Biology subjects

Carmeliet, P.

Publications and source records attributed to Carmeliet, P..

11 recordsLinked to original sources

bioRxiv2026

BfBio: a graph-based tool for the prediction of Angiogenic Stalk Cell genes using a Personalized PageRank algorithm

Although most human protein coding genes have functional annotations in databases, such as GeneCards, many remain poorly characterized. To address this gap, computational tools can be leveraged to predict the functional roles of under-annotated genes by extracting patterns from complex biological networks. Here we introduce Brain-for-Biotech (BfBio), a framework designed to identify genes important for vascular endothelial cells (EC), which are crucial cells for vessel formation (angiogenesis), vascular homeostasis, hemostasis and blood/tissue barrier function but also critical mediators of immunity and cancer progression. BfBio utilizes a Personalized PageRank (PPR) algorithm on an integrated network of different omics datasets and publicly available gene-gene/protein-protein interaction databases. In this study, we apply the predictive capabilities of BfBio to infer angiogenic stalk cell phenotype function in genes for which this function was not known before. By leveraging a set of genes characterizing the stalk cell cluster in lung tumor EC models previously identified, we have achieved a high Area Under Receiver Operative Characteristic (AUC-ROC) performance (0.837). Enrichment analysis, coupled with a text mining application, further confirmed that among the 49 predicted genes four of them were poorly characterized yet possessed biologically relevant properties and were linked to cancer, thereby validating BfBio as a robust tool for prioritizing novel therapeutic targets in vascular biology.

cancer biology↗

bioRxiv2025

Astrocytic glycolysis attenuates mitochondrial efficiency to preserve cognition

Astrocytic glycolysis is tightly coupled to neurotransmission and thought to be essential for neurological health. However, the metabolic adaptations that enable astrocytes to maintain a durable glycolytic profile without compromising viability are elusive. Here, using in vivo approaches including cell-specific gene expression disruption, metabolic flux analyses and behavioral tests in mice, we addressed this issue. We found that Pfkfb3 (6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-3) is instrumental in maintaining the astrocytic glycolytic phenotype. Importantly, astrocytic glycolysis sustained by Pfkfb3 is required for normal cognitive performance. Mechanistically, ATP generated through glycolysis is consumed by mitochondria, via the reverse mode of ATP synthase, to conserve the proton gradient across the inner mitochondrial membrane. This enables mitochondria to attenuate pyruvate decarboxylation, tricarboxylic acid cycle and electron transport chain activity, thereby preserving pyruvate for conversion into lactate and delivery to neurons. These findings reveal that astrocytes sacrifice mitochondrial bioenergetic efficiency as a previously underappreciated strategy to support cognition.

cell biology↗

bioRxiv2025

Hypernatremia Enhances Transient Outward Potassium and Late Sodium Currents in a Mouse Model of Long QT Syndrome Type 3

Cardiac voltage-gated sodium channel gain-of-function (NavGOF) is characterized by action potential duration (APD) prolongation. Hypernatremia and perinexal widening synergistically prolong cardiac APD in guinea pig. However, guinea pig lack the transient outward potassium current (Ito), which could be increased by hypernatremia and thereby shorten APD. ObjectiveDetermine whether hypernatremia and perinexal expansion synergistically prolong APD in an animal model functionally expressing Ito. MethodsWhole-cell Ito was measured in isolated genetically-modified NavGOF ({Delta}KPQ) mouse ventricular myocytes. Ventricular APD at 30 (APD30) and 90 (APD90) percent repolarization were measured from optically mapped, Langendorff-perfused wild-type (WT) and {Delta}KPQ mouse hearts at different perfusate sodium concentrations (145 or 160mM), without and with the perinexal adhesion antagonist {beta}adp1. ResultsIn isolated myocytes, hypernatremia (160mM sodium) increased Ito. In whole-heart, hypernatremia significantly decreased both APD30 and APD90 in WT but only APD30 in {Delta}KPQ preparations. Perinexal disruption with {beta}adp1 did not change APD30 or APD90 in WT hearts, however it decreased APD30 and increased APD90 in {Delta}KPQ hearts. Combination of hypernatremia and {beta}adp1 did not synergistically change APD in {Delta}KPQ hearts. Computational models predict that Ito activation can prevent synergistic APD prolongation in mouse during hypernatremia and perinexal expansion that was observed previously in a guinea pig NavGOF model lacking Ito. ConclusionsHypernatremia during NavGOF prevents early ventricular repolarization due to Ito activation (mouse) and prolongs repolarization in the absence of Ito (guinea pig). Future studies in animal models electrophysiologically similar to humans are needed to determine if hypernatremia and perinexal expansion are proarrhythmic during NavGOF.

