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Introduction
The absence of a functional vasculature in cultured human induced pluripotent stem cell (hiPSC)-derived kidney organoids is a major limitation. It precludes full maturation and functionality of organoid nephron structures, thus restricting their applicability. In organoids maintained in vitro, the sparse endothelial cells (ECs) that are initially formed during differentiation perish over time, suggesting the absence of essential cues for EC maintenance in the culture conditions. We have previously shown that transplantation in the coelomic cavity of chicken embryos induces functional vascularization and enhanced maturation of kidney organoids. Interestingly, the vasculature in these transplanted organoids is chimeric, consisting of both chicken and human ECs which, contrary to the in vitro situation, are maintained in vivo. To date, it is unclear what leads to the demise of human ECs in vitro as well as what sustains them upon transplantation.
Renal ECs are known to display extensive molecular and phenotypic heterogeneity to support the specific functions of each nephron segment1, 2, 3–4. Glomerular ECs display fenestrae without diaphragms and play a role in glomerular filtration. ECs in peritubular capillaries have bridged fenestrae and contribute to the tubular reabsorption of solutes. In the larger renal vessels, the EC lining is continuous. A recent study demonstrated that renal EC heterogeneity is even more extensive than was previously thought by identifying at least 24 distinct renal EC phenotypes in the adult mouse kidney based on single-cell RNA sequencing (scRNAseq) analysis1. However, it is unknown whether and to what extent kidney organoid-derived ECs resemble renal ECs. Although previous studies provided scRNA-seq data of kidney organoid cells, the absence or paucity of ECs recovered from the organoids precluded the extensive analysis of this critical cell population. In this study, we isolated a total of 11,966 high-quality human ECs from untransplanted and transplanted kidney organoids and analyzed them using scRNAseq. We demonstrate that organoid ECs in vitro display phenotypic heterogeneity, but lack similarity to fetal kidney ECs. Transplantation in chicken embryos stimulates EC proliferation and induces a new, more mature, arterial EC phenotype, resembling fetal kidney arterial-afferent arteriolar ECs.
Results
Transplantation greatly affects the kidney organoid endothelial cell transcriptome
To evaluate the molecular effects of transplantation on kidney organoid ECs, we compared human ECs from organoids cultured in...