João Pinho Monteiro

All Publications

• A cell and transcriptome atlas of the human arterial vasculature. bioRxiv : the preprint server for biology Zhao, Q., Pedroza, A., Sharma, D., Gu, W., Dalal, A., Weldy, C., Jackson, W., Li, D. Y., Ryan, Y., Nguyen, T., Shad, R., Palmisano, B. T., Monteiro, J. P., Worssam, M., Berezwitz, A., Iyer, M., Shi, H., Kundu, R., Limbu, L., Kim, J. B., Kundaje, A., Fischbein, M., Wirka, R., Quertermous, T., Cheng, P. 2024

  Abstract

  Vascular beds show different propensities for different vascular pathologies, yet mechanisms explaining these fundamental differences remain unknown. We sought to build a transcriptomic, cellular, and spatial atlas of human arterial cells across multiple different arterial segments to understand this phenomenon. We found significant cell type-specific segmental heterogeneity. Determinants of arterial identity are predominantly encoded in fibroblasts and smooth muscle cells, and their differentially expressed genes are particularly enriched for vascular disease-associated loci and genes. Adventitial fibroblast-specific heterogeneity in gene expression coincides with numerous vascular disease risk genes, suggesting a previously unrecognized role for this cell type in disease risk. Adult arterial cells from different segments cluster not by anatomical proximity but by embryonic origin, with differentially regulated genes heavily influenced by developmental master regulators. Non-coding transcriptomes across arterial cells contain extensive variation in lnc-RNAs expressed in cell type- and segment-specific patterns, rivaling heterogeneity in protein coding transcriptomes, and show enrichment for non-coding genetic signals for vascular diseases.

  View details for DOI 10.1101/2024.09.10.612293

  View details for PubMedID 39314359

  View details for PubMedCentralID PMC11419041

• Deciphering the impact of genomic variation on function. Nature 2024; 633 (8028): 47-57

  Abstract

  Our genomes influence nearly every aspect of human biology-from molecular and cellular functions to phenotypes in health and disease. Studying the differences in DNA sequence between individuals (genomic variation) could reveal previously unknown mechanisms of human biology, uncover the basis of genetic predispositions to diseases, and guide the development of new diagnostic tools and therapeutic agents. Yet, understanding how genomic variation alters genome function to influence phenotype has proved challenging. To unlock these insights, we need a systematic and comprehensive catalogue of genome function and the molecular and cellular effects of genomic variants. Towards this goal, the Impact of Genomic Variation on Function (IGVF) Consortium will combine approaches in single-cell mapping, genomic perturbations and predictive modelling to investigate the relationships among genomic variation, genome function and phenotypes. IGVF will create maps across hundreds of cell types and states describing how coding variants alter protein activity, how noncoding variants change the regulation of gene expression, and how such effects connect through gene-regulatory and protein-interaction networks. These experimental data, computational predictions and accompanying standards and pipelines will be integrated into an open resource that will catalyse community efforts to explore how our genomes influence biology and disease across populations.

  View details for DOI 10.1038/s41586-024-07510-0

  View details for PubMedID 39232149

  View details for PubMedCentralID 7405896

• Smooth muscle expression of RNA editing enzyme ADAR1 controls vascular integrity and progression of atherosclerosis. bioRxiv : the preprint server for biology Weldy, C. S., Li, Q., Monteiro, J. P., Guo, H., Galls, D., Gu, W., Cheng, P. P., Ramste, M., Li, D., Palmisano, B. T., Sharma, D., Worssam, M. D., Zhao, Q., Bhate, A., Kundu, R. K., Nguyen, T., Li, J. B., Quertermous, T. 2024

  Abstract

  Mapping the genomic architecture of complex disease has been predicated on the understanding that genetic variants influence disease risk through modifying gene expression. However, recent discoveries have revealed that a significant burden of disease heritability in common autoinflammatory disorders and coronary artery disease is mediated through genetic variation modifying post-transcriptional modification of RNA through adenosine-to-inosine (A-to-I) RNA editing. This common RNA modification is catalyzed by ADAR enzymes, where ADAR1 edits specific immunogenic double stranded RNA (dsRNA) to prevent activation of the double strand RNA (dsRNA) sensor MDA5 ( IFIH1 ) and stimulation of an interferon stimulated gene (ISG) response. Multiple lines of human genetic data indicate impaired RNA editing and increased dsRNA sensing to be an important mechanism of coronary artery disease (CAD) risk. Here, we provide a crucial link between observations in human genetics and mechanistic cell biology leading to progression of CAD. Through analysis of human atherosclerotic plaque, we implicate the vascular smooth muscle cell (SMC) to have a unique requirement for RNA editing, and that ISG induction occurs in SMC phenotypic modulation, implicating MDA5 activation. Through culture of human coronary artery SMCs, generation of a conditional SMC specific Adar1 deletion mouse model on a pro-atherosclerosis background, and with incorporation of single cell RNA sequencing cellular profiling, we further show that Adar1 controls SMC phenotypic state, is required to maintain vascular integrity, and controls progression of atherosclerosis and vascular calcification. Through this work, we describe a fundamental mechanism of CAD, where cell type and context specific RNA editing and sensing of dsRNA mediates disease progression, bridging our understanding of human genetics and disease causality.

