Part 2 Phenotypic Diversity And Metabolic Specialization Of Renal Endothelial Cells

Contact: Audrey Hu Whatsapp/hp: 0086 13880143964 Email: audrey.hu@wecistanche.com

Sébastien J. Dumas^1,6, Elda Meta^1,6, Mila Borri ^1,6, Yonglun Luo ^2,3, Xuri Li ⁴, Ton J. Rabelink⁵and Peter Carmeliet ^1,4

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Key points

• The endothelium differs between different organs, probably to support distinct organ functions.

• Multiple specialized endothelial cell phenotypes coexist within the renal glomeruli, cortex, and medulla; these function to support glomerular filtration, the reabsorption and secretion of ions and metabolites, and urine concentration.

• The different local microenvironments in the kidney shape the molecular and metabolic heterogeneity of the renal endothelium; conversely, endothelial cell-derived endocrine factors sustain the niches of different kidney microenvironments.

• The metabolism of renal endothelial cells can be altered in the context of kidney injury and diseases, partly as a result of changes in the microenvironment.

• Greater understanding of the phenotypic diversity and metabolic specialization of renal endothelial cells may aid the identification of new targets for the treatment of kidney diseases and kidney regeneration.

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REC responses to changes in shear stress ECs are constantly exposed to a force of stretch-induced by the pulsatility of blood flow and a frictional force parallel to the vessel wall, the fluid shear stress¹⁹⁵. These cells are equipped to sense these forces and to transduce them into biochemical signals that can affect vessel homeostasis by regulating vascular tone and EC remodeling, which adjusts the blood flow to meet tissue requirements195. Interestingly, ECs from different portions of the vascular bed are exposed to specific flow types and respond accordingly195 (Fig. 5a). In arteries and arterioles, blood flow is highly pulsatory, whereas in capillaries it is of similar magnitude but less pulsatory, and in venules and veins blood flow is about three-fold to tenfold lower and pulsatility is minimal195. In the kidney, the vasculature of the cortex receives >94% of renal blood flow¹⁹⁶ suggesting that the medullary vasculature is exposed to a relatively low shear stress environment. By contrast, gRECs are exposed to a relatively high shear stress environment (estimated to be from 1dyn/cm2 to as high as 95dyn/cm2 ), as a result of high blood flow and pressure combined with increased blood viscosity as a result of the filtration process197 (Fig. 5b). Exposure of gREC to shear stress is critical since it prevents platelet aggregation, in part by inducing the conformational unfolding of the blood glycoprotein von Willebrand factor (vWF) which enhances its susceptibility to cleavage by the ADAMTS13 metalloprotease^198,199. The importance of this phenomenon to gREC health is demonstrated by the pro-thrombotic phenotype observed in Shiga toxin-associated hemolytic uraemic syndrome. Shiga toxin promotes the secretion of vWF by gRECs and the formation of ultra-large vWF multimers that are resistant to cleavage by ADAMTS13 and induce thrombotic microangiopathy in glomeruli and the kidney microvasculature200, eventually leading to AKI. In thrombotic thrombocytopenic purpura, low ADAMTS13 activity results in a similar outcome¹⁹⁹.

The importance of shear stress is also illustrated by the development of atherosclerotic lesions in renal arteries, which can result in renal artery stenosis — the single largest cause of secondary hypertension201,202. These lesions develop in atheroprone regions in the arteries and arterioles that are exposed to lower laminar shear stress than other regions, such as regions of arterial bifurcation, where blood flow is typically disturbed201 (Fig. 5c).