physiology↗

bioRxiv2024

Prenatal VEGF Nano-Delivery Reverses Congenital Diaphragmatic Hernia-Associated Pulmonary Abnormalities

RationaleCongenital diaphragmatic hernia (CDH) results in lung hypoplasia. In severe cases, tracheal occlusion (TO) can be offered to promote lung growth. However the benefit is limited, and novel treatments are required to supplement TO. Vascular endothelial growth factor (VEGF) is downregulated in animal models of CDH and could be a therapeutic target, but its role in human CDH is not known. ObjectivesTo investigate whether VEGF supplementation could be a suitable treatment for CDH-associated lung pathology. MethodsFetal lungs from CDH patients were used to determine pulmonary morphology and VEGF expression. A novel human ex vivo model of fetal lung compression recapitulating CDH features was developed and used to determine the effect of exogenous VEGF supplementation (Figure 1A). A nanoparticle-based approach for intra-pulmonary delivery of VEGF was developed by conjugating it on functionalized nanodiamonds (ND-VEGF) and was tested in experimental CDH in vivo. O_FIG O_LINKSMALLFIG WIDTH=138 HEIGHT=200 SRC="FIGDIR/small/581170v1_fig1.gif" ALT="Figure 1"> View larger version (77K): org.highwire.dtl.DTLVardef@2ba882org.highwire.dtl.DTLVardef@4413d2org.highwire.dtl.DTLVardef@17155aborg.highwire.dtl.DTLVardef@1af7c73_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 1C_FLOATNO (A) Schematic of the study design based on the analysis of the early-stage CDH fetal lung sections to assess early hypoplasia and VEGF impairment. In parallel, a human compression-based CDH model will be developed to validate the CDH phenotype and test a therapeutical approach to rescue lung hypoplasia. (B) H&E analysis of 18 and 22 pcw post-mortem CDH human lung tissue reveals impaired epithelial development compared to control healthy tissue at matching developmental stages. Scale bar 100m. (C) Morphometric analysis of CDH and healthy lung tissue at 18-22 pcw reveals decreased values of volume densities of air space (Vair%) and corresponding higher values of volume densities of alveolar septa (Vsep%) in CDH tissues compared to healthy tissue at the same developmental stages. A similar trend can be observed for the mean linear intercept of air space (Lma%) and its corresponding mean linear intercept of septal thickness (Lmw%). (D) Histologic analysis of VEGFA, KDR, Ki67 and NKX2-1 markers in normal and CDH fetal lung tissue at 18 and 22 pcw. Scale bar 100m. (E) Quantification of epithelial VEGFA and KDR expression reported in D shows a significant decrease for both markers in CDH compared to healthy samples. Correspondingly, the percentage of Ki67-positive epithelial cells is significantly lower in CDH compared to healthy samples, with no significant differences in the number of NKX2-1-positive cells. ***<p=0.001. (F) Immunofluorescence analysis of ECAD in normal and CDH fetal lung tissue at 18 and 22 pcw. Scale bar 100m. (G) Quantification of epithelial nuclear density based on results in Figure F shows no significant differences between CDH and healthy tissue. C_FIG Measurements and Main ResultsVEGF expression was downregulated in distal pulmonary epithelium of human CDH fetuses in conjunction with attenuated cell proliferation. The compression model resulted in impaired branching morphogenesis similar to CDH and downregulation of VEGF expression in conjunction with reduced proliferation of terminal bud epithelial progenitors; these could be reversed by exogenous supplementation of VEGF. Prenatal delivery of VEGF with the ND-VEGF platform in CDH fetal rats resulted in lung growth and pulmonary arterial remodelling that was complementary to that achieved by TO alone with appearances comparable to healthy controls. ConclusionsThis innovative approach could have a significant impact on the treatment of CDH.

