AJAP1 Regulates Human Epithelial Cell Behaviors Required for Epicardium and EPDC Formation During Cardiac Development
Authors: Shemon Claire , Ashby Elizabeth , Green Rebecca , Castaneda Christopher , Gerick Alexander , Wilson Lauren , Hatcher Cathy J 
Published: May 2024

Introduction: The epicardium forms the outermost layer of the heart. It is composed of a single layer of specialized epithelial cells that make important cellular contributions to cardiovascular development. During embryonic development, the epicardium is a source of multipotent progenitor cells, growth factors and extracellular matrix (ECM) components. The epicardium gives rise to multipotent epicardial-derived cells (EPDCs) which undergo epithelial-to-mesenchymal transition and differentiate into non-myocyte cardiac lineages. It is also a source of paracrine signals essential for myocardial growth and coronary vessel formation that become dormant after gestation. Adult epicardial cells can be reactivated in response to myocardial injury or ischemia to express a fetal gene program for regenerative cardiac repair. The drive to harness epicardial cells for cardiac regeneration is tethered to its roles in cardiovascular development. Thus, it is necessary to identify molecules that regulate epicardial cell behaviors required for epicardium formation and EPDC differentiation. Using our Tbx5-deficient mouse model of defective epicardium formation ( Tbx5epi−/−), we identified a transcript that is novel to the heart. Adherens junction associated protein 1 is a cell adhesion molecule that is encoded by the AJAP1 transcript which was downregulated in Tbx5epi−/− mice with impaired epicardium formation. Our preliminary studies revealed that AJAP is expressed in epithelial cells of the epicardium during heart development. Silencing of AJAP1 in primary cultured human epithelial cells reduces their ability to migrate. A role for AJAP1 in the epicardium during cardiac development remains to be identified. Study Objective: This study was designed to investigate the impact of AJAP1 on epithelial cell functions that are required for formation of the epicardium and differentiation of EPDCs. Methods: Studies were conducted in a primary human epithelial cell line, HMEpiC, which served as an in vitro model system to examine the contributions of AJAP1 to epithelial cell behaviors. HMEpiCs were transfected with small interfering RNAs (siRNAs) targeting either human AJAP1 to knock down its endogenous expression or a non-targeting negative control siRNA. AJAP1 knockdown was confirmed by quantitative PCR and immunofluorescence. We conducted assays on control and AJAP1 knockdown (KD) cells to examine the effects on epithelial cell behaviors contributing to epicardium formation and EPDC differentiation. This included analysis of cell proliferation, apoptosis, and adhesion to extracellular matrices. Results: Endogenous AJAP1 mRNA expression was reduced significantly by siRNA-mediated knockdown in human epithelial cells. We observed changes in cell morphology that correlated with a significant increase in apoptosis in AJAP1 KD cells compared to negative control cells. Proliferation of HMEpiCs was unchanged by AJAP1 knockdown. Also, we observed a significant increase in the adhesion of AJAP1 KD cells to certain extracellular matrices versus control cells. Conclusion: These findings provide evidence for the AJAP1 cell adhesion molecule to regulate epithelial cell behaviors that are crucial to formation of the epicardium and differentiation of EPDCs during cardiac development. PCOM Center for Chronic Disorders of Aging; PCOM Division of Research. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.