Generation and characterisation of scalable and stable human pluripotent stem cell-derived microvascular-like...

Area of Science:

• Cardiovascular Biology
• Stem Cell Biology
• Vascular Biology

Background:

• Coronary microvascular disease (CMD) is a significant health challenge, necessitating accurate in vitro models for investigation.
• Current human pluripotent stem cell-derived endothelial cells (hPSC-ECs) lack scalability and microvascular specificity for CMD research.
• Developing phenotypically stable, cardiac-specific microvascular cells is crucial for advancing CMD understanding.

Purpose of the Study:

• To develop a scalable 3D protocol for generating phenotypically stable cardiac microvascular-like endothelial cells (hPSC-CMVECs) and cardiac pericyte-like cells.
• To characterize these generated cells for their suitability as a robust in vitro model for CMD.
• To explore the potential applications of these cells in cardiac tissue engineering.

Main Methods:

• Generation of vascular organoids from human pluripotent stem cells (hPSCs) in 3D stirred tank bioreactors.
• Treatment of emerging 3D hPSC-ECs with high-concentration VEGF-A (3DV) to promote phenotypic stability and microvascular characteristics.
• Comprehensive characterization using single-cell RNA sequencing (scRNA-seq) and marker analysis.

Main Results:

• The 3D stirred tank bioreactor protocol with VEGF-A treatment yielded phenotypically stable 3DV hPSC-ECs.
• scRNA-seq confirmed pronounced expression of cardiac endothelial and microvascular genes in 3DV hPSC-ECs.
• Generated mural cells displayed markers characteristic of cardiac pericytes, forming a suitable cell model for CMD.

Conclusions:

• A scalable 3D protocol successfully generated phenotypically stable hPSC-CMVECs and cardiac pericytes.
• This novel cell model is suitable for investigating CMD mechanisms and cardiac microvasculature.
• The cells' stability, cardiac specificity, and angiogenic potential make them valuable for cardiac tissue engineering.