bioengineering↗

bioRxiv2023

Weak neuronal glycolysis sustains cognition and organismal fitness

The energy cost of neuronal activity is mainly sustained by glucose1,2. However, in an apparent paradox, neurons only weakly metabolize glucose through glycolysis3,4,5,6, a circumstance that can be accounted for by the constant degradation of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-3 (Pfkfb3)3,7,8, a key glycolysis-promoting enzyme. To evaluate the in vivo physiological significance of this hypo-glycolytic metabolism, here we genetically engineered mice with their neurons transformed into active glycolytic cells through Pfkfb3 expression. In vivo molecular, biochemical, and metabolic flux analyses of these neurons revealed an accumulation of anomalous mitochondria, complex I disassembly, bioenergetic deficiency and mitochondrial redox stress. Notably, glycolysis-mediated NAD+ reduction impaired sirtuin-dependent autophagy. Furthermore, these mice displayed cognitive decline and a metabolic syndrome that was mimicked by confining Pfkfb3 expression to hypothalamic neurons. Neuron-specific genetic ablation of mitochondrial redox stress corrected these alterations. Thus, the weak glycolytic nature of neurons is required to sustain higher-order organismal functions.

cell biology↗

bioRxiv2021

Molecular atlas of the human brain vasculature at the single-cell level

A broad range of brain pathologies critically relies on the vasculature, and cerebrovascular disease is a leading cause of death worldwide. However, the cellular and molecular architecture of the human brain vasculature remains poorly understood. Here, we performed single-cell RNA sequencing of 599,215 freshly isolated endothelial, perivascular and other tissue-derived cells from 47 fetuses and adult patients to construct a molecular atlas of the developing fetal, adult control and diseased human brain vasculature. We uncover extensive molecular heterogeneity of healthy fetal and adult human brains and across eight vascular-dependent CNS pathologies including brain tumors and brain vascular malformations. We identify alteration of arteriovenous differentiation and reactivated fetal as well as conserved dysregulated pathways in the diseased vasculature. Pathological endothelial cells display a loss of CNS-specific properties and reveal an upregulation of MHC class II molecules, indicating atypical features of CNS endothelial cells. Cell-cell interaction analyses predict numerous endothelial-to-perivascular cell ligand-receptor crosstalk including immune-related and angiogenic pathways, thereby unraveling a central role for the endothelium within brain neurovascular unit signaling networks. Our single-cell brain atlas provides insight into the molecular architecture and heterogeneity of the developing, adult/control and diseased human brain vasculature and serves as a powerful reference for future studies.

neuroscience↗

bioRxiv2021

CD8+ T Cell Metabolic Rewiring Defined by Single-Cell RNA-Sequencing Identifies a Critical Role of ASNS Expression Dynamics in T Cell Differentiation