Effects of laminar shear stress on ECs. Laminar shear stress induces upregulation of the transcription factors Krüppel-like factor 2 (KLF2) and KLF4 in ECs, in part through the release of ATP and subsequent activation of P2X4 purinergic receptors203, and in part via activation of the MEK5–ERK5–MEF2 signaling pathway^204,205 (Fig. 5d). In the kidney, Klf2, and Klf4 together with the KLF4 target gene, Thbd (which encodes thrombomodulin), have been reported to be markers of gRECs derived from the efferent arterioles in adult mice^10,11 (Fig. 3b). The location of these flow-responsive markers is consistent with the fact that RECs located at the immediate exit site of glomeruli are exposed to high laminar shear stress, potentially related to the high blood viscosity of this region. The upregulation and activation of KLF2 mediate EC quiescence, characterized by an increase in the expression of VE-cadherin and β-catenin to aid the maintenance of a tight vascular barrier206, the alignment of ECs in the direction of flow207, the inhibition of inflammation and maintenance of an anti-atherogenic phenotype, an upregulation of antioxidants and a decrease in vascular tone secondary to the endothelial production of NO and prostacyclins208–210 (Fig. 5d). Activation of KLF2 in gRECs protected them against injury and disease progression in animal models of CKD^211,212. Accordingly, KLF2 is upregulated in the glomerular endothelium in response to shear stress in vitro, where it promotes an anticoagulant and anti-inflammatory phenotype and triggers an endothelial-dependent decrease in podocyte barrier resistance, required for proper filtration function204. The upregulation of gREC KLF2 secondary to glomerular hyperfiltration conferred protection against EC dysfunction and attenuated the progression of CKD in a model of unilateral nephrectomy212. Conversely, loss of endothelial KLF2 exacerbated glomerular hypertrophy and proteinuria in a model of streptozotocin-induced diabetic kidney disease²¹¹. Endothelial KLF4 is also renoprotective in AKI213. EC-specific loss of KLF4 exacerbated kidney injury in a model of ischaemic AKI by promoting EC acquisition of a pro-inflammatory phenotype²¹³. However, it is worth noting that KLF2 and KLF4 have context-specific roles in the endothelium214. For instance, they can promote the activation of ECs and the formation of lesions leading to cerebral cavernous malformations in development^214,215.

Exposure to laminar shear stress reduces glucose uptake²¹⁶ and stimulates mitochondrial biogenesis in ECs^217–219. KLF2 activation downregulates the expression of PFKFB3 together with that of other glycolytic genes such as HK2 (which encodes hexokinase 2) and PFK1 (which encodes phosphofructokinase 1 (PFK1)), resulting in a decrease in glycolysis216, and the shuttling of available early glycolytic intermediates to the hexosamine and glucuronic acid biosynthetic pathway for UDP-GlcUA and UDP-GlcNAc synthesis, respectively, which are the limiting substrates of hyaluronan synthase (HAS2)^{133,220–222}. KLF2 also induces the expression and membrane translocation of HAS2 and the consequent synthesis of the glycocalyx component hyaluronan^133,222. Thus, ECs exposed to laminar flow display a much thicker glycocalyx than ECs exposed to disturbed flow133 (Fig. 5d,e). EC-specific deletion of Has2 has profound effects on the kidney, including impairment of the glycocalyx structure of capillary gRECs, disruption of the glomerular endothelial fenestrations40, albuminuria indicative of filtration barrier dysfunction, disturbed glycocalyx-dependent angiopoietin 1 signaling, and abnormal podocyte structures as a result of abnormal EC–podocyte crosstalk, resulting in glomerular capillary rarefaction and glomerulosclerosis⁴⁰.

Mitochondrial respiration and ATP generation are also increased in ECs under unidirectional flow compared with those exposed to disturbed flow223,224. Blockade of mitochondrial ATP generation inhibits shear stress-induced ATP release whereas, by contrast, inhibition of glycolysis has no effect, suggesting that EC mitochondrial respiration is required for purinergic receptor activation, which in turn induces KLF2 expression in response to shear stress^203,218,224. Furthermore, mitochondrial biogenesis is upregulated in response to shear stress, owing to the activation of a SIRT1–PGC1a– TFAM signaling cascade217,219, whereas the expression of antioxidant genes, such as haem oxygenase 1 and glutaredoxin 1, is increased to protect ECs from ROS225,226 (Fig. 5d). Inhibiting the electron transport chain in ECs exposed to laminar flow resulted in EC inflammation, suggesting that mitochondrial respiration prevents EC activation²²³. In the kidney, activation of the 5-HT1F receptor to stimulate mitochondrial biogenesis in RECs may reduce vascular rarefaction and promote recovery from injury as has been shown in a model of AKI227.

