All organisms growing beyond the oxygen diffusion limit critically depend on a functional vasculature for survival. Yet blood vessels are far more than passive, uniform conduits for oxygen and ...nutrient supply. A remarkable organotypic heterogeneity is brought about by tissue-specific differentiated endothelial cells (lining the blood vessels’ lumen) and allows blood vessels to deal with organ-specific demands for homeostasis. On the flip side, when blood vessels go awry, they promote life-threatening diseases characterized by endothelial cells inappropriately adopting an angiogenic state (eg, tumor vascularization) or becoming dysfunctional (eg, diabetic microvasculopathies), calling respectively for antiangiogenic therapies and proangiogenic/vascular regenerative strategies. In solid tumors, despite initial enthusiasm, growth factor–based (mostly anti-VEGF vascular endothelial growth factor) antiangiogenic therapies do not sufficiently live up to the expectations in terms of efficiency and patient survival, in part, due to intrinsic and acquired therapy resistance. Tumors cunningly deploy alternative growth factors than the ones targeted by the antiangiogenic therapies to reinstigate angiogenesis or revert to other ways of securing blood flow, independently of the targeted growth factors. In trying to alleviate tissue ischemia and to repair dysfunctional or damaged endothelium, local in-tissue administration of (genes encoding) proangiogenic factors or endothelial (stem) cells harnessing regenerative potential have been explored. Notwithstanding evaluation in clinical trials, these approaches are often hampered by dosing issues and limited half-life or local retention of the administered agents. Here, without intending to provide an all-encompassing historical overview, we focus on some recent advances in understanding endothelial cell behavior in health and disease and identify novel molecular players and concepts that could eventually be considered for therapeutic targeting.
In 2009, it was postulated that endothelial cells (ECs) would only be able to execute the orders of growth factors if these cells would accordingly adapt their metabolism. Ten years later, it has ...become clear that ECs, often differently from other cell types, rely on distinct metabolic pathways to survive and form new blood vessels; that manipulation of EC metabolic pathways alone (even without changing angiogenic signaling) suffices to alter vessel sprouting; and that perturbations of these metabolic pathways can underlie excess formation of new blood vessels (angiogenesis) in cancer and ocular diseases. Initial proof of evidence has been provided that targeting (normalizing) these metabolic perturbations in diseased ECs and delivery of metabolites deserve increasing attention as novel therapeutic approaches for inhibiting or stimulating vessel growth in multiple disorders.
Angiogenesis research has been focused for decades on growth factors and has largely overlooked the role of endothelial cell metabolism. Li et al. overview the breakthroughs in endothelial cell metabolism over the past decade and discusses possible therapeutic implications for stimulating or inhibiting vessel growth by manipulating endothelial cell metabolism.
More than a decade ago, when we first embarked on our joumey to delineate the biological function of vascular endothelial growth factor B （VEGF-B）, we had a hard time comprehending why VEGF-B was ...needed. In mice, genetic deletion of VEGF-B seemed to be harmless, since the VEGF-B null mice, to a large extent, can still live a fairly normal life 1. Moreover, overexpression of VEGF-B in different mouse tissues, such as the skin or skeletal muscle,
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-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.
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•We single-cell RNA-sequenced 56,771 endothelial cells (ECs) from human, mouse, and cultured lung tumor models•Tip ECs were resolved into migratory and basement-membrane remodeling phenotypes•Capillary and venous ECs expressed immunoregulatory gene signatures•Integrated analysis identified collagen modification as an angiogenic pathway
Goveia et al. use single-cell RNA sequencing to provide an inventory of tumor endothelial cell (TEC) phenotypes from human and mouse non-small cell lung cancer and validate them functionally. Specific TEC phenotypes are associated with prognosis and response to anti-angiogenic therapy.
