Vascular Morphogenesis of Human Umbilical Vein Endothelial Cells on Cell-Derived Macromolecular Matrix Microenvironment

Abstract

Extracellular matrix (ECM) is a highly organized network of proteins and other macromolecules that plays a critical role in cell adhesion, migration, and differentiation. In this study, we hypothesize that ECM derived from in-vitro-cultured cells possesses unique surface texture, topography, and mechanical property, and consequently carries some distinct cues for vascular morphogenesis of human umbilical vein endothelial cells (ECs). Cell-derived matrix (CDM) was obtained by culturing fibroblasts, preosteoblasts, and chondrocytes, respectively, on coverslips and then by decellularizing them using detergents and enzymes. These matrices were named fibroblast-derived matrix (FDM), preosteoblast-derived matrix (PDM), and chondrocyte-derived matrix (CHDM). Immunofluorescence of each CDM shows that some of the matrix components are fibronectin (FN), type I collagen, and laminin. Atomic force microscopy analysis presented that average fiber diameter ranged from 2 to 7 mm and FDM holds much larger fibers. The matrix elasticity measurements revealed that average Young's modulus of CHDM (17.7 -4.2 kPa) was much greater than that of PDM (10.5 -1.1 kPa) or FDM (5.7 -0.5 kPa). During 5-day culture, EC morphologies were dramatically changed on PDM and FDM, but those on CHDM and gelatin were rather stable, regardless of time lapse. Cell migration assay discovered quicker repopulation of the scratched areas on PDM and FDM than on gelatin and CHDM. A capillary-like structure (CLS) assembly was also notable only in the PDM and FDM, as compared with CHDM, gelatin, or FN that were very poor in CLS formation. Quantitative analysis of mean CLS branch points and branch lengths demonstrated much better angiogenic activity of ECs on PDM and FDM. Interestingly, CLS formation was closely associated with matrix remodeling by ECs and the matrix clearance on PDM with time was sharply contrasted with that on CHDM that majority of the matrix FN was reserved. It was notable that membrane type 1-matrix metalloprotease was deeply involved in the process of matrix remodeling. This study indicates that specific matrix microenvironments are very critical for vascular morphogenesis of ECs, and thus, provide a nice platform for angiogenesis study as well as vascular tissue engineering.

Human vascular model with defined stimulation medium – a characterization study

“…The electron microscopic analysis as well as immunocytochemistry results of this study showed that hASC and HUVEC co-culture actively produces extracellular matrix (ECM) components, including fibrillins, thus creating natural 3D scaffold around them. The reciprocal interaction between ECM stroma and vascular network is important in directing vessel growth (Hoying et al, 2014;Du et al, 2014). The hASC and HUVEC co-culture gives mechanical support for other target cells, e.g., cardiomyocytes, and, additionally, the microenvironment formed by the co-culture enhances target cell viability as reported previously (Vuorenpää et al, 2014).…”

Cardiomyoblast (H9c2) Differentiation on Tunable Extracellular Matrix Microenvironment

“…22 In another study using human umbilical vein endothelial cells, both FDM and preosteoblast-derived matrix were excellent in forming an assembly of capillary-like structure and in inducing a significant matrix remodeling for vasculogenesis. 23 Those experiences have prompted us to explore the potential of FDM as a novel culture platform for H9c2 cardiomyoblasts; a cell line isolated from embryonic rat heart tissue described in length by Hescheler et al 16 In fact, many studies have shown that ECM plays a significant role in cardiomyocyte differentiation. 5,24,25 Hence, in this study, we have investigated the benefits of FDM in cellular behaviors of H9c2 cardiomyoblasts, primarily focusing on their differentiation into cardiomyogenic cells.…”

Pivotal role for decorin in angiogenesis

“…Indeed, today we know that the structure of the ECM in itself has a great impact on angiogenesis via directly or indirectly regulating endothelial cell (EC) behavior [8–11]. Angiogenesis requires the generation of “activated migratory” ECs (tip cells) which guide the developing vascular sprout [12–15].…”