Human dermal microvascular arterial and venous blood endothelial cells and their use in bioengineered dermo-epidermal skin substitutes in vitro and in vivo

The bio-engineering of vascular networks is pivotal to create complex tissues and

organs in vitro for regenerative medicine applications. The vascular plexus is needed for a

sufficient and fast blood supply after transplantation, and, thus, required for the survival and

function of the engineered tissue or organ. Hence, human endothelial cells are an attractive

source for bio-engineering purposes, for example human dermal microvascular endothelial

cells (HDMECs).

So far, a discrimination between arterial and venous blood endothelial cells after

isolation of HDMECs from skin biopsies and if arterial and/or venous capillaries are formed in

pre-vascularized bio-engineered substitutes was not investigated.

In this study, we investigated employedby single cell sequencing for to

investigate/compare human arterial and venous endothelial cell markers in human fetal and

juvenile skin. Further, we analyzed if these markers are present after isolation of human skin

derived endothelial cells under 2D culture conditions. In additionFinally, we investigated

assessed if human endothelial cells form distinct arterial and venous capillaries in 3D

collagen type I hydrogels, and if these capillaries retain their identity after transplantation.

We determinedOur results showed that arterial and venous endothelial cell markers

such as NRP1 and NR2F2 are expressed both in fetal and juvenile skin, and are retained after

isolation in culture. We could show demonstrate that arterial and venous endothelial cells

maintain their differentiation status and form arterial and venous capillaries in 3D in vitro

culture systems and that the capillaries inosculate after transplantation.

In summary, we could show that we could bio-engineer human arterial, venous, and

lymphatic capillaries in a human skin substitute in view of regenerative medicine approaches

for clinical applications.