Tissue spreading on implantable substrates is a competitive outcome of cell–cell vs. cell–substratum adhesivity

Abstract

While the interactions of cells with polymeric substrata are widely studied, the influence of cell-cell cohesivity on tissue spreading has not been rigorously investigated. Here we demonstrate that the rate of tissue spreading over a two-dimensional substratum reflects a competition or ''tug-of-war'' between cell-cell and cellsubstratum adhesions. We have generated both a ''library'' of structurally related copolymeric substrata varying in their adhesivity to cells and a library of genetically engineered cell populations varying only in cohesivity. Cell-substratum adhesivity was varied through the poly(ethylene glycol) content of a series of copolymeric substrata, whereas cell-cell cohesivity was varied through the expression of the homophilic cohesion molecules Nand R-cadherin by otherwise noncohesive L929 cells. In the key experiment, multicellular aggregates containing about 600 cells were allowed to spread onto copolymeric surfaces. We compared the spreading behavior of aggregates having different levels of cell-cell cohesivity on a series of copolymeric substrata having different levels of cell-substratum adhesivity. In these experiments, cell-cell cohesivity was measured by tissue surface tensiometry, and cell-substratum adhesivity was assessed by a distractive method. Tissue spreading was assayed by confocal microscopy as the rate of cell emigration from similar-sized, fluorescencelabeled, multicellular aggregates deposited on each of the substrata. We demonstrate that either decreasing substratum adhesivity or increasing cell-cell cohesivity dramatically slowed the spreading rate of cell aggregates.T issue spreading over a substratum is a fundamental process in animal development, wound healing, and malignancy. Understanding the process of tissue spreading is critically important for the emerging field of tissue engineering and for the future use of biomaterials scaffolds for tissue or organ regeneration. Because spreading of a three-dimensional tissue over a surface must come at the expense of associations between cells, greater cell-cell cohesivity should restrain cell emigration (1). Evaluating the expectation that tissue spreading involves a competition between cell-cell and cell-substratum attachments requires the isolation of these two variables from all others. Toward that end, a ''library'' of four cell lines identical except in their cadherin expression and consequently in their cohesivity has been generated. To vary the cell-substratum adhesivity, a library of six structurally related copolymers differing in poly-(ethylene glycol) (PEG) content has been prepared. We then compared the adhesion-related behavior of each of these cell populations on each of these substrata. Materials and MethodsReagents. DMEM, antibiotics (penicillin-streptomycin, gentamycin, G418) and culture supplements (sodium pyruvate; nonessential amino acids, L-glutamine) were purchased from BRL Products (Life Technologies, Gaithersburg, MD). FBS was obtained from HyClone. Fibronectin, laminin, trypsin-EDTA, and PKH2 green fl...

Engineering biological structures of prescribed shape using self-assembling multicellular systems

“…We used gels with differing chemical composition. The relative importance of cell-cell and cell-matrix interactions has been investigated quantitatively (11).…”

“…Cell aggregates have traditionally been used as a powerful tool to understand the principles of cell-cell (10) and cell-matrix adhesion (11), as well as cell sorting (12). In addition, rapid prototyping technology has successfully been applied for computer-aided deposition of cells in gels to create 3D tissue constructs (13,14).…”

Cell and organ printing 2: Fusion of cell aggregates in three‐dimensional gels

“…Recently, important insights have been gained regarding the possible mechanism of aggregate fusion in the presence of extracellular matrix (Ryan et al, 2001). It was demonstrated that competition exists between cell-to-cell forces vs. cell-to-substrate forces.…”

Vascular Endothelial-Cadherin Regulates Cytoskeletal Tension, Cell Spreading, and Focal Adhesions by Stimulating RhoA

“…Although studies have reported that cell-cell cohesivity through cadherins competitively decreases cell-substratum adhesivity, a process known as contact inhibition of cell adhesion and spreading (Yap and Manley, 1993;Ryan et al, 2001), an explanation for the underlying mechanism remains elusive. Cadherin engagement between cells leads to the formation of adherens junctions, the cytoplasmic face of which consists of an adhesion plaque containing many of the same structural proteins as are found in FAs (Pokutta and Weis, 2002).…”

“…Likewise, changes in the engagement of vascular endothelial (VE)-cadherin, the principal junctional molecule in endothelial cells (Dejana et al, 1999), alter both biochemical pathways, such as vascular endothelial growth factor (VEGF) and cell survival signaling (Carmeliet et al, 1999), as well as mechanical states by altering cytoskeletal organization and the local transfer of mechanical stress (Shay-Salit et al, 2002). Although the biochemical and mechanical consequences of integrin-and cadherin-mediated adhesion each have been described, how these adhesions cross talk and cooperate is less well understood.Although studies have reported that cell-cell cohesivity through cadherins competitively decreases cell-substratum adhesivity, a process known as contact inhibition of cell adhesion and spreading (Yap and Manley, 1993;Ryan et al, 2001), an explanation for the underlying mechanism remains elusive. Cadherin engagement between cells leads to the formation of adherens junctions, the cytoplasmic face of which consists of an adhesion plaque containing many of the same structural proteins as are found in FAs (Pokutta and Weis, 2002).…”