KLF4 also induces the upregulation of cholesterol-25- hydroxylase (CH25H) and liver X receptor upon exposure to an atheroprotective pulsatile shear stress²²⁸. CH25H catalyzes the production of 25-hydroxycholesterol, which prevents the activation of sterol regulatory element-binding protein 2 (SREBP2), an important mediator in the EC response to disturbed blood flow (see below)229,230. Hence, modulation of KLF2 and KLF4 expression and activation by laminar shear stress and their consequent metabolic responses may have a critical role in the maintenance of REC quiescence and glomerular filtration (Fig. 5d).

Effects of disturbed blood flow on ECs. ECs exposed to disturbed blood flow such as in arterial bifurcations or curvatures are activated and exhibit a pro-inflammatory and atherogenic phenotype231 (Fig. 5e). Thus, the activation of arterial and afferent arteriolar RECs located in such atheroprone regions might promote the development of atherosclerotic renal artery stenosis.

KLF4 expression is repressed by disturbed blood flow, with increased methylation of the KLF4 promoter region preventing MEF2 binding and subsequent KLF4 transcription²³². Furthermore, microRNA (miR)-92a, which is induced in atheroprone regions in response to low shear stress, represses endothelial expression of KLF2, KLF4, and SIRT1, and downregulates phosphatidic acid phosphatase type 2B (PPAP2B)216,233,234. Under conditions of normal laminar shear stress, PPAP2B dephosphorylates circulating lysophosphatidic acid (LPA), preventing its binding to the LPAR1 receptor, which otherwise induces pro-inflammatory signalling233 (Fig. 5e). Endothelial loss of PPAP2B leads to exacerbated local and systemic inflammation associated with an increase in endothelial permeability235. LPA signaling is involved in kidney disease through the induction of ROS, inflammatory cytokines and fibrosis236

Endothelial exposure to low shear stress and disturbed flow induces the expression of glycolytic enzymes and of pyruvate dehydrogenase kinase 1 (PDK1), thereby uncoupling glycolysis from mitochondrial metabolism and decreasing mitochondrial respiration in ECs^223,237 (Fig. 5e). Mechanistically, disturbed flow induces the production of NAD(P)H oxidase 4 (NOX4)-derived ROS and activates NF-κB, leading to upregulation and stabilization of HIF1α by preventing its degradation^223,237. Activation of the NF-κB pathway has been linked to heparinase activity and consequent glycocalyx degradation^238. In line with these findings, activation of NF-κB is repressed by chronic shear stress in the glomerular endothelium239. HIF1α activation in response to disturbed flow enhances arterial EC proliferation and expression of inflammatory markers, whereas inhibition of glycolysis prevents these responses223,237. Moreover, activation of mitochondrial oxidative phosphorylation prevents the pro-inflammatory phenotype induced by disturbed flow in arterial ECs²²³.

Disturbances in low shear stress also induce activation of the pro-inflammatory transcription factors YAP and TAZ, whereas laminar shear stress inhibits these in an integrin-dependent manner^240,241. Activation of YAP and TAZ modulates EC metabolism by stimulating glycolysis and mitochondrial activity in an MYC-dependent manner²⁴², and by upregulating glutaminolysis²⁴³. Conversely, the glycolytic enzyme PFK1 stimulates YAP and TAZ activity in a positive feedback loop244. YAP and TAZ are mechanoregulators of the TGFβ–SMAD signaling pathway in the kidney. They have been demonstrated to promote kidney fibrosis in an experimental model of unilateral ureteral obstruction, although the role of RECs in this fibrotic response as a result of endothelial-to-mesenchymal transition has been poorly investigated^245,246

