Enhancement of collagen deposition and cross-linking by coupling lysyl oxidase with bone morphogenetic protein-1 and its application in tissue engineering

Cultured cell-derived extracellular matrices (ECM)-based biomaterials exploit the inherent capacity of cells to create highly sophisticated supramolecular assemblies. However, standard cell culture conditions are far from ideal given the fact that the diluted microenvironment does not favor the production of ECM components, a circumstance particularly relevant for collagen. An incomplete conversion of procollagen by C-proteinase/bone morphogenetic protein 1 (BMP1) has been proposed to severely limit in vitro collagen deposition. BMP1 also catalyzes the proteolytic activation of the precursor of the collagen cross-linking enzyme, lysyl oxidase (LOX) to yield the active form, suggesting a deficit in cross-linking activity under standard conditions. We hypothesized that the implementation of fibroblast cultures with LOX and BMP1 may be an effective way to increase collagen deposition. To test it, we have generated stable cell lines overexpressing LOX and BMP1 and studied the effect of supernatants enriched in LOX and BMP1 on collagen synthesis and deposition from fibroblasts. Herein, we demonstrate that the supplementation with LOX and BMP1 strongly increased the deposition of collagen onto the insoluble matrix at the expense of the soluble fraction in the extracellular medium. Using decellularization protocols, we also show that fibroblast-derived matrices regulate adipogenic and osteogenic differentiation of human mesenchymal stem cells (MSC), and this effect was modulated by LOX/BMP1. Collectively, these data demonstrate that we have developed a convenient protocol to enhance the capacity of in vitro cell cultures to deposit collagen in the ECM, representing this approach a promising technology for application in tissue engineeringTis work was supported by grants from Ministerio de Economía y Competitividad (Plan Nacional de I+D+I: SAF2012-34916, and SAF2015-65679-R to F.R-P