High-Resolution Imaging and Assessment of Vascular Network in Tri-Lineage hiPSC-Derived Cardiac Microtissues

Area of Science:

• Cardiovascular Biology
• Stem Cell Biology
• Biomedical Engineering

Background:

• Cardiac microtissues (cMTs) are 3D models mimicking native heart tissue composition.
• Induced pluripotent stem cells (iPSCs) enable the generation of cardiomyocytes, endothelial cells, and cardiac fibroblasts for cMTs.
• Endothelial cells in cMTs promote microvascular network formation, enhancing tissue function and disease modeling relevance.

Purpose of the Study:

• To present detailed protocols for generating scaffold-free, vascularized cMTs from human iPSCs.
• To outline methods for high-resolution imaging and analysis of cMTs, focusing on vascular-like structures.
• To provide a framework for studying cell interactions and microenvironment in engineered cardiac tissues.

Main Methods:

• Aggregation of pre-differentiated iPSC-derived cardiomyocytes, endothelial cells, and cardiac fibroblasts.
• Fluorescence labeling of whole-mount and sectioned cMTs.
• Confocal microscopy for image acquisition, visualization, and 3D analysis using ImageJ (FIJI).

Main Results:

• Successful generation of tri-lineage vascularized cMTs from human iPSCs.
• Detailed visualization and characterization of spontaneous microvascular-like networks within cMTs.
• Quantification of capillary-like networks using ImageJ software with 3D modeling.

Conclusions:

• The presented protocols facilitate the creation and examination of vascularized cMTs.
• These models offer valuable insights into cardiac cell interactions and microenvironment.
• Vascularized cMTs are promising tools for cardiac disease modeling and drug screening.