  View details for DOI 10.1101/2024.07.08.602569

  View details for PubMedID 39026721

  View details for PubMedCentralID PMC11257488

• Genome-Wide Genetic Associations Prioritize Evaluation of Causal Mechanisms of Atherosclerotic Disease Risk. Arteriosclerosis, thrombosis, and vascular biology Quertermous, T., Li, D. Y., Weldy, C. S., Ramste, M., Sharma, D., Monteiro, J. P., Gu, W., Worssam, M. D., Palmisano, B. T., Park, C. Y., Cheng, P. 2024; 44 (2): 323-327

  Abstract

  The goal of this review is to discuss the implementation of genome-wide association studies to identify causal mechanisms of vascular disease risk.The history of genome-wide association studies is described, the use of imputation and the creation of consortia to conduct meta-analyses with sufficient power to arrive at consistent associated loci for vascular disease. Genomic methods are described that allow the identification of causal variants and causal genes and how they impact the disease process. The power of single-cell analyses to promote genome-wide association studies of causal gene function is described.Genome-wide association studies represent a paradigm shift in the study of cardiovascular disease, providing identification of genes, cellular phenotypes, and disease pathways that empower the future of targeted drug development.

  View details for DOI 10.1161/ATVBAHA.123.319480

  View details for PubMedID 38266112

• Extracellular vesicles from differentiated stem cells contain novel proangiogenic miRNAs and induce angiogenic responses at low doses. Molecular therapy : the journal of the American Society of Gene Therapy Kesidou, D., Bennett, M., Monteiro, J. P., McCracken, I. R., Klimi, E., Rodor, J., Condie, A., Cowan, S., Caporali, A., Wit, J. B., Mountford, J. C., Brittan, M., Beqqali, A., Baker, A. H. 2023

  Abstract

  Extracellular vesicles (EVs) released from healthy endothelial cells (ECs) have shown potential for promoting angiogenesis, but their therapeutic efficacy remains poorly understood. We have previously shown that transplantation of a human embryonic stem cell-derived endothelial cell product (hESC-ECP), promotes new vessel formation in acute ischemic disease in mice, likely via paracrine mechanism(s). Here, we demonstrated that EVs from hESC-ECPs (hESC-eEVs) significantly increased EC tube formation and wound closure in vitro at ultralow doses, whereas higher doses were ineffective. More important, EVs isolated from the mesodermal stage of the differentiation (hESC-mEVs) had no effect. Small RNA sequencing revealed that hESC-eEVs have a unique transcriptomic profile and are enriched in known proangiogenic microRNAs (miRNAs, miRs). Moreover, an in silico analysis identified three novel hESC-eEV-miRNAs with potential proangiogenic function. Differential expression analysis suggested that two of those, miR-4496 and miR-4691-5p, are highly enriched in hESC-eEVs. Overexpression of miR-4496 or miR-4691-5p resulted in increased EC tube formation and wound closure in vitro, validating the novel proangiogenic function of these miRNAs. In summary, we demonstrated that hESC-eEVs are potent inducers of EC angiogenic response at ultralow doses and contain a unique EV-associated miRNA repertoire, including miR-4496 and miR-4691-5p, with novel proangiogenic function.

  View details for DOI 10.1016/j.ymthe.2023.11.023

  View details for PubMedID 38096818

• Discovery of Transacting Long Noncoding RNAs That Regulate Smooth Muscle Cell Phenotype. Circulation research Shi, H., Nguyen, T., Zhao, Q., Cheng, P., Sharma, D., Kim, H. J., Brian Kim, J., Wirka, R., Weldy, C. S., Monteiro, J. P., Quertermous, T. 2023

  Abstract

  Smooth muscle cells (SMCs), the major cell type in atherosclerotic plaques, are vital in coronary artery diseases (CADs). Smooth muscle cell (SMC) phenotypic transition, which leads to the formation of various cell types in atherosclerotic plaques, is regulated by a network of genetic and epigenetic mechanisms and governs the risk of disease. The involvement of long noncoding RNAs (lncRNAs) has been increasingly identified in cardiovascular disease. However, SMC lncRNAs have not been comprehensively characterized, and their regulatory role in SMC state transition remains unknown.A discovery pipeline was constructed and applied to deeply strand-specific RNA sequencing from perturbed human coronary artery SMC with different disease-related stimuli, to allow for the detection of novel lncRNAs. The functional relevance of a select few novel lncRNAs were verified in vitro.We identified 4579 known and 13 655 de novo lncRNAs in human coronary artery SMC. Consistent with previous long noncoding RNA studies, these lncRNAs overall have fewer exons, are shorter in length than protein-coding genes (pcGenes), and have relatively low expression level. Genomic location of these long noncoding RNA is disproportionately enriched near CAD-related TFs (transcription factors), genetic loci, and gene regulators of SMC identity, suggesting the importance of their function in disease. Two de novo lncRNAs, ZEB-interacting suppressor (ZIPPOR) and TNS1-antisense (TNS1-AS2), were identified by our screen. Combining transcriptional data and in silico modeling along with in vitro validation, we identified CAD gene ZEB2 as a target through which these lncRNAs exert their function in SMC phenotypic transition.Expression of a large and diverse set of lncRNAs in human coronary artery SMC are highly dynamic in response to CAD-related stimuli. The dynamic changes in expression of these lncRNAs correspond to alterations in transcriptional programs that are relevant to CAD, suggesting a critical role for lncRNAs in SMC phenotypic transition and human atherosclerotic disease.