Cytotoxic T cells dynamically rewire their metabolism during the course of an immune response. While T-cell metabolism has been extensively studied at phenotypic endpoints of activation and differentiation, the underlying dynamics remain largely elusive. Here, we leverage on single-cell RNA-sequencing (scRNA-seq) measurements of in vitro activated and differentiated CD8+ T cells cultured in physiological media to resolve these metabolic dynamics. We find that our scRNA-seq analysis identifies most metabolic changes previously defined in in vivo experiments, such as a rewiring from an oxidative to an anabolism-promoting metabolic program during activation to an effector state, which is later reverted upon memory polarization. Importantly, our scRNA-seq data further provide a dynamic description of these changes. In this sense, our data predict a differential time-dependent reliance of CD8+ T cells on the synthesis versus uptake of various non-essential amino acids during T-cell activation, which we corroborate with additional functional in vitro experiments. We further exploit our scRNA-seq data to identify metabolic genes that could potentially dictate the outcome of T-cell differentiation, by ranking them based on their expression dynamics. Among the highest-ranked hits, we find asparagine synthetase (Asns), whose expression sharply peaks for effector CD8+ T cells and further decays towards memory polarization. We then confirm that these in vitro Asns expression dynamics are representative of an in vivo situation in a mouse model of viral infection. Moreover, we find that disrupting these expression dynamics in vitro, by depleting asparagine from the culture media, delays central-memory polarization. Accordingly, we find that preventing the decay of ASNS by stable overexpression at the protein level in vivo leads to a significant increase in effector CD8+ T-cell expansion, and a concomitant decrease in central-memory formation, in a mouse model of viral infection. This shows that ASNS expression dynamics dictate the fate of CD8+ T-cell differentiation. In conclusion, we provide a resource of dynamic expression changes during CD8+ T-cell activation and differentiation that is expected to increase our understanding of the dynamic metabolic requirements of T cells progressing along the immune response cascade.

immunology↗

bioRxiv2021

Repression of Hypoxia-Inducible Factor-1 Contributes to Increased Mitochondrial Reactive Oxygen Species Production in Diabetes

BackgroundExcessive production of mitochondrial reactive oxygen species (ROS) is a central mechanism for the development of diabetes complications. Recently, hypoxia has been identified to play an additional pathogenic role in diabetes. In this study, we hypothesized that ROS overproduction was secondary to the impaired responses to hypoxia due to the inhibition of hypoxia-inducible factor-1 (HIF-1) by hyperglycemia. MethodsThe dynamic of ROS levels was analysed in the blood of healthy subjects and individuals with type 1 diabetes after exposure to hypoxia (ClinicalTrials.gov registration no. NCT02629406). The relation between HIF-1, glucose levels, ROS production and its functional consequences were analyzed in renal mIMCD-3 cells and in kidneys of mouse models of diabetes. ResultsExposure to hypoxia increased circulating ROS in subjects with diabetes, but not in subjects without diabetes. High glucose concentrations repressed HIF-1 both in hypoxic cells and in kidneys of animals with diabetes, through a HIF prolyl-hydroxylase (PHD) - dependent mechanism. The impaired HIF-1 signaling contributed to excess production of mitochondrial ROS through increased mitochondrial respiration that was mediated by Pyruvate dehydrogenase kinase 1 (PDK1) and was followed by functional consequences. The restoration of HIF-1 function attenuated ROS overproduction despite persistent hyperglycemia, and conferred protection against apoptosis and renal injury in diabetes. ConclusionsWe conclude that the repression of HIF-1 plays a central role in mitochondrial ROS overproduction in diabetes and is a potential therapeutic target for diabetic complications. These findings are highly significant and timely since the first PHD inhibitor that can activate HIF-1 has been newly approved for clinical use.

cell biology↗

bioRxiv2021

Heterogeneity in PHGDH protein expression potentiates cancer cell dissemination and metastasis

