Epithelial Cell Requirement for AJAP1 in Cardiac Development
Authors: Stella Amanda , Adjei‐Danquah Edith , Bunting Clayton , Stewart Jonathan , Van Wyk Albertus , Scott Ellen , Hatcher Cathy J 
Published: June 2021

Introduction

Cardiovascular development occurs through a series of molecular and cellular processes. Disruption of these processes result in congenital heart defects that compromise normal cardiovascular function. During mammalian cardiovascular development, the transitory proepicardium (PE) is a progenitor cell source for the epicardium and a subset of coronary vascular cells. The PE is composed of epithelial‐like mesothelial cells that migrate onto the myocardium to form the epicardium. These epicardial cells proliferate, and undergo apoptosis and epithelial‐mesenchymal transition to become epicardial‐derived cells (EPDCs). These EPDCs delaminate and migrate into the myocardium to form a subset of primitive coronary vascular cells. Molecular signals that contribute to formation of the epicardium and coronary vessels are being examined. Immunofluorescent studies from our lab indicate that the adherens junction‐associated protein (Ajap1) is expressed in the transitory PE, epicardium and primitive coronary vessels of mouse hearts. AJAP1 is a cell adhesion molecule that associates with β‐catenin in the E‐catenin‐cadherin complex to form cell adherens junctions in extra‐cardiac tissues. However, its role in the heart remains unknown.

Study Objective

We sought to elucidate how AJAP1 influences epithelial cell behaviors including cell proliferation, apoptosis, migration and epithelial‐mesenchymal transition (EMT). Previous investigations indicated that cell cycle regulatory properties often associate with EMT. Based on this idea, we hypothesized that AJAP1 regulates establishment of the epithelial‐like layer of epicardial cells by influencing epithelial cell proliferation, apoptosis, migration and EMT.

Methods

We utilized primary human mammary epithelial cells (HMEpiCs) as an in vitro system to explore contributions of AJAP1 to epithelial cell behavior. A small interfering RNA (siRNA)‐mediated approach was used to silence endogenous AJAP1 gene expression and reduce its protein expression. siRNA‐transfected HMEpiCs were subjected to gene expression analyses and cell‐based assays to examine changes in epithelial cell behavior respondent to AJAP1 silencing. We assessed epithelial cell morphology, proliferation, apoptosis, migration and EMT induction via TGFβ1 stimulation.

Results

AJAP1‐silenced HMEpiCs exhibited a >75% decrease in AJAP1 mRNA expression in comparison to non‐targeting, negative control HMEpiCS. Reduced AJAP1 expression produced discernable changes in cell morphology prompting us to explore other cell behaviors. We observed alterations in expression of cell cycle and apoptotic markers in AJAP1‐silenced HMEpiCs versus control cells. These findings led us to investigate how these cell cycle alterations influenced epithelial cell migration and EMT via marker analysis.

Conclusion

We conclude that AJAP1 contributes to pertinent behaviors of epithelial cells, including cell morphology, viability and differentiation. These findings will potentially elucidate how AJAP1 contributes to epicardium formation and subsequent development of coronary vessels.

Support or Funding Information

PCOM Center for Chronic Disorders of Aging

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.