Porous honeycomb film membranes enhance endothelial barrier integrity in human vascular wall bilayer model compared to standard track‐etched membranes

Abstract

In vitro vascular wall bilayer models for drug testing and disease modeling must emulate the physical and biological properties of healthy vascular tissue and its endothelial barrier function. Both endothelial cell (EC)‐vascular smooth muscle cell (SMC) interaction across the internal elastic lamina (IEL) and blood vessel stiffness impact endothelial barrier integrity. Polymeric porous track‐etched membranes (TEM) typically represent the IEL in laboratory vascular bilayer models. However, TEM stiffness exceeds that of diseased blood vessels, and the membrane pore architecture limits EC‐SMC interaction. The mechanical properties of compliant honeycomb film (HCF) membranes better simulate the Young's modulus of healthy blood vessels, and HCFs are thinner (4 vs. 10 μm) and more porous (57 vs. 6.5%) than TEMs. We compared endothelial barrier integrity in vascular wall bilayer models with human ECs and SMCs statically cultured on opposite sides of HCFs and TEMs (5 μm pores) for up to 12 days. Highly segregated localization of tight junction (ZO‐1) and adherens junction (VE‐cadherin) proteins and quiescent F‐actin cytoskeletons demonstrated superior and earlier maturation of interendothelial junctions. Quantifying barrier integrity based on transendothelial electrical resistance (TEER), membranes showed only minor but significant TEER differences despite enhanced junctional protein localization on HCF. Elongated ECs on HCF likely experienced greater paracellular diffusion than blocky ECs on TEM. Also, larger populations of plaques of connexin 43 subunit‐containing gap junctions suggested enhanced EC‐SMC communication across the more porous, thinner HCF. Compared with standard TEMs, engineered vascular wall bilayers cultured on HCFs better replicate physiologic endothelial barrier integrity.