ERRγ agonist under mechanical stretching manifests hypertrophic cardiomyopathy phenotypes of engineered cardiac tissue through maturation.

Engineered cardiac tissue (ECT) using human induced pluripotent stem cell-derived cardiomyocytes is a promising tool for modeling heart disease. However, tissue immaturity makes robust disease modeling difficult. Here, we established a method for modeling hypertrophic cardiomyopathy (HCM) malignant (MYH7 R719Q) and nonmalignant (MYBPC3 G115∗) pathogenic sarcomere gene mutations by accelerating ECT maturation using an ERRγ agonist, T112, and mechanical stretching. ECTs treated with T112 under 10% elongation stimulation exhibited more organized and mature characteristics. Whereas matured ECTs with the MYH7 R719Q mutation showed broad HCM phenotypes, including hypertrophy, hypercontraction, diastolic dysfunction, myofibril misalignment, fibrotic change, and glycolytic activation, matured MYBPC3 G115∗ ECTs displayed limited phenotypes, which were primarily observed only under our new maturation protocol (i.e., hypertrophy). Altogether, ERRγ activation combined with mechanical stimulation enhanced ECT maturation, leading to a more accurate manifestation of HCM phenotypes, including non-cardiomyocyte activation, consistent with clinical observations.

A Method for Contraction Force Measurement of hiPSC-Derived Engineered Cardiac Tissues.

Engineered cardiac tissue (ECT) derived from human induced pluripotent stem cells (iPSCs) can replicate human heart in vitro and be applied to drug discovery and heart disease models. The contraction force of ECT is an important indicator of its function and of the disease phenotype. Here we describe a construction method of ECT using the Flexcell® Tissue Train® culture system and a contraction force measurement method based on the Frank-Starling law.

ERRγ enhances cardiac maturation with T-tubule formation in human iPSC-derived cardiomyocytes.

One of the earliest maturation steps in cardiomyocytes (CMs) is the sarcomere protein isoform switch between TNNI1 and TNNI3 (fetal and neonatal/adult troponin I). Here, we generate human induced pluripotent stem cells (hiPSCs) carrying a TNNI1EmGFP and TNNI3mCherry double reporter to monitor and isolate mature sub-populations during cardiac differentiation. Extensive drug screening identifies two compounds, an estrogen-related receptor gamma (ERRγ) agonist and an S-phase kinase-associated protein 2 inhibitor, that enhances cardiac maturation and a significant change to TNNI3 expression. Expression, morphological, functional, and molecular analyses indicate that hiPSC-CMs treated with the ERRγ agonist show a larger cell size, longer sarcomere length, the presence of transverse tubules, and enhanced metabolic function and contractile and electrical properties. Here, we show that ERRγ-treated hiPSC-CMs have a mature cellular property consistent with neonatal CMs and are useful for disease modeling and regenerative medicine.

Identification and Characterization of a New Series of Ghrelin O-Acyl Transferase Inhibitors.

Ghrelin O-acyl transferase (GOAT; MBOAT4) catalyzes O-acylation at serine-3 of des-acyl ghrelin. Acyl ghrelin is secreted by stomach X/A-like cells and plays a role in appetite and metabolism. Therefore, GOAT has been expected to be a novel antiobesity target because it is responsible for acyl ghrelin production. Here, we report homogeneous time-resolved fluorescence (HTRF) and enzyme-linked immunosorbent assay (ELISA) methods utilizing human GOAT-expressing microsomes as a novel high-throughput assay system for the discovery of hit compounds and optimization of lead compounds. Hit compounds exemplified by compound A (2-[(2,4-dichlorobenzyl)sulfanyl]-1,3-benzoxazole-5-carboxylic acid) were identified by high-throughput screening using the HTRF assay and confirmed to have GOAT inhibitory activity using the ELISA. Based on the hit compound information, the novel lead compound (compound B, (4-chloro-6-{[2-methyl-6-(trifluoromethyl)pyridin-3-yl]methoxy}-1-benzothiophen-3-yl)acetic acid) was synthesized and exhibited potent GOAT inhibition with oral bioavailability. Both the hit compound and lead compound showed octanoyl-CoA competitive inhibitory activity. Moreover, these two compounds decreased acyl ghrelin production in the stomach of mice after their oral administration. These novel findings demonstrate that GOAT is a druggable target, and its inhibitors are promising antiobesity drugs.