Cancer metastasis requires the transient activation of cellular programs enabling dissemination and seeding in distant organs. Genetic, transcriptional and translational intra-tumor heterogeneity contributes to this dynamic process. Beyond this, metabolic intra-tumor heterogeneity has also been observed, yet its role for cancer progression remains largely elusive. Here, we discovered that intra-tumor heterogeneity in phosphoglycerate dehydrogenase (PHGDH) protein expression drives breast cancer cell dissemination and metastasis formation. Specifically, we observed intra-tumor heterogeneous PHGDH expression in primary breast tumors, with low PHGDH expression being indicative of metastasis in patients. In mice, Phgdh protein, but not mRNA, expression is low in circulating tumor cells and early metastatic lesions, leading to increased dissemination and metastasis formation. Mechanistically, low PHGDH protein expression induces an imbalance in glycolysis that can activate sialic acid synthesis. Consequently, cancer cells undergo a partial EMT and show increased p38 as well as SRC phosphorylation, which activate cellular programs of dissemination. In turn, inhibition of sialic acid synthesis through knock-out of cytidine monophosphate N-acetylneuraminic acid synthetase (CMAS) counteracts the increased cancer cell dissemination and metastasis induced by low PHGDH expression. In conclusion, we find that heterogeneity in PHGDH protein expression promotes cancer cell dissemination and metastasis formation.

cancer biology↗

bioRxiv2020

Hierarchical imaging and computational analysis of three-dimensional vascular network architecture in the entire postnatal and adult mouse brain

The formation of new blood vessels and the establishment of vascular networks are crucial during brain development, in the adult healthy brain, as well as in various diseases of the central nervous system (CNS). Here, we describe a method that enables hierarchical imaging and computational analysis of vascular networks in postnatal- and adult mouse brains. Resin-based vascular corrosion casting, scanning electron microscopy, synchrotron radiation and desktop {micro}CT imaging, and computational network analysis are used. Combining these methods enables detailed visualization and quantification of the three-dimensional (3D) brain vasculature. Network features such as vascular volume fraction, branch point density, vessel diameter, - length, -tortuosity, and -directionality as well as extravascular distance can be obtained at any developmental stage from the early postnatal to the adult brain. Our method allows characterizing brain vascular networks separately for capillaries and non-capillaries. The entire protocol, from mouse perfusion to vessel network analysis, takes approximately 10 days. Online summaryThis protocol uses vascular corrosion casting, hierarchical synchrotron radiation {micro}CT imaging, and computational image analysis to assess the three-dimensional vascular network architecture.

neuroscience↗

bioRxiv2020

DNA Methylation Repels Binding of HIF Transcription Factors to Maintain Tumour Immunotolerance

AO_SCPLOWBSTRACTC_SCPLOWO_ST_ABSBackgroundC_ST_ABSHypoxia is pervasive in cancer and other diseases. Cells sense and adapt to hypoxia by activating hypoxia-inducible transcription factors (HIFs), but it is still an outstanding question why cell types differ in their transcriptional response to hypoxia. ResultsHere, we report that HIFs fail to bind CpG dinucleotides that are methylated in their consensus binding sequence, both in in vitro biochemical binding assays and in vivo studies of differentially methylated isogenic cell lines. Based on in silico structural modelling, we show that 5-methylcytosine indeed causes steric hindrance in the HIF binding pocket. A model wherein cell-type-specific methylation landscapes, as laid-down by the differential expression and binding of other transcription factors under normoxia control cell-type-specific hypoxia responses is observed. We also discover ectopic HIF binding sites in repeat regions which are normally methylated. Genetic and pharmacological DNA demethylation, but also cancer-associated DNA hypomethylation, expose these binding sites, inducing HIF-dependent expression of cryptic transcripts. In line with such cryptic transcripts being more prone to cause double-stranded RNA and viral mimicry, we observe low DNA methylation and high cryptic transcript expression in tumours with high immune checkpoint expression, but not in tumours with low immune checkpoint expression, where they would compromise tumour immunotolerance. In a low-immunogenic tumour model, DNA demethylation upregulates cryptic transcript expression in a HIF-dependent manner, causing immune activation and reducing tumour growth. ConclusionsOur data elucidate the mechanism underlying cell-type specific responses to hypoxia, and suggest DNA methylation and hypoxia to underlie tumour immunotolerance.

genomics↗