Interestingly, ECs exposed to a disturbed blood flow also show activation of SREBP2, which upregulates genes involved in cholesterol synthesis including the rate-limiting enzyme of the mevalonate pathway HMG-CoA reductase (Fig. 5e) and decreases cholesterol efflux^230,247, increasing the intracellular level of cholesterol in ECs^230,247. Interestingly, inhibition of HMG-CoA reductase by statins induces an endothelial increase in KLF2 expression, and reduces pro-inflammatory signaling by NF-κB, HIF1α, and YAP–TAZ, thereby inducing an EC-like response to laminar flow^241,248,249. Furthermore, SREBP2 activation promotes the transcription of miR-92a, upregulates the expression of NOX2, which induces production of ROS, and increases the expression of the NLRP3 inflammasome, ultimately promoting endothelial inflammation and atherosclerosis216,230. Thus, SREBP2 may be one of the key drivers in the renal arterial and afferent arteriolar endothelial response to a disturbed flow.

REC responses to changes in osmolarity

The kidneys can produce urine of widely varying osmolarity depending on hydration status. The countercurrent multiplier of the loop of Henle generates a medullary osmolarity gradient that underlies the urine concentration mechanism and determines the final urine osmolarity. Briefly, the TALH — a tubule segment that is mostly impermeable to water — actively shuttles NaCl from the filtrate to the medullary interstitium, establishing a 20 mOsm/kg H2O osmolality difference across the ascending and descending flow²⁵⁰ (Fig. 6a). As an osmotic response, water is reabsorbed through the thin descending limb of Henle (TDLH), thereby increasing filtrate osmolarity250. As this filtrate progresses from the TDLH to the TALH, active reabsorption of NaCl by the TALH re-establishes the 20 mOsm/kg H2O osmolality difference between the TALH and interstitium, further increasing medullary interstitium osmolarity250. The multiplication of these small osmolality differences between countercurrent flows leads to a large corticomedullary osmolality gradient (a countercurrent multiplier)²⁵⁰. At the level of the vasa recta, water efflux facilitated by aquaporins occurs in parallel with the reabsorption of urea and NaCl within the DVR, driven by the difference between the blood and medullary osmolarity, resulting in an increased blood osmolarity towards the papilla²⁵⁰ (Fig. 6a). By contrast, the highly fenestrated AVR reabsorb medullary water and release NaCl into the interstitium, since the blood coming from the papilla is of higher osmolarity than the medullary interstitium²⁵⁰ (Fig. 6a). This countercurrent exchange between the DVR and AVR maintains the medullary osmolarity gradient created by the countercurrent multiplier system (Fig. 6a). The high osmolarity of the medulla is also sustained by the collecting ducts, which actively export urea within the inner medullary interstitium while concentrating urine according to the medullary osmolarity gradient via aquaporin-facilitated water transport. As a consequence, medullary cells including mRECs are exposed to extreme levels of hyperosmolarity, especially under conditions of dehydration wherein osmolality can rise as high as 1,400mOsm/kg in humans²⁵⁰ (Fig. 6a). As described below, available evidence suggests that mRECs have adapted to these extreme conditions by activating protective mechanisms and developing a specific metabolic profile10. Of note, other (R)ECs can be exposed to hyperosmolar conditions as a consequence of hyperglycemia in the context of diabetes mellitus²⁵¹.

The response of ECs to conditions of hyperosmolarity has been poorly investigated, with most of the research in this field focusing on renal epithelial cells, in which hyperosmolarity triggers cell cycle arrest, the production of ROS, and DNA damage²⁵². Specifically, the epithelial response to hyperosmolarity is characterized by a reorganization of the cytoskeletal actin via a process dependent on integrins and the Rho family of GTPases253, activation of the Na+ channels NHE4 (ref. 254), NKCC1, and NKCC2 (ref. 255), and the resulting influx of Na+ ions for maintenance of cell volume. These responses trigger the expression of heat shock proteins to maintain the correct folding of proteins and activation of the hyperosmolarity-sensitive transcription factor TonEBP (also known as NFAT5); under conditions of prolonged hyperosmolarity, such as in the papilla, these responses ultimately result in the accumulation of inert organic osmolytes²⁵².

