the experimental observations of the threshold cell-cell gap, a mathematical model will be created to simulate the experimental observations. Each cell remodels the matrix around them by adhering to the matrix and by pulling the fibers as the cell filopodia generate contractile forces. Each cell forms a zone of remodeled matrix with higher stiffness under tension. If cells are far apart, their zones remain independent of each other. In a system with many cells, such non-interacting zones maintain global symmetry. When the cells are close to each other, their zones overlap, and symmetry is broken. The stiffness between the cells becomes higher, and the cells form a stiffer fibrous bridge between them. More filopodia tend to extend along the bridge compared to other directions, the cells become polarized and elongated while remaining contractile. This results in an attractive interaction between the cells that will compact the matrix. Three types of fibroblasts from mouse (embryonic fibroblast, NIH/3T3), monkey (kidney fibroblasts) and human (lung) will be used to test hypothesis on matrix compaction, and slime mold cells (Dictyostelium discoideum) will be used to test for generality. All of these cell types have comparable size, ~ 10 micrometer in diameter. Hence, we predict similar critical distances for all cell types.