Endothelial Characteristics of Cardiac Resident Stem Cell Antigen-1+ and c-kit+ Cells

Abstract

Background: Stem cell antigen-1 (Sca-1) and c-kit have been extensively used as cell surface markers to identify stem cells in various organs. In the heart, these were originally suggested to be markers for the intrinsic cardiac myocyte stem cells; however, a growing body of evidence now suggests that the majority of Sca-1+ or c-kit+ cells in the heart are of endothelial lineage. These cells may play an important role in vascular adaptation during cardiac remodelling and pathophysiological heart diseases. However, endothelial functions of these cells have not been studied. Furthermore, there is no information regarding differences in the abundance of this population across ventricles, sexes, strains of rat, and in right ventricle pathological conditions. Purpose: The purpose of this thesis was to examine the endothelial cell characteristics of cardiac Sca-1+ cells and c-kit+ expressing cells, and assess the distribution across ventricles, sexes, strains, and in a model of RV overload. Hypothesis: We hypothesize that cardiac resident Sca-1+ cells and c-kit+ cells are endothelial precursor/progenitor cells and have angiogenic function. Biological sex, genetic background, and disease state affect abundance of these cells. Methods: Hearts from adult male and female Sprague Dawley (SD) and Fischer CDF rats were excised and digested to obtain a single-cell suspension. Cells were then stained with stem cell (Sca-1 and c-kit) and endothelial cell (CD31) markers, and analyzed using flow cytometry to determine the abundance of the cell population. Sca-1+ cells were isolated through magnetic sorting to assess endothelial cell characteristics, such as cell morphology, acetylated low density lipoprotein uptake, lectin binding, and network formation, when cultured on Matrigel. To study the changes during pathological cardiac remodelling, flow cytometry analysis of the Fischer CDF rat hearts was performed at 4 weeks after monocrotaline injection (MCT; 60 mg/kg, sc). Results: A majority of Sca-1+ cells and c-kit+ cells in the heart were CD31+. Sca-1+ cells isolated from SD rat hearts demonstrated lectin binding and of uptake acetylated low- density lipoprotein. The right ventricle (RV) of the heart had more c-kit+CD31+ cells and Sca-1+CD31+ cells than the left ventricle (LV). In normal healthy rats, the abundance of c-kit+CD31+ cells and Sca- 1+CD31+ cells in the RV of male rats were similar to the RV of female rats. MCT treatment led to an increase in RV systolic pressure (RVSP) and RV hypertrophy, which was associated with a marked reduction in abundance of c-kit+CD31+ or Sca-1+CD31+ cells in the RV. Conclusions and Clinical Significance: The c-kit or Sca-1 expressing cells in the heart are resident endothelial progenitor-like cells. These cells are highly abundant in the RV, and a marked reduction in abundance of these cells is observed during pathological RV remodelling. Considering the higher abundance of these cells in the RV, these cells may also play a role in vascular adaptation during RV remodeling. It is important to note that RV angiogenesis has been shown to be critical for maintenance of RV function.