A Rapid-Patterning 3D Vessel-on-Chip for Imaging and Quantitatively Analyzing Cell-Cell Junction Phenotypes.

Area of Science:

• Neuroscience
• Biotechnology
• Cell Biology

Background:

• The blood-brain barrier (BBB) controls molecular traffic between the brain and blood.
• BBB permeability relies on junction proteins in brain endothelial cells.
• Existing in vitro BBB models lack quantitative 3D microvessel junction analysis.

Purpose of the Study:

• To create a simple 3D vessel-on-chip model for quantitative analysis of junction presentation in microvessels.
• To investigate how 3D microvessel structures influence endothelial cell junction phenotypes.
• To evaluate the model's utility in assessing BBB integrity under different conditions.

Main Methods:

• Fabrication of 3D microvessels using polydimethylsiloxane and microneedles.
• Culturing human iPSC-derived brain microvascular endothelial-like cells (iBMEC-like cells) within the 3D structure.
• Developing an image analysis pipeline to 'unwrap' 3D vessels for 2D quantitative analysis of cell-cell junctions.
• Treating the model with tumor necrosis factor α to assess disruption of cell-cell junction integrity.

Main Results:

• 3D cylindrical structures altered the phenotype of tight junction proteins and cell morphology.
• The 3D vessel-on-chip model successfully demonstrated disruption of cell-cell junction integrity upon TNF-α treatment.
• The developed analysis pipeline enabled quantitative evaluation of junction presentation in 3D microvessels.

Conclusions:

• A rapid and straightforward 3D vessel-on-chip model and analysis pipeline were established.
• This model allows for quantitative screening and evaluation of endothelial cell-cell junction integrity.
• The system is suitable for assessing the impact of various microenvironments and treatments on BBB integrity.