Promotion of Vascular Morphogenesis of Endothelial Cells Co-Cultured with Human Adipose-Derived Mesenchymal Stem Cells Using Polycaprolactone/Gelatin Nanofibrous Scaffolds.

Yun-Min Kook,Kangwon Lee,Hyerim Kim,Sujin Kim,Min Hee Park,Chan Yeong Heo,Won-Gun Koh

doi:10.3390/nano8020117

Abstract

New blood vessel formation is essential for tissue regeneration to deliver oxygen and nutrients and to maintain tissue metabolism. In the field of tissue engineering, in vitro fabrication of new artificial vessels has been a longstanding challenge. Here we developed a technique to reconstruct a microvascular system using a polycaprolactone (PCL)/gelatin nanofibrous structure and a co-culture system. Using a simple electrospinning process, we fabricated three-dimensional mesh scaffolds to support the sprouting of human umbilical vein endothelial cells (HUVECs) along the electrospun nanofiber. The co-culture with adipose-derived mesenchymal stem cells (ADSCs) supported greater sprouting of endothelial cells (ECs). In a two-dimensional culture system, angiogenic cell assembly produced more effective direct intercellular interactions and paracrine signaling from ADSCs to assist in the vascular formation of ECs, compared to the influence of growth factor. Although vascular endothelial growth factor and sphingosine-1-phosphate were present during the culture period, the presence of ADSCs was the most important factor for the construction of a cell-assembled structure in the two-dimensional culture system. On the contrary, HUVECs co-cultured on PCL/gelatin nanofiber scaffolds produced mature and functional microvessel and luminal structures with a greater expression of vascular markers, including platelet endothelial cell adhesion molecule-1 and podocalyxin. Furthermore, both angiogenic factors and cellular interactions with ADSCs through direct contact and paracrine molecules contributed to the formation of enhanced engineered blood vessel structures. It is expected that the co-culture system of HUVECs and ADSCs on bioengineered PCL/gelatin nanofibrous scaffolds will promote robust and functional microvessel structures and will be valuable for the regeneration of tissue with restored blood vessels.

Highlights

New blood vessel formation is crucial to supplying nutrients and oxygen to cells and to remove waste products during the engineering of new tissues
The morphology of co-cultured human umbilical vein endothelial cells (HUVECs) was significantly different in terms of cell length and aspect ratio compared to HUVECs cultured alone at all experimental days regardless of the presence or absence of growth factors (Figure 4E,F). These findings indicated that the growth factors did not affect vascular structure formation and that co-culture of adipose-derived mesenchymal stem cells (ADSCs) only promoted the assembly of HUVECs into the periphery of stem cells in the 2D culture system
Quantitative cell length and aspect ratio measurements revealed that the addition of vascular endothelial growth factor (VEGF) was associated with more extensive cell assembly and elongation than other groups in both the monoculture and co-culture groups (Figure 5E,F). These results indicated that when cells are cultured in the nanofibrous environment, co-culturing with ADSCs and growth factors is effective in enhancing the blood-vessel-like structure formation of HUVECs

Summary

Introduction

New blood vessel formation is crucial to supplying nutrients and oxygen to cells and to remove waste products during the engineering of new tissues. The collective data indicate the capability of co-cultures with MSCs and ECs in the development of blood vessels in vivo after cell transplantation Both cell types are appealing sources for generating functional vascular networks for tissue regeneration applications [16]. One of the MSCs, adipose-derived mesenchymal stem cells (ADSCs) have emerged as a novel source of neovascularization that secretes various angiogenic factors (including VEGF, angiopoietin, and a variety of microRNAs) more than bone-marrow-derived MSCs [17,18]. This finding suggests that ADSCs may be a powerful candidate for blood vessel regeneration

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