Evidence from a transcriptome study in mice exposed to water deprivation suggests that a similar process occurs in the renal medullary endothelium10 (Fig. 6b). These mRECs demonstrate increased expression of genes related to glycolytic and oxidative phosphorylation pathways10 (Fig. 6b). Moreover, transcriptome analyses have suggested that these cells engage enzymatic and transport processes to aid the accumulation of inert organic osmolytes — such as the sugar derivatives, polyols (in particular, sorbitol, which is derived from the polyol pathway and Myo-inositol, which can be taken up or synthesized from glycolytic intermediates), amino acids (in particular, taurine) and possibly methylamines such as betaine and glycerophosphocholine — to balance the osmolarity between the intracellular and extracellular compartments10 (Fig. 6b). Upon prolonged water deprivation, mRECs also upregulate the Na+/K+ ATPase, presumably to limit Na+ intracellular accumulation (Fig. 6b). In this setting, oxidative phosphorylation, which yields 32 ATP molecules per glucose molecule, would represent a more efficient strategy than anaerobic glycolysis, which releases only two ATP molecules per glucose molecule, to sustain a higher Na+/K+ ATPase activity. Furthermore, some of the glycolysis intermediates that accumulate in ECs exposed to hyperosmolar conditions are likely to deviate from their glycolytic fate to enter osmolyte synthesis pathways, in particular the polyols, sorbitol, and myo-inositol¹⁰. Together, these metabolic adaptations of mRECs to changes in osmolarity would enable their survival in extreme hyperosmolar environments and enable them to sustain kidney function^10,256.

EC adaptation in tissue homeostasis

ECs are instructed by angiogenic growth factor signaling from supporting cells such as pericytes, epithelial cells, and podocytes. Conversely, the endothelium instructs its direct neighboring cells through autocrine signaling.

REC exposure to angiogenic factors

Although ECs are considered quiescent in adulthood under physiological conditions, one of the most intriguing findings obtained from the available single-cell RNA-seq studies on RECs is the presence of some typical angiogenic ECs in the kidney cortex and medulla in healthy adult mice6,10. These RECs are characterized by a genetic program resembling that of angiogenic migratory ECs (also known as tip cells) (Fig. 7a). These cells are probably exposed to angiogenic growth factors and may contribute to vascular regeneration in the adult vasculature, although their role is still unknown. Cells within the kidney can produce angiogenic growth factors, either in a constitutive manner (for example, in the case of podocyte-derived VEGFA) or in response to changes in microenvironmental conditions such as hypoxia, oxidative stress, shear stress, or inflammation. Angiogenic growth factors drive vascularization of the glomeruli and tubules during kidney development in a process that mainly depends on the cooperation of sprouting and intussusceptive angiogenesis257 (Fig. 7a,b). For instance, angiopoietins and their cognate endothelial receptor Tie2 support lymphangiogenesis and are critical for the development of the AVR in mouse embryos, and are necessary for urine concentration ability30. VEGF signaling is also required for the proper development of glomerular architecture. For instance, antibody-mediated neutralization of VEGF in newborn mice impairs vessel formation in the superficial renal cortex and glomeruli, and ultimately impairs nephron development258, whereas postnatal deletion of VEGFR2 damages the glomerular endothelium48. Moreover, loss of podocyte-derived VEGFA precludes glomerular vascularization and leads to perinatal death, whereas heterozygotic loss of Vegfa results in glomerular endothelins and proteinuria49. Podocyte-derived VEGF is therefore a critical cue for establishing the glomerular endothelium and maintaining it in adulthood257. Similarly, tubular epithelial cell-derived VEGFA is necessary for the development and maintenance of peritubular capillaries⁵⁰.

