A New Versatile Platform for Assessment of Improved Cardiac Performance in Human-Engineered Heart Tissues

Abstract

Cardiomyocytes derived from human pluripotent stem cells (hPSC-CMs) hold a great potential as human in vitro models for studying heart disease and for drug safety screening. Nevertheless, their associated immaturity relative to the adult myocardium limits their utility in cardiac research. In this study, we describe the development of a platform for generating three-dimensional engineered heart tissues (EHTs) from hPSC-CMs for the measurement of force while under mechanical and electrical stimulation. The modular and versatile EHT platform presented here allows for the formation of three tissues per well in a 12-well plate format, resulting in 36 tissues per plate. We compared the functional performance of EHTs and their histology in three different media and demonstrated that tissues cultured and maintained in maturation medium, containing triiodothyronine (T3), dexamethasone, and insulin-like growth factor-1 (TDI), resulted in a higher force of contraction, sarcomeric organization and alignment, and a higher and lower inotropic response to isoproterenol and nifedipine, respectively. Moreover, in this study, we highlight the importance of integrating a serum-free maturation medium in the EHT platform, making it a suitable tool for cardiovascular research, disease modeling, and preclinical drug testing.

Automated assessment of human engineered heart tissues using deep learning and template matching for segmentation and tracking

“…The contraction trajectory created by calculating the distances between the brighter points detected on the left and right pillars showed a square contraction wave (Figure 2e) (Equations 4 and 5). It is worthy to note that this is not the expected smooth contraction wave according to literature 35–37 …”

Contractility analysis of human engineered 3D heart tissues by an automatic tracking technique using a standalone application

“…Initially, to highlight the pillars tip location and simplify the tracking technic, the transparent PDMS tip of the pillars were painted using carbon black [ 25 ]. The carbon black allows to achieve a high level of blackness and reduces the light that passes through the material, resulting in clear black circles in the image ( Fig 4A ).…”

“…Therefore, we have applied this image analysis software to study this biological question. We successfully made EHTs from the hPSC line (NKX2.5 EGFP/+- COUP-TFII mCherry/+ ) with different ratios of HCF (0%, 1%, 2%, 3%, 5% and 10%), by using our previously developed platform [ 25 ]. First, we evaluated the success rate of tissue formation over time, which is the homogenous formation of the 3D cardiac tissue around the pillars without any gap of cells in the middle ( S3 Fig ).…”

“…Cardiac EHTs were made as previously described from Ribeiro et al [ 25 ]. Briefly, three tissues were made per well in a 12 well plate format with five different ratios of HCF: 0%, 1%, 3%, 5% and 10%.…”

“…Subsequently, cells were mixed with an extracellular matrix (ECM) mixture consisting of 2X MM medium, fibrinogen (final concentration 2 mg/mL, Sigma-Aldrich F8630), Matrigel (final concentration 1 mg/mL) and aprotinin (final concentration 2.5 μg/mL, Sigma-Aldrich, A1153). Then, 0.6 U/mL of thrombin (Sigma, T7513) was added to the mix and quickly after mixing, 15 μl was used to make each one of the three tissues per well [ 25 ]. After 24 hours, the first refreshment was done and after that, the refreshments were done every 2 or 3 days and after contraction measurements.…”