The role of phosphoglycerate dehydrogenase (PHGDH), a key enzyme of the serine synthesis pathway (SSP), in endothelial cells (ECs) remains poorly characterized. We report that mouse neonates with ...EC-specific PHGDH deficiency suffer lethal vascular defects within days of gene inactivation, due to reduced EC proliferation and survival. In addition to nucleotide synthesis impairment, PHGDH knockdown (PHGDHKD) caused oxidative stress, due not only to decreased glutathione and NADPH synthesis but also to mitochondrial dysfunction. Electron transport chain (ETC) enzyme activities were compromised upon PHGDHKD because of insufficient heme production due to cellular serine depletion, not observed in other cell types. As a result of heme depletion, elevated reactive oxygen species levels caused EC demise. Supplementation of hemin in PHGDHKD ECs restored ETC function and rescued the apoptosis and angiogenesis defects. These data argue that ECs die upon PHGDH inhibition, even without external serine deprivation, illustrating an unusual importance of serine synthesis for ECs.
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•PHGDH loss in endothelial cells causes lethal angiogenesis defects•PHGDH inhibition impairs heme synthesis and reduces purine and pyrimidine synthesis•Heme deficiency causes mitochondrial respiration defects and lethal oxidative stress•Hemin supplementation rescues loss of PHGDH in endothelial cells
Vandekeere et al. highlight a unique role for serine metabolism in endothelial cells (ECs). ECs utilize the serine synthetic pathway (SSP) differently from cancer cells and rely exclusively on the SSP for heme synthesis to maintain mitochondrial respiration and homeostasis.
The role of fatty acid synthesis in endothelial cells (ECs) remains incompletely characterized. We report that fatty acid synthase knockdown (FASNKD) in ECs impedes vessel sprouting by reducing ...proliferation. Endothelial loss of FASN impaired angiogenesis in vivo, while FASN blockade reduced pathological ocular neovascularization, at >10-fold lower doses than used for anti-cancer treatment. Impaired angiogenesis was not due to energy stress, redox imbalance, or palmitate depletion. Rather, FASNKD elevated malonyl-CoA levels, causing malonylation (a post-translational modification) of mTOR at lysine 1218 (K1218). mTOR K-1218 malonylation impaired mTOR complex 1 (mTORC1) kinase activity, thereby reducing phosphorylation of downstream targets (p70S6K/4EBP1). Silencing acetyl-CoA carboxylase 1 (an enzyme producing malonyl-CoA) normalized malonyl-CoA levels and reactivated mTOR in FASNKD ECs. Mutagenesis unveiled the importance of mTOR K1218 malonylation for angiogenesis. This study unveils a novel role of FASN in metabolite signaling that contributes to explaining the anti-angiogenic effect of FASN blockade.
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•Fatty acid synthase inhibition impairs physiological and pathological angiogenesis•FASN inhibition selectively reduces endothelial cell proliferation, not migration•By elevating malonyl-CoA levels, FASN inhibition promotes mTOR lysine malonylation•The resultant decrease in mTORC1 kinase activity contributes to angiogenic defects
Bruning et al. report that blocking fatty acid synthase (FASN) in endothelial cells (ECs) reduces angiogenesis by impairing EC proliferation. Mechanistically, FASN inhibition elevates the malonyl-CoA substrate pool, thereby increasing post-translational malonylation of mTOR and decreasing the pro-angiogenic mTORC1 activity.
Single-cell RNA sequencing (scRNA-seq) has become a powerful tool for scientists of many research disciplines due to its ability to elucidate the heterogeneous and complex cell-type compositions of ...different tissues and cell populations. Traditional cell-type identification methods for scRNA-seq data analysis are time-consuming and knowledge-dependent for manual annotation. By contrast, automatic cell-type identification methods may have the advantages of being fast, accurate, and more user friendly. Here, we discuss and evaluate thirty-two published automatic methods for scRNA-seq data analysis in terms of their prediction accuracy, F1-score, unlabeling rate and running time. We highlight the advantages and disadvantages of these methods and provide recommendations of method choice depending on the available information. The challenges and future applications of these automatic methods are further discussed. In addition, we provide a free scRNA-seq data analysis package encompassing the discussed automatic methods to help the easy usage of them in real-world applications.
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.
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.
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
and in dehydrated mice
.
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.
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.
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