As with endothelial activation in immune activation, the physiological response of RECs to angiogenic signaling may become dysregulated in disease conditions²⁵⁹. For example, dysregulated angiogenesis is observed in rejected kidney allografts²⁶⁰. Moreover, the loss of angiogenic factors, mainly VEGF, has been implicated in capillary loss after AKI261; conversely, VEGF upregulation improves kidney function, and VEGF administration reduces capillary rarefaction in the context of ischemia-reperfusion injury^262,263. Experimental and clinical evidence suggests that in CKD, the balance between pro-angiogenic and anti-angiogenic factors is disturbed^259,264. Although CKD is characterized by the rarefaction of peritubular capillaries, which promotes kidney hypoxia, prevents the redifferentiation of regenerating tubules, and is likely to be associated with endothelial-to-mesenchymal transition, which contributes to kidney fibrosis, proangiogenic signaling in capillary gRECs favors the disruption of the filtration barrier259. For instance, evidence from animal models suggests that in early phases of diabetic kidney disease, the glomerular endothelium and ECs found within the vicinity of the glomerulus undergo VEGF-mediated abnormal angiogenesis; blockade of angiogenesis in these models improves kidney function²⁶⁵.

While little is known about intussusceptive vessel growth, sprouting angiogenesis has been intensively characterized1. The sprouting process is orchestrated through the activation of EC surface receptors and downstream signaling pathways by angiogenic growth factors, and the emergence of two major angiogenic EC phenotypes — proliferating ECs (also known as stalk cells) and migratory ECs (also known as tip cells; described in detail elsewhere^1,266,267). At the metabolic level, each angiogenic EC type has specific needs (energetic and biomass demands), according to their proliferative or migratory phenotypes, and adapts their metabolism accordingly144 (Box 1). Whether angiogenic RECs, identified by single-cell RNA-seq studies in healthy adult mice exhibit phenotypic and metabolic activity similar to that of tip cells remains to be determined. Moreover, the potential ability of metabolic strategies such as metabolic supplementation to promote angiogenesis or lymphangiogenesis for kidney repair and regeneration also requires further investigation.

Angiocrine signaling in kidney

ECs also directly instruct their neighboring cells through the release of growth factors and cytokines, and the expression of adhesion molecules268. In the kidney, the vasculature establishes a niche that sustains the development and maintenance of the kidney structure and function4,11. Direct proof for this concept is provided by studies of induced pluripotent stem cell-derived kidney organoids, in which maturation of glomerular and tubular structures can be induced by promoting vascularization and blood flow through transplantation of the organoid under the kidney capsule of mice269. In addition, disruption of the glomerular endothelium in embryonic mouse kidneys prevents podocyte maturation, in particular, the acquisition of foot processes and slit diaphragms⁴⁹. Interestingly, both the developing and adult kidney vasculature demonstrate spatial differences in the expression of endocrine factors. During the development of the glomerulus, EC-derived matrix metalloproteinase 2 seems to regulate the interaction of ECs with podocytes for glomerular assembly²⁷⁰, whereas expression of PDGFB stimulates the recruitment of mesangial cells^271,272. The developing glomerular endothelium also expresses growth factor genes such as Fgf1 and Vegfa, the Notch ligand Dll4, the neuronal guidance cue Sema5a, and the Wnt antagonist Dkk2 (refs4,11); Dkk2 is also enriched in afferent arteriole gRECs that are associated with the JGA in adulthood10. The developing renal arteries and arterioles also express growth factors, including Pdgfb, Tgfb2, and Ltbp4 (ref. 11); expression of Tgfb2 and Ltbp4 remain enriched in the same REC subsets in adulthood10. Strikingly, the insulin-like growth factor signaling system shows a clear zonation within the adult kidney vasculature: Igf1 is mainly expressed in the kidney medulla by RECs of the AVR in the papilla, Igfbp7 is considered a broad marker of mRECs, Igfbp4 is found in the afferent arteriolar gRECs, Igfbp3 is mostly expressed by peritubular capillary cRECs and Igfbp5 is enriched in both cortical peritubular and glomerular capillaries10,11. Multiple other endocrine factors also show heterogeneous expression among the different EC phenotypes in adult mouse kidneys10. Their roles in the maintenance of kidney tissue and functions remain to be investigated.

