HiPS-Endothelial Cells Acquire Cardiac Endothelial Phenotype in Co-culture With hiPS-Cardiomyocytes

Abstract

Cell-cell interactions are crucial for organ development and function. In the heart, endothelial cells engage in bidirectional communication with cardiomyocytes regulating cardiac development and growth. We aimed to elucidate the organotypic development of cardiac endothelial cells and cardiomyocyte and endothelial cell crosstalk using human induced pluripotent stem cells (hiPSC). Single-cell RNA sequencing was performed with hiPSC-derived cardiomyocytes (hiPS-CMs) and endothelial cells (hiPS-ECs) in mono- and co-culture. The presence of hiPS-CMs led to increased expression of transcripts related to vascular development and maturation, cardiac development, as well as cardiac endothelial cell and endocardium-specific genes in hiPS-ECs. Interestingly, co-culture induced the expression of cardiomyocyte myofibrillar genes and MYL7 and MYL4 protein expression was detected in hiPS-ECs. Major regulators of BMP- and Notch-signaling pathways were induced in both cell types in co-culture. These results reflect the findings from animal studies and extend them to human endothelial cells, demonstrating the importance of EC-CM interactions during development.

Fate and state transitions during human blood vessel organoid development

“…Hence, in order to find appropriate culture conditions for organ-specific hBVOs, one needs to first decipher the major drivers of the differences between vascular cells of different organs. Epigenetic memory needs to be taken into consideration to improve approaches which have already been applied for certain tissues, such as co-culture of PSC-derived ECs with the desired tissue (Giacomelli et al, 2017; Helle et al, 2021; Sances et al, 2018), culture in tissue-conditioned media (Puech et al, 2018), addition of small molecules (Roudnicky et al, 2020a) or synergistic overexpression of transcription factors (Dailamy et al, 2021; Roudnicky et al, 2020b). We highlight the high degree of intraorgan heterogeneity displayed by ECs, which likely governs the difference in development and severity of vascular disorders.…”

Why is endothelial resilience key to maintain cardiac health?

“…In the heart, ECs show open chromatin of certain cardiomyocyte signature loci, express myofibrillar genes and cardiac-specific transcription factor MEF2C [70,140]. Co-culture of human-induced pluripotent stem cell-derived cardiomyocytes with ECs induced MYL7 and MYL4 expression as well as NOTCH and BMP signaling in endothelial cells [35]. The functions of cardiomyocyte gene expression in healthy or injured endothelium however are unclear.…”

Genomic and transcriptomic data analyses highlight KPNB1 and MYL4 as novel risk genes for congenital heart disease

“…All control cell lines were male. The cell lines were differentiated to hiPS-CMs as previously described (Helle et al, 2021). Briefly, Wnt-pathway activation and inactivation were used to induce differentiation, and metabolic selection with media containing lactate instead of glucose was used to ensure purity of the CM culture.…”

“…Briefly, Wnt-pathway activation and inactivation were used to induce differentiation, and metabolic selection with media containing lactate instead of glucose was used to ensure purity of the CM culture. The cells were processed for single cell sequencing as described previously (Helle et al, 2021)…”

“…The KPNB1 homolog Kpnb1 was expressed in all cells of the region, whereas the MYL4 homolog Myl4 was mainly expressed in the myocardium. Furthermore, we analyzed the expression of MYL4 and KPNB1 in hiPSC-derived endothelial cells (ECs) and cardiomyocytes (CMs) from a healthy individual grown in monoculture or in a coculture containing both cell types (Helle et al, 2021)(Figure 2B). The hiPS-ECs express the EC marker KDR/VEGFR2, and the hiPS-CMs express the cardiomyocyte marker TNNT2, defining the different cell types in the scRNAseq data (Figure 2B).…”