Single cell analysis of endothelial cells in male and female developing heart

Background:

Sex differences exist in the circulatory system and the onset of cardiovascular diseases. While these differences are primarily attributed to sex hormones, it remains unclear when sex differences emerge in the cardiovascular system and when they begin to cause functional disparities. Our previous study using embryonic mice found that sex differences in the morphology of coronary vessels already exist at E17.5. It also revealed that female coronary vessels exhibit greater vasodilation capacity. However, it remains unclear how these sex differences form and how they relate to the sex differences observed later in development.

Methods:

Embryonic mouse hearts were sorted using FACS to obtain cardiac endothelial cells for single-cell RNA sequencing. Each sample was labeled with cell-multiplexing oligos, allowing us to distinguish between male and female cells within a single dataset. Differentially expressed genes between E17.5 male and female embryos were used to determine sex in a public dataset.

Results:

E17.5 mouse cardiac vascular endothelial cells were identified as artery, capillary1, capillary2, and vein. Females had a moderately higher proportion of arterial cells. Gene set analysis revealed that male vascular endothelial cells exhibited upregulated cell motility-associated genes, whereas those of females did not, but rather endothelial cell differentiation and response to laminar fluid shear stress. This result suggests that male and female endothelial cells exhibit distinct characteristics in achieving vascular maturation. To understand when the sex difference may have shown, E12.5 cardiac vascular endothelial cells were also analyzed. The gene expression was similar to that of E17.5, suggesting that the sex difference may be evident in earlier developmental stages.

Conclusion:

This study suggests that gene expression patterns in cardiac vascular endothelial cells showed sex differences at E17.5 or earlier. This may explain the functional sex differences in coronary artery development at embryonic stages.

Ms. Shion Nagasawa is a researcher specializing in heart development and cardiovascular diseases. She holds a master’s degree in Pharmaceutical Sciences from University of Shizuoka, Japan. She is currently a PhD student at Chiba University in Japan.