Angiocrine signaling is also critical for the repair processes after kidney injury. For instance, RECs may contribute to renal recovery after AKI by regulating inflammatory responses or providing pro-tubulogenic signals²⁷³. In cocultures, renal proximal tubule cells stimulate RECs to express a number of angiocrine factors, including VEGF, TGFβ, its antagonist α2-macroglobulin and hepatocyte growth factor, which in turn promote the survival, proliferation, and differentiation of the proximal tubule cells274. It is likely that capillary rarefaction induces ‘chronic injury–dysregulated repair’ processes, which result in impaired epithelial regeneration, nephron loss, and pro-inflammatory and fibrotic signals, possibly due to insufficient angiocrine signals273. In support of this hypothesis is evidence demonstrating that endothelial progenitor cell extracts possess renoprotective activity in animal models of AKI by preventing capillary loss and reducing fibrosis, with endocrine factors such as leukemia inhibitory factor preventing and reversing fibroblast-to-myofibroblast transition²⁷⁵.

Further investigation of the physiological and pathological microenvironments that shape the endocrine profiles of RECs is warranted. Moreover, the mechanisms by which REC metabolic processes interlink with these distinct endocrine signatures remain to be determined. Interestingly, a 2020 study described endocrine metabolite communication in muscle regeneration after ischemia. That study demonstrated that EC-derived lactate induces macrophages to adopt a pro-regenerative M2-like phenotype, raising the question of whether a similar mechanism might occur in the kidney to promote regeneration after ischaemic insults²⁷⁶.

Conclusions

The mammalian vascular system has long been considered to be a passive conduit for the delivery of oxygen and nutrients to the different organs, as well as for waste removal, with the endothelium serving as a blood–tissue barrier. This paradigm is no longer valid since intensive research in the vascular field has revealed an active role for the endothelium not only in the regulation of vasotone and inflammation, but also in organ maturation through the production of endocrine mediators, immunity, and organ function. The importance of ECs to these processes is evidenced by the fact that each organ benefits from a unique endothelium, tailored to its specific needs, and shaped by the cellular and non-cellular microenvironment.

Endothelium in the kidney is unique to that organ and exhibits vast heterogeneity to sustain the multiple activities of the kidney in fluid and blood pressure homeostasis. Importantly, REC dysfunction is not only associated with kidney disease but can drive disease progression, identifying the kidney endothelium as a relevant therapeutic target. In the past decade, extensive analyses have revealed that the metabolism of ECs is tightly regulated at the cellular and subcellular levels and that different EC phenotypes display unique metabolic profiles. The plasticity of EC metabolism provides a mechanism to orchestrate EC phenotypic behavior, enabling ECs to actively respond to changes in their microenvironment, but also providing a potential opportunity for therapeutic targeting. For example, inhibition of endothelial PFKFB3 in tumor ECs normalized glycolytic flux and improved vessel maturation are associated with decreased endothelial activation and tightening of the endothelial barrier²⁷⁷. Moreover, supplementation of ECs with the acetyl-CoA precursor, acetate, promotes (lymph)angiogenesis, in vitro and in vivo^278–280. Specific diets may also offer new therapeutic avenues: a 2019 study demonstrated that a ketogenic diet promotes lymphangiogenesis by increasing the pool of acetyl-CoA in LECs, and decreasing lymphoedema in an animal model²⁸¹

Similarly, in the kidney, the phenotypic heterogeneity of RECs is thought to be driven by metabolic adaptations to cellular needs and signals from the microenvironment in both health and disease. Metabolic targeting of endothelial dysfunction, as well as angiogenesis and lymphangiogenesis, are potential strategies to improve kidney function in the context of kidney disease. Kidney organoids have been proposed as a regenerative approach to replace kidney function but their lack of developed vasculature is a major limitation^282–284. Strategies (including metabolic strategies) to stimulate kidney organoid vascularization and promote EC maturation may promote the maturation of these organoids and ultimately facilitate their therapeutic use²⁶⁹. Despite the need for further investigation into the specificities of the renal endothelium, the available evidence supports a key role for these cells in kidney physiological processes and suggests that (metabolic) targeting of RECs could be a highly beneficial strategy for promoting renal recovery and regeneration.

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