Cell-to-Cell Communication as a Strategy to Regenerate Three-dimensional Tissue.

Abstract

Cell-to-cell communication is ubiquitous in the majority of cells and is indispensable for proper development of most tissues. Because cells within tissues constantly interact intercellularly to coordinate normal tissue functions and homeostasis, it was hypothesized that increasing gap junction intercellular communication (GJIC), exogenously and endogenously, could improve cell signaling and differentiation in interior regions of tissue engineering equivalents, thereby regenerating larger and more uniform volumes of tissue. The exogenous approach consists of altering the initial cell seeding strategy and biomimetic environment while in the endogenous approach cells are genetically engineered to over-express gap junction proteins.

To test this hypothesis, we used bone as a model tissue, because of the inherent limitation achieving spatially uniform bone regeneration. Bone marrow stromal cells (BMSCs) where seeded using in polymeric and biomimetic (mineralized) scaffolds by two novel seeding techniques, filtration and micromass, and assessed for (a) GJIC, (b) differentiation in three-dimensions and (c) regeneration of bone in-vivo. The results demonstrated significant increases in cell differentiation and regeneration when the alternative seeding techniques and mineralized templates were used over the commonly used static seeding technique and polymer templates, respectively. Both seeding techniques demonstrated increased GJIC over the statically seeded analogue.

To assess specifically for the effects of GJIC without the interference of other factors, BMSCs were transduced with a lentivirus containing the Connexin43 gene, the most prominent gap junction protein in bone. The results showed that transduced cells expressed significantly higher levels of Connexin43, had higher GJIC, expressed differentiation markers at higher levels throughout 3D cell cultures and regenerated larger volumes of spatially uniform tissue. GJIC and differentiation were significantly greater at the core regions of the scaffold, suggesting an important role for GJIC in overcoming compromised regeneration in the core of tissue engineered equivalents.

This research, therefore, presents a novel platform to obtain full 3D equivalents of engineered tissue that were previously unachievable. These experiments also imply that both targeted delivery and cell-based strategies can be used as treatments to enhance communication in 3D living tissue.