Environmental Specification of Pluripotent Stem Cell Derived Endothelial Cells Toward Arterial and Venous Subtypes

Abstract

Endothelial cells (ECs) are required for a multitude of cardiovascular clinical applications, such as revascularization of ischemic tissues or endothelialization of tissue engineered grafts. Patient derived primary ECs are limited in number, have donor variabilities and their in vitro phenotypes and functions can deteriorate over time. This necessitates the exploration of alternative EC sources. Although there has been a recent surge in the use of pluripotent stem cell derived endothelial cells (PSC-ECs) for various cardiovascular clinical applications, current differentiation protocols yield a heterogeneous EC population, where their specification into arterial or venous subtypes is undefined. Since arterial and venous ECs are phenotypically and functionally different, inappropriate matching of exogenous ECs to host sites can potentially affect clinical efficacy, as exemplified by venous graft mismatch when placed into an arterial environment. Therefore, there is a need to design and employ environmental cues that can effectively modulate PSC-ECs into a more homogeneous arterial or venous phenotype for better adaptation to the host environment, which will in turn contribute to better application efficacy. In this review, we will first give an overview of the developmental and functional differences between arterial and venous ECs. This provides the foundation for our subsequent discussion on the different bioengineering strategies that have been investigated to varying extent in providing biochemical and biophysical environmental cues to mature PSC-ECs into arterial or venous subtypes. The ability to efficiently leverage on a combination of biochemical and biophysical environmental cues to modulate intrinsic arterio-venous specification programs in ECs will greatly facilitate future translational applications of PSC-ECs. Since the development and maintenance of arterial and venous ECs in vivo occur in disparate physio-chemical microenvironments, it is conceivable that the application of these environmental factors in customized combinations or magnitudes can be used to selectively mature PSC-ECs into an arterial or venous subtype.

Surface markers: An identity card of endothelial cells

“…For many years, it was thought that blood flow was responsible for establishing vascular identity. The discovery that members of the Eph family are differentially expressed in arteries and veins from very early stages of development, prior to development of a functional circulation, was one of the first indications that artery‐vein identity is intrinsically programmed . Arterial and venous specification in the developing embryo can be genetically pre‐determined.…”

Effects of shear stress on differentiation of stem cells into endothelial cells

“…Further studies investigated whether EPCs differentiate into arterial or venous ECs in response to shear stress[ 3 ]. Obi et al [ 41 ] have demonstrated that controlled levels of shear stress in a flow-loading device increased the expression levels of the arterial EC markers ephrinB2, Notch1/3, Hey1/2, and activin receptor-like kinase 1, but decreased expression of the venous markers EphB4 and neuropilin 2.…”

“…Most of the previous experimental systems involved the use of bioreactors or viscometers, like parallel plate flow and conical flow reactors[ 3 ]. To overcomes the limitations of macroperfusion systems in shear application throughput and precision, Toh and Voldman[ 54 ] used a multiplex microfluidic array that applied shear stresses varying by > 1000 times (1.6 × 10 -7 -1.6 × 10 -4 N/cm 2 ) to mouse ESCs.…”

“…Endothelial cells (ECs) derived from donors or differentiated from stem cells are required for various clinical applications, such as promoting angiogenesis in ischemic areas or re-endothelialization of tissue-engineered grafts. Patient-derived primary ECs are limited in number, have donor variabilities, and their in vitro phenotypes and functions can deteriorate over time[ 3 ]. This necessitates the exploration of alternative EC sources.…”

Selection of different endothelialization modes and different seed cells for tissue-engineered vascular graft

“…There are significant differences in morphology between arterial and venous endothelium [ 182 , 183 ]. Arterial ECs are long and narrow or oval, while venous ECs are short and wide, which is related to the fact that the velocity of blood flow in the venous circulation is significantly lower than that in the arterial circulation.…”