Abstract
Mesenchymal stem cells (MSC) have demonstrated efficacy for improving cardiomyocyte (CM) function
in vitro, in vivo
and in clinical trials, but the mechanism of this enhancement remains elusive. The objective of this study was to test the hypothesis that human engineered cardiac tissues (hECT) offer a viable model system to investigate the effects of human MSC on CM contractile function.
Human CM (hCM) were produced from embryonic stem cells (hESC, H7 line) using a small-molecule based differentiation approach. Blebbistatin and BMP4 were added to hESC suspended in StemPro34 differentiation media for 24 h, followed by BMP4 and Activin A to day 4.5, followed by addition of IWR-1 Wnt inhibitor for at least 4 days. To create hECT, approximately 1 million hCM were mixed with 2.0 mg/ml bovine type I collagen and 0.9 mg/ml Matrigel, and pipetted into a mold fabricated from polydimethylsiloxane with integrated cantilever end-posts. To model hMSC cell therapy, two types of hECT were created: hCM-only control hECT, and hMSC-CM hybrid hECT containing hCM mixed with 5-10% of human bone marrow-derived MSC. Over several days in culture, the hECT self-assembled and started beating; end-post deflection was tracked in real time to compute twitch force using beam theory.
Human CMs were produced with high efficiency (>70% cTnT+) with a predominantly ventricular phenotype (MLC2v+). Resulting hECTs exhibited spontaneous beating (1.3±0.4 Hz), cellular alignment, registered sarcomeres, and expression of cardiac specific genes cTnT, α-MHC, β-MHC and SERCA2a. After 11±2 days in culture, developed stress (force/area) was over 10-fold higher in hMSC-CM hybrid tissues (0.27±0.048 mN/mm
2
) compared to hCM-only controls (0.02±0.006 mN/mm
2
; p=0.04, n=5 per group). This reflected significantly greater twitch force (0.11±0.004 mN vs 0.033±0.016 mN, p=0.016) and smaller cross-sectional area (0.19±0.12 mm
2
vs 0.49±0.10 mm
2
; p=0.003) in hMSC-CM hybrid vs hCM-only hECT.
In conclusion, human ECT offer a novel system to study MSC-CM interactions. The findings suggest hMSC supplementation improves contractility compared to CM-only hECT. Investigating the mechanisms of hMSC-mediated enhancement of hECT function may yield insights into MSC-based therapies for cardiac regeneration.