Integrating Vascular Phenotypic and Proteomic Analysis in an Open Microfluidic Platform.

Area of Science:

• Vascular Biology
• Proteomics
• Drug Discovery

Background:

• Angiogenesis, the formation of new blood vessels, is crucial for development and disease.
• Understanding the molecular mechanisms and drug effects on angiogenesis is vital for therapeutic development.
• Current methods for analyzing angiogenesis lack high-throughput capabilities and integrated morphological and proteomic analysis.

Purpose of the Study:

• To introduce and validate a novel vascular phenotypic and proteomic analysis (VPT) platform for high-throughput studies of vascular development.
• To investigate the effects of specific anti-angiogenic agents on endothelial cell behavior and protein expression.
• To elucidate the relationship between cytoskeletal integrity, protein expression changes, and angiogenic responses.

Main Methods:

• Development of a vascular phenotypic and proteomic analysis (VPT) platform with an open-channel configuration for endothelial cell alignment.
• High-throughput screening of anti-angiogenic agents (bevacizumab, ramucirumab, cabozantinib, regorafenib, wortmannin, chloroquine, paclitaxel) on vascular sprouting.
• Quantitative proteomic analysis using LC-MS/MS to identify global protein expression changes in response to drug treatment.

Main Results:

• The VPT platform facilitates 3D morphological examination and protein analysis of vascular development.
• Anti-angiogenic agents caused an approximate 50% reduction in angiogenic sprouting at higher concentrations.
• LC-MS/MS analysis identified significant protein expression changes related to cell cycle, cytoskeleton, senescence, and angiogenesis pathways in response to four drugs.

Conclusions:

• The VPT platform provides a powerful tool for analyzing drug impacts on vascular biology, integrating morphological and proteomic data.
• Cytoskeletal integrity is intricately linked to angiogenic responses, highlighting the importance of combined analytical approaches.
• The VPT platform is versatile for studying proteome and morphology in various biological models beyond blood vessels.