cells Article Quantitative Evaluation of Cardiac Cell Interactions and Responses to Cyclic Strain Richard Duc Hien Tran 1,2,† , Tessa Altair Morris 1,3,4,† , Daniela Gonzalez 1,2 , Ali Hatem Salaheldin Hassan Ahmed Hetta 1,2 and Anna Grosberg 1,2,3,4,5, *   Citation: Tran, R.D.H.; Morris, T.A.; Gonzalez, D.; Hetta, A.H.S.H.A.; Grosberg, A. Quantitative Evaluation of Cardiac Cell Interactions and Responses to Cyclic Strain. Cells 2021, 10, 3199. https://doi.org/10.3390 /cells10113199 Academic Editor: Wayne Carver Received: 27 August 2021 Accepted: 27 October 2021 Published: 17 November 2021 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affil- iations. Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). 1 Edwards Lifesciences Center for Advanced Cardiovascular Technology, University of California, Irvine, CA 92617-2700, USA; rdtran1@uci.edu (R.D.H.T.); tessam@uci.edu (T.A.M.); gonzad14@uci.edu (D.G.); asalahel@uci.edu (A.H.S.H.A.H.) 2 Department of Biomedical Engineering, University of California, Irvine, CA 92617, USA 3 Center for Complex Biological Systems, University of California, Irvine, CA 92697, USA 4 NSF-Simons Center for Multiscale Cell Fate Research, University of California, Irvine, CA 92697, USA 5 Department of Chemical and Biomolecular Engineering, University of California, Irvine, CA 92617, USA * Correspondence:grosberg@uci.edu † These authors contributed equally to this work. Abstract: The heart has a dynamic mechanical environment contributed by its unique cellular composition and the resultant complex tissue structure. In pathological heart tissue, both the mechanics and cell composition can change and influence each other. As a result, the interplay between the cell phenotype and mechanical stimulation needs to be considered to understand the biophysical cell interactions and organization in healthy and diseased myocardium. In this work, we hypothesized that the overall tissue organization is controlled by varying densities of cardiomyocytes and fibroblasts in the heart. In order to test this hypothesis, we utilized a combination of mechanical strain, co-cultures of different cell types, and inhibitory drugs that block intercellular junction formation. To accomplish this, an image analysis pipeline was developed to automatically measure cell type-specific organization relative to the stretch direction. The results indicated that cardiac cell type-specific densities influence the overall organization of heart tissue such that it is possible to model healthy and fibrotic heart tissue in vitro. This study provides insight into how to mimic the dynamic mechanical environment of the heart in engineered tissue as well as providing valuable information about the process of cardiac remodeling and repair in diseased hearts. Keywords: heart tissue organization; cell type classification; cyclic strain; intercellular junctions 1. Introduction The two dominant cell types in the myocardium are cardiomyocytes and cardiac fibroblasts; cardiomyocytes generate contractile force [1–3], while fibroblasts play vital roles in maintaining functions within the heart, such as extracellular matrix production, cardiac remodeling, cell–cell signaling, promoting blood vessel formation, and secretion of growth factors and cytokines [4–6]. In a healthy heart, cardiomyocytes and fibroblasts are organized along the direction of contraction [7–9]. However, in the event of myocardial infarction or other cardiac diseases, there is increased migration of fibroblasts into the regions of damaged tissue as well as changes to the morphology and viability of the myocytes [10–15]. The alterations in cellular composition and structure result in disorganization and loss of efficient heart function [16–18]. Although cellular disorganization, as well as a shift in the dominant cell type as a result of injury or remodeling, such as fibrosis, has been observed, the mechanisms that drive the organization of cardiomyocytes and fibroblasts and how they influence each other are not fully understood. Investigating the mechanisms responsible for organization in the heart is imperative to create accurate in vitro models of infarcted or diseased hearts, Cells 2021, 10, 3199. https://doi.org/10.3390/cells10113199 https://www.mdpi.com/journal/cells