Cardiomyocyte Differentiation via Microcarrier-Based Techniques for hiPSC Culture
Board Location: #20
Discipline: Biological Sciences
Subcategory: Biomedical Engineering
Session: 4
Jacob Cagan - Arizona State University
Co-Author(s): Carlos Barreto, University of Puerto Rico Mayagüez Mara Domenech, University of Puerto Rico Mayagüez
Cardiovascular disease (CVD) accounts for over 30% of global deaths, underscoring the urgent need for innovative therapies. Stem cell therapy has emerged as a promising approach; however, scalable and reliable differentiation methods remain a critical bottleneck. This study aimed to evaluate the differentiation of human induced pluripotent stem cells (hiPSCs) into cardiomyocytes using microcarriers (μCs) under static conditions, as a step toward scalable suspension cultures. Protocols optimized for planar cultures were adapted to μCs, focusing on the effects of CHIR99021, a GSK-3 inhibitor and Wnt pathway activator, and cell density on differentiation outcomes.
hiPSCs (WTC-11) were seeded at varying CHIR99021 concentrations (5 μM, 7.5 μM, and 10 μM) for both planar Matrigel-coated surfaces (control group) and Collagen I μCs coated with Vitronectin (test group) in ultra-low attachment 48-well plates. Microcarrier surface areas were matched to Matrigel well surface areas (1.1 cm²) to ensure comparable conditions. Flow cytometry was used to assess the expression of key biomarkers, including OCT4 (pluripotency), Nkx2.5 (early cardiac differentiation), cTnT (cardiomyocyte differentiation), and cTnI (cardiomyocyte maturation).
The μC experiments revealed no statistically significant differences in Nkx2.5 expression among the CHIR99021 concentrations. However, cTnT and cTnI expression were significantly improved with higher CHIR99021 concentrations. These findings indicate that higher CHIR99021 concentrations improve cardiomyocyte differentiation on μCs, highlighting the potential for optimizing suspension-based differentiation protocols. Future work will refine these protocols and evaluate the scalability and functional outcomes of μC-based cardiomyocyte differentiation.
Funder Acknowledgement(s): CMaT, University of Puerto Rico Mayagüez
Faculty Advisor: Carlos Barreto, carlos.barreto7@upr.edu
Role: REU Student