Enhanced Osteogenesis in Cocultures with Human Mesenchymal Stem Cells and Endothelial Cells on Polymeric Microfiber Scaffolds

Abstract

In this work, human mesenchymal stem cells (hMSCs) and their osteogenically precultured derivatives were directly cocultured with human umbilical vein endothelial cells (HUVECs) on electrospun three-dimensional poly(e-caprolactone) microfiber scaffolds to evaluate the coculture's effect on the generation of osteogenic constructs. Specifically, cells were cultured on scaffolds for up to 3 weeks, and the cellularity, alkaline phosphatase (ALP) activity, and bone-like matrix formation were assessed. Constructs with cocultures and monocultures had almost identical cellularity after the first week, however, lower cellularity was observed in cocultures compared to monocultures during the subsequent 2 weeks of culture. Scaffolds with cocultures showed a significantly higher ALP activity, glycosaminoglycan and collagen production, as well as greater calcium deposition over the course of study compared to monocultures of hMSCs. Furthermore, the osteogenic outcome was equally robust in cocultures containing osteogenically precultured and non-precultured hMSCs. The results demonstrate that the combination of MSC and HUVEC populations within a porous scaffold material under osteogenic culture conditions is an effective strategy to promote osteogenesis.

Flow Perfusion Co-culture of Human Mesenchymal Stem Cells and Endothelial Cells on Biodegradable Polymer Scaffolds

“…3, 12, 21, 32 Furthermore, previous studies from our laboratories have demonstrated that co-culturing osteogenically precultured hMSCs with HUVECs in a 1:1 ratio under static conditions led to enhanced osteogenic differentiation compared to cultures of hMSCs alone. 9 Both cell populations have previously been shown to be highly responsive to mechanical forces, 1, 16, 22 and the effect of mechanical stimulation on the behavior of endothelial cells and vascularization in co-cultures has been investigated. 18, 24 However, the effect of mechanical forces on the osteogenic differentiation of hMSC/HUVEC co-cultures has not been evaluated.…”

“…9 In order to better recapitulate the bone marrow perivascular niche, the dynamic mechanical environment in vivo should also be considered in an in vitro co-culture system. 13 Flow perfusion bioreactor systems can be used to apply mechanical stresses to three-dimensional cell cultures.…”

Tissue engineering strategies for promoting vascularized bone regeneration

“…166–168 It has been suggested that MSCs are the major producer of VEGF during vascular bone regeneration. 101,167,169 These MSC secreted pro-angiogenic factors stimulate EPC proliferation and differentiation, which in turn causes EPCs to secrete osteogenic growth factors that stimulate MSC differentiation.…”

3D tissue-engineered model of Ewing's sarcoma

“…Unexpectedly, even before a mature vascular network is formed, ECs can affect co-cultured cells and influence the surrounding microenvironment. When mixed with MSCs, for example, ECs enhance osteogenic matrix production within 3D PCL scaffolds [210, 211]. …”

“…A step in this direction, human MSCs, which could serve as a precursor for ES, and human umbilical vein endothelial cells (HUVECs) were co-cultured in 3D PCL-scaffolds under conditions of flow perfusion. Superior proliferation of both cell lines was achieved and the spatial distribution was more uniform throughout 3D-PCL scaffolds [213]. Since VEGF secretion by human ES tumors has been shown to encourage CD38- primitive MSCs to migrate from the bone marrow to the tumor, where they differentiate into ECs and/or pericytes capable of angiogenesis and vasculogenesis [198-201], it would be intriguing to know whether a similar effect could be replicated within a tissue-engineered ES model.…”