Abstract
IntroductionRapid establishment of functional blood vessels is a prerequisite for successful tissue engineering. During vascular development, endothelial cells (ECs) and perivascular cells assemble into a complex regulating proliferation of ECs, vessel diameter and production of extracellular matrix proteins. The aim of this study was to evaluate the ability of mesenchymal stem cells (MSCs) to establish an endothelial-perivascular complex in tissue-engineered constructs comprising ECs and MSCs.MethodsPrimary human ECs and MSCs were seeded onto poly(L-lactide-co-1,5-dioxepan-2-one) (poly(LLA-co-DXO)) scaffolds and grown in dynamic culture before subcutaneous implantation in immunocompromised mice for 1 and 3 weeks. Cellular activity, angiogenic stimulation and vascular assembly in cell/scaffold constructs seeded with ECs or ECs/MSCs in a 5:1 ratio was monitored with real-time RT-PCR, ELISA and immunohistochemical microscopy analysis.ResultsA quiescent phenotype of ECs was generated, by adding MSCs to the culture system. Decreased proliferation of ECs, in addition to up-regulation of selected markers for vascular maturation was demonstrated. Baseline expression of VEGFa was higher for MSCs compared with EC (P <0.001), with subsequent up-regulated VEGFa-expression for EC/MSC constructs before (P <0.05) and after implantation (P <0.01). Furthermore, an inflammatory response with CD11b + cells was generated from implantation of human cells. At the end of the 3 week experimental period, a higher vascular density was shown for both cellular constructs compared with empty control scaffolds (P <0.01), with the highest density of capillaries being generated in constructs comprising both ECs and MSCs.ConclusionsInduction of a quiescent phenotype of ECs associated with vascular maturation can be achieved by co-seeding with MSCs. Hence, MSCs can be appropriate perivascular cells for tissue-engineered constructs.
Highlights
Rapid establishment of functional blood vessels is a prerequisite for successful tissue engineering
Mesenchymal stem cells induce endothelial cell quiescence The expression of endothelial specific ANGs was altered by the addition of mesenchymal stem cells (MSCs) to the culture system (Figure 1A)
Cell proliferation was assessed through expression of proliferating cell nuclear antigen (PCNA) and ki67, both being downregulated in endothelial cells (ECs)/MSC constructs relative to the monocultured ECs (Figure 1B)
Summary
Rapid establishment of functional blood vessels is a prerequisite for successful tissue engineering. Milestone discoveries in the study of MSCs are the fibroblastic colony-forming units initiated by single cells [2], the multilineage potential [3] and, more recently, a shift in view towards MSCs as tissue-specific progenitors residing in close proximity to the microvasculature of various organs [4]. The latter might serve as a rationale for MSCs as a potential perivascular cell in prevascularized constructs for tissue regeneration
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