Human induced pluripotent stem cell-derived kidney organoids have potential for disease modeling and to be developed into clinically transplantable auxiliary tissue. However, they lack a functional... Show moreHuman induced pluripotent stem cell-derived kidney organoids have potential for disease modeling and to be developed into clinically transplantable auxiliary tissue. However, they lack a functional vasculature, and the sparse endogenous endothelial cells (ECs) are lost upon prolonged culture in vitro, limiting maturation and applicability. Here, we use intracoelomic transplantation in chicken embryos followed by single-cell RNA sequencing and advanced imaging platforms to induce and study vasculogenesis in kidney organoids. We show expansion of human organoid-derived ECs that reorganize into perfused capillaries and form a chimeric vascular network with host-derived blood vessels. Ligand-receptor analysis infers extensive potential interactions of human ECs with perivascular cells upon transplantation, enabling vessel wall stabilization. Perfused glomeruli display maturation and morphogenesis to capillary loop stage. Our findings demonstrate the beneficial effect of vascularization on not only epithelial cell types, but also the mesenchymal compartment, inducing the expansion of ' on target ' perivascular stromal cells, which in turn are required for further maturation and stabilization of the neo-vasculature. The here described vasculogenic capacity of kidney organoids will have to be deployed to achieve meaningful glomerular maturation and kidney morphogenesis in vitro. Show less
Dumas, S.J.; Meta, E.; Borri, M.; Luo, Y.L.; Li, X.R.; Rabelink, T.J.; Carmeliet, P. 2021
The adult kidney vasculature comprises diverse populations of endothelial cells that support specific functions according to their microenvironment. This Review summarizes our current understanding... Show moreThe adult kidney vasculature comprises diverse populations of endothelial cells that support specific functions according to their microenvironment. This Review summarizes our current understanding of the phenotypic, molecular and metabolic heterogeneity of renal endothelial cells in relation to their microenvironment and the potential application of targeting renal endothelial cell metabolism as a therapeutic strategy for kidney diseases or kidney regeneration.Complex multicellular life in mammals relies on functional cooperation of different organs for the survival of the whole organism. The kidneys play a critical part in this process through the maintenance of fluid volume and composition homeostasis, which enables other organs to fulfil their tasks. The renal endothelium exhibits phenotypic and molecular traits that distinguish it from endothelia of other organs. Moreover, the adult kidney vasculature comprises diverse populations of mostly quiescent, but not metabolically inactive, endothelial cells (ECs) that reside within the kidney glomeruli, cortex and medulla. Each of these populations supports specific functions, for example, in the filtration of blood plasma, the reabsorption and secretion of water and solutes, and the concentration of urine. Transcriptional profiling of these diverse EC populations suggests they have adapted to local microenvironmental conditions (hypoxia, shear stress, hyperosmolarity), enabling them to support kidney functions. Exposure of ECs to microenvironment-derived angiogenic factors affects their metabolism, and sustains kidney development and homeostasis, whereas EC-derived angiocrine factors preserve distinct microenvironment niches. In the context of kidney disease, renal ECs show alteration in their metabolism and phenotype in response to pathological changes in the local microenvironment, further promoting kidney dysfunction. Understanding the diversity and specialization of kidney ECs could provide new avenues for the treatment of kidney diseases and kidney regeneration. Show less
Rohlenova, K.; Goveia, J.; Garcia-Caballero, M.; Subramanian, A.; Kalucka, J.; Treps, L.; ... ; Carmeliet, P. 2020
