Abstract 14156: Promotion of Human Pluripotent Stem Cell-Engineered Cardiac Tissue Maturation by Non-Physiological Intermittent Hydrostatic Pressure

Background: The immaturity of human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) and hPSC-engineered cardiac tissues (ECTs) hinder their potential for cardiac regenerative therapy. Multiple studies have demonstrated the crucial role of ventricular loading pressure in cardiac repair and regeneration.

Hypothesis: We postulated that hydrostatic pressure could enhance the maturation and functionality of the ECTs.

Methods: To create ECTs, a mixture of hPSC-derived cardiovascular cells was co-cultured with Collagen I and Matrigel, then subjected to hydrostatic pressure. The functionality of the ECTs was assessed using a video-based system to measure contractile capacity, a flux analyzer to evaluate metabolic function, and a calcium indicator. Histological assessments were conducted to examine the structural maturation. CM maturation were evaluated through scRNA-seq. The trained ECTs were transplanted onto a rat myocardial infarction (MI) model followed by echocardiogram (N=3).

Results: The optimized protocol consisting of intermittent pressure (1 hour/day) at 50 kPa (equivalent to 375 mmHg), for a duration of 3 days, preceded by a 2-week pre-culture period, remarkably enhanced the CM component and collagen alignment of the ECTs. This training led to increased expression of various genes associated with CM maturation. The trained ECTs exhibited a positive force-frequency relationship in terms of contractility, along with an elevation in peak calcium flux, indicating functional maturation. The training resulted in increased mitochondrial content and maximum respiration capacity, indicating metabolic maturation. Combination of the hydrostatic training and dynamic culture promoted vascular network formation of the ECTs. The maturation was dependent on the presence of endothelial cells (ECs), as ECTs without ECs failed to exhibit the maturation. The ECTs demonstrated a significant increase in ejection fraction of a rat MI model at 4 weeks post-transplantation (control vs training: 42.8±13.0 vs 80.7±8.5 %, P= 0.0045).

Conclusion: The application of non-physiological and intermittent hydrostatic pressure training has led to significant maturation of ECTs, showcasing their therapeutic potential.