  View details for DOI 10.1161/CIRCRESAHA.122.321960

  View details for PubMedID 36852690

• Molecular mechanisms of coronary artery disease risk at the PDGFD locus. Nature communications Kim, H., Cheng, P., Travisano, S., Weldy, C., Monteiro, J. P., Kundu, R., Nguyen, T., Sharma, D., Shi, H., Lin, Y., Liu, B., Haldar, S., Jackson, S., Quertermous, T. 2023; 14 (1): 847

  Abstract

  Genome wide association studies for coronary artery disease (CAD) have identified a risk locus at 11q22.3. Here, we verify with mechanistic studies that rs2019090 and PDGFD represent the functional variant and gene at this locus. Further, FOXC1/C2 transcription factor binding at rs2019090 is shown to promote PDGFD transcription through the CAD promoting allele. With single cell transcriptomic and histology studies with Pdgfd knockdown in an SMC lineage tracing male atherosclerosis mouse model we find that Pdgfd promotes expansion, migration, and transition of SMC lineage cells to the chondromyocyte phenotype. Pdgfd also increases adventitial fibroblast and pericyte expression of chemokines and leukocyte adhesion molecules, which is linked to plaque macrophage recruitment. Despite these changes there is no effect of Pdgfd deletion on overall plaque burden. These findings suggest that PDGFD mediates CAD risk by promoting deleterious phenotypic changes in SMC, along with an inflammatory response that is primarily focused in the adventitia.

  View details for DOI 10.1038/s41467-023-36518-9

  View details for PubMedID 36792607

• Molecular mechanisms of coronary artery disease risk at the PDGFD locus. bioRxiv : the preprint server for biology Kim, H., Cheng, P., Travisano, S., Weldy, C., Monteiro, J. O., Kundu, R., Nguyen, T., Sharma, D., Shi, H., Lin, Y., Liu, B., Haldar, S., Jackson, S., Quertermous, T. 2023

  Abstract

  Platelet derived growth factor (PDGF) signaling has been extensively studied in the context of vascular disease, but the genetics of this pathway remain to be established. Genome wide association studies (GWAS) for coronary artery disease (CAD) have identified a risk locus at 11q22.3, and we have verified with fine mapping approaches that the regulatory variant rs2019090 and PDGFD represent the functional variant and putative functional gene. Further, FOXC1/C2 transcription factor (TF) binding at rs2019090 was found to promote PDGFD transcription through the CAD promoting allele. Employing a constitutive Pdgfd knockout allele along with SMC lineage tracing in a male atherosclerosis mouse model we mapped single cell transcriptomic, cell state, and lesion anatomical changes associated with gene loss. These studies revealed that Pdgfd promotes expansion, migration, and transition of SMC lineage cells to the chondromyocyte phenotype and vascular calcification. This is in contrast to protective CAD genes TCF21 , ZEB2 , and SMAD3 which we have shown to promote the fibroblast-like cell transition or perturb the pattern or extent of transition to the chondromyocyte phenotype. Further, Pdgfd expressing fibroblasts and pericytes exhibited greater expression of chemokines and leukocyte adhesion molecules, consistent with observed increased macrophage recruitment to the plaque. Despite these changes there was no effect of Pdgfd deletion on SMC contribution to the fibrous cap or overall lesion burden. These findings suggest that PDGFD mediates CAD risk through promoting SMC expansion and migration, in conjunction with deleterious phenotypic changes, and through promoting an inflammatory response that is primarily focused in the adventitia where it contributes to leukocyte trafficking to the diseased vessel wall.

  View details for DOI 10.1101/2023.01.26.525789

  View details for PubMedID 36747745

• From novel discovery tools and biomarkers to precision medicine - basic cardiovascular science highlights of 2021/2022. Cardiovascular research Evans, P. C., Davidson, S. M., Wojta, J., Back, M., Bollini, S., Brittan, M., Catapano, A. L., Chaudhry, B., Cluitmans, M., Gnecchi, M., Guzik, T. J., Hoefer, I., Madonna, R., Monteiro, J. P., Morawietz, H., Osto, E., Padro, T., Sluimer, J. C., Tocchetti, C. G., Van der Heiden, K., Vilahur, G., Waltenberger, J., Weber, C. 2022

  Abstract

  Here we review the highlights of cardiovascular basic science in published in 2021 and early 2022 on behalf of the European Society of Cardiology Council for Basic Cardiovascular Science. We begin with non-coding RNAs which have emerged as central regulators cardiovascular biology, and then discuss how technological developments in single-cell 'omics are providing new insights in cardiovascular development, inflammation and disease. We also review recent discoveries on the biology of extracellