(J-364) Engineering a human endothelium platform for disease modeling

Endothelial tissue is a semipermeable barrier that lines blood vessels and regulates the transport of circulating moieties, cells, and biomolecules between the vascular and extravascular compartments. Altered molecular and structural properties of this vascular barrier often precede several pathological conditions, leading to aberrant transport of signaling macromolecules and cells. The exact mechanisms triggering a pathological decline in the vascular barrier properties, and its contributions to disease progression remain ill-defined predominantly due to the lack of a relevant model system for investigation. We have engineered an acrylic-based human isogenic biomimetic platform. The platform consists of individually addressable top and bottom compartments separated by a transwell membrane with endothelium on one side and tissue-specific cells (i.e. pancreatic islets) on the other side. Our study models the interactions of immune cells and biomolecules with an endothelium-derived from  type-1-diabetic patient vs healthy donor.

The platform is fabricated in two parts: the top and the bottom using CNC milling from an acrylic sheet, which interlock with each other. The top part holds the silicone-resin-based gasket which forms a fluidic seal when both parts are interlocked and closed using a commercially available mechanical clamp. Our method entails maturation of hiPSC-derived endothelial barrier onto a static transwell-based culture and confirming the establishment through (transendothelial electrical resistance) TEER values. A plateau in TEER values indicates the establishment of a mature barrier. The mature barrier is then transferred into our engineered platform reversibly to simulate vascular vessels. The transwell membrane is then transferred to the chip reversibly. To recapitulate physiological hemodynamics a differential pressure will be generated via pressure-driven flow across the vascular and extravascular compartments. The isogeneity of the system will be attained by differentiating human induced pluripotent stem cells (hiPSC) from the same donor into endothelial cells and T cells sourced from donor blood.

Result and discussion:
The barrier properties of the endothelium in the platform were characterized by measuring hydraulic conductivity, permeability as a function of solute size, charge (using FITC-dextran) and type (protein/non-protein entity), expression of junctional proteins (VE-cadherin). A healthy endothelium was compared with a diseased endothelium to evaluate the changes in barrier properties in pathological conditions. The endothelial barrier thus formed was maintained in chip culture for five days. FITC-dextran-based permeability assay showed an increase in permeability in the presence of diseased vs healthy endothelium. The immune cells extravasation increased in response to target antigens in the presence of a diseased endothelium as compared to the healthy endothelium, suggesting a dysfunctional endothelial barrier.

Conclusion:
This platform will have the ability to i) study the interactions between the tissue-specific barrier and selected circulating moieties with disease-relevant cellular genotypes, and ii) investigate the transendothelial transport dynamics spatiotemporally. Additionally, it can be utilized as a device for testing drugs targeted to endothelial cell receptors that interact with immune cells.

The research has been funded by NIDDK(UH3DK122638)