Endothelial cell (EC) metabolism is an emerging target for anti-angiogenic therapy in tumor angiogenesis and choroidal neovascularization (CNV), but little is known about individual EC metabolic... Show moreEndothelial cell (EC) metabolism is an emerging target for anti-angiogenic therapy in tumor angiogenesis and choroidal neovascularization (CNV), but little is known about individual EC metabolic transcriptomes. By single-cell RNA sequencing 28,337 murine choroidal ECs (CECs) and sprouting CNV-ECs, we constructed a taxonomy to characterize their heterogeneity. Comparison with murine lung tumor ECs (TECs) revealed congruent marker gene expression by distinct EC phenotypes across tissues and diseases, suggesting similar angiogenic mechanisms. Trajectory inference predicted that differentiation of venous to angiogenic ECs was accompanied by metabolic transcriptome plasticity. ECs displayed metabolic transcriptome heterogeneity during cell- cycle progression and in quiescence. Hypothesizing that conserved genes are important, we used an integrated analysis, based on congruent transcriptome analysis, CEC-tailored genome-scale metabolic modeling, and gene expression meta-analysis in cross-species datasets, followed by in vitro and in vivo validation, to identify SQLE and ALDH18A1 as previously unknown metabolic angiogenic targets. Show less
Heterogeneity of lung tumor endothelial cell (TEC) phenotypes across patients, species (human/mouse), and models (in vivo/in vitro) remains poorly inventoried at the single-cell level. We single... Show moreHeterogeneity of lung tumor endothelial cell (TEC) phenotypes across patients, species (human/mouse), and models (in vivo/in vitro) remains poorly inventoried at the single-cell level. We single-cell RNA (scRNA)-sequenced 56,771 endothelial cells from human/mouse (peri)-tumoral lung and cultured human lung TECs, and detected 17 known and 16 previously unrecognized phenotypes, including TECs putatively regulating immune surveillance. We resolved the canonical tip TECs into a known migratory tip and a putative basement-membrane remodeling breach phenotype. Tip TEC signatures correlated with patient survival, and tip/breach TECs were most sensitive to vascular endothelial growth factor blockade. Only tip TECs were congruent across species/models and shared conserved markers. Integrated analysis of the scRNA-sequenced data with orthogonal multi-omics and meta-analysis data across different human tumors, validated by functional analysis, identified collagen modification as a candidate angiogenic pathway. Show less
Dumas, S.J.; Meta, E.; Borri, M.; Goveia, J.; Rohlenova, K.; Conchinha, N.V.; ... ; Carmeliet, P. 2020
Background Renal endothelial cells from glomerular, cortical, and medullary kidney compartments are exposed to different microenvironmental conditions and support specific kidney processes. However... Show moreBackground Renal endothelial cells from glomerular, cortical, and medullary kidney compartments are exposed to different microenvironmental conditions and support specific kidney processes. However, the heterogeneous phenotypes of these cells remain incompletely inventoried. Osmotic homeostasis is vitally important for regulating cell volume and function, and in mammals, osmotic equilibrium is regulated through the countercurrent system in the renal medulla, where water exchange through endothelium occurs against an osmotic pressure gradient. Dehydration exposes medullary renal endothelial cells to extreme hyperosmolarity, and how these cells adapt to and survive in this hypertonic milieu is unknown.Methods We inventoried renal endothelial cell heterogeneity by single-cell RNA sequencing >40,000 mouse renal endothelial cells, and studied transcriptome changes during osmotic adaptation upon water deprivation. We validated our findings by immunostaining and functionally by targeting oxidative phosphorylation in a hyperosmolarity model in vitro and in dehydrated mice in vivo.Results We identified 24 renal endothelial cell phenotypes (of which eight were novel), highlighting extensive heterogeneity of these cells between and within the cortex, glomeruli, and medulla. In response to dehydration and hypertonicity, medullary renal endothelial cells upregulated the expression of genes involved in the hypoxia response, glycolysis, and-surprisingly-oxidative phosphorylation. Endothelial cells increased oxygen consumption when exposed to hyperosmolarity, whereas blocking oxidative phosphorylation compromised endothelial cell viability during hyperosmotic stress and impaired urine concentration during dehydration.Conclusions This study provides a high-resolution atlas of the renal endothelium and highlights extensive renal endothelial cell phenotypic heterogeneity, as well as a previously unrecognized role of oxidative phosphorylation in the metabolic adaptation of medullary renal endothelial cells to water deprivation. Show less
