**1.2 Spatiotemporal regulation of cell–cell adhesions**

Altering the number of cellular adhesions is critical to many biological processes during tissue development and cancer progression. For instance, the interconnected nature of epithelial cells, which line the surface of organs, tissues, and blood vessels, designates their polarity, which is critical to their function. EMT takes place when epithelial cells lose the adhesions to other cells and therefore their basal-apical polarity. The resulting mesenchymal cell has increased cellular motility and invasiveness. This process takes place naturally to produce the mesoderm, one of the germ layers, during embryonic development [8, 9], pro-inflammatory wound healing [10], and during cancer cell metastasis [11–13].

Before the development of the germ layers, the embryonic stem cells in the inner mass of the blastocyst are largely epithelial in characteristic; however, during germ layer development, gastrulation, the epithelial-like cells undergo EMT to form the

### **Figure 1.**

*E-cadherin dependent cell–cell adhesion. The E-cadherin consists of five extracellular domains, one transmembrane domain and an intracellular domain. During binding of two E-cadherin molecules the proteins p120, β-catenin, ɑ-catenin, and vinculin get recruited to the intracellular domain leading to cytoskeletal adhesion and actomyosin based activation.*

### *Spatiotemporal Regulation of Cell–Cell Adhesions DOI: http://dx.doi.org/10.5772/intechopen.97009*

mesoderm. I*n vitro* culturing of embryonic stem cells or epiblast cell colonies, shows that they lose expression of E-cadherin, vimentin, and N-cadherin, thus giving rise to cells with a mesenchymal phenotype. The opposite of EMT, mesenchymal-epithelial transition (MET) also occurs naturally and can be seen in the procedure by which induced pluripotent stem cells are formed from fully differentiated cells. This process requires the transition from a cmesenchymal phenotype to an epithelial phenotype, and the activation of epithelial genes encoding epithelial cell junction proteins [8].

EMT extends to carcinomas as well, where a subpopulation of self-renewing cells, known as cancer stem cells, can efficiently generate new tumors. This can be seen in mammary carcinomas following the induction of EMT, which promotes the generation of clusters of invasive mammary gland cells [14]. The extent of these epithelial connections can also be seen in metastatic experiments involving the mammary cancer cell line MCF-7, which maintains an epithelial-like phenotype. In these experiments, MCF-7 is added on top of mammary endothelial cell sheets, and the invasiveness of MCF-7's was evaluated over increasing crossflow, it was revealed that the majority of MCF-7 cells could not form strong adhesions thereby failing to invade. Instead, the MCF-7 s remained rounded and rolled across the surface of the endothelial sheet [13].

Cadherin connections also guide cell migration through their intracellular connection to the cytoskeleton. For instance, in experiments examining the effect of cadherin adhesions in binary cell systems, it was revealed that single adhesions quickly recruit more cadherins to the initial contact site. Additionally, each recruited cadherin binds to the actin cytoskeleton preventing its depolymerization and enabling actomyosin-based mechanical signals [2, 15–17]. Additionally, cadherin-based stabilization of actin in migrating cells leads to *in situ* blebbing of the plasma membrane. These develop the leading edge for the cell, which in turn coordinates the migration of the cell [18]. In tissues with lots of interlocking cadherins, these effects lead to the development of leader cells, which migrate in front of the main body of follower cells. This is an event very common in angiogenesis, where sprouting endothelial cells lead to the development of new blood vessels [19].

### **1.3 Bottom-up tissue engineering**

Another aspect for which controlled cell–cell adhesions are crucial is in bottom-up tissue engineering, in which single cells are organized into either planar or threedimensional structures [20]. Since bottom-up engineering does not rely on external matrices to sequester the cells and instruct cellular arrangement the ability to spatiotemporally control the cell–cell connections is critical to building the desired structure. Techniques for creating bottom-up tissues include bioprinting, construction of cell sheets, and self-assembly of multicellular aggregates [20–23].

Self-assembled multicellular aggregates form by mixing multiple cell types such that microtissues with desired organization form. Generally, these structures form based on minimizing the potential internal energy resulting from cell–cell adhesions [24, 25]. Self-assembled aggregates have been used to construct multicell neuroorganoids comprised of cortical neural progenitor cells, endothelial cells, and mesenchymal stem cells. Different aggregates of each or a mix of two cells were first created in low-attachment 96-well plates. Following aggregate production, aggregates were then mixed to fuse the three cell populations. The resultant aggregate then sorted to form discrete layers within the aggregate. The cortical neural progenitor and endothelial cells developed into vascularized cortical brain tissue, while the mesenchymal

stem cells took on a supportive role in the core of the aggregate [26]. With the ability to spatiotemporally control cell–cell adhesions it becomes possible to self-assemble cells together to produce more complex tissues that better recapitulate the *in vivo* structure.
