Perfused 3D angiogenic sprouting in a high-throughput in vitro platform.

Area of Science:

• Biomedical Engineering
• Cell Biology
• Vascular Biology

Background:

• Current in vitro models lack key microenvironmental cues like perfusion and stable gradients essential for studying angiogenic sprouting.
• Angiogenic sprouting is a complex process involving new blood vessel growth from existing ones, regulated by biochemical and physical factors.

Purpose of the Study:

• To develop and validate an in vitro platform that integrates perfusion and stable biomolecular gradients for studying physiologically relevant angiogenic sprouting.
• To investigate the role of specific factors and perfusion in microvascular stabilization and maturation.

Main Methods:

• A microfluidic platform with 40 individually addressable units was designed to culture perfused microvessels within a 3D collagen-1 matrix.
• Pro-angiogenic factors, including vascular endothelial growth factor-165 (VEGF-165), phorbol 12-myristate 13-acetate (PMA), and sphingosine-1-phosphate (S1P), were used to induce angiogenesis.
• Perfusion with FITC-Dextran was used to assess microvessel leakiness and maturation.

Main Results:

• The platform successfully induced endothelial cells to form tip and stalk cells, leading to matrix invasion and lumen formation.
• A combination of VEGF-165, PMA, and S1P demonstrated optimal pro-angiogenic effects, with S1P crucial for sprout guidance and repetitive formation.
• Anastomosis of angiogenic sprouts with other perfusion channels led to stabilization and maturation, evidenced by reduced leakiness.

Conclusions:

• The developed platform provides a more physiologically relevant in vitro model for studying angiogenic sprouting and microvascular stabilization.
• Perfusion acts as a critical factor for the survival, maturation, and stabilization of newly formed blood vessels.
• This platform offers a robust tool for in-depth investigation of angiogenesis in a controlled 3D microenvironment.