A Versatile Microfluidic Extrusion-Based Hydrogel Platform for Self-Organization and Long-Term Maintenance of...

Area of Science:

• Biomedical Engineering
• Cell Biology
• Vascular Biology

Background:

• Lymphatic endothelium is crucial for fluid balance and immune function.
• Existing in vitro models lack the complex 3D structure and long-term stability of lymphatic vessels.

Purpose of the Study:

• To develop a novel microfluidic platform for generating 3D human lymphatic endothelium.
• To create a stable, long-term in vitro model that mimics lymphatic vessel architecture and function.

Main Methods:

• Utilized coaxial extrusion to create size-tunable 3D tubular constructs from primary human lymphatic endothelial cells and a specialized hydrogel.
• Cultured engineered lymphatic tubes for 30 days to assess viability, marker expression, barrier function, and cellular remodeling.
• Performed RNA sequencing to compare gene expression profiles of 3D lymphatic tubes and 2D monolayers.

Main Results:

• Successfully generated 3D lymphatic endothelial tubes with tunable diameters (50-300 µm) that self-assembled into lumen-forming monolayers within one week.
• Demonstrated long-term viability, sustained lymphatic marker expression, and selective macromolecular permeability for at least 30 days.
• Observed cellular remodeling of the hydrogel core into a fibronectin-rich perivascular zone and identified enrichment of lymphatic morphogenesis and maturation markers in 3D tubes compared to 2D cultures.

Conclusions:

• The microfluidic platform provides a robust and tunable system for engineering 3D lymphatic endothelium.
• This model system enables detailed investigation of lymphatic structure-function relationships and holds potential for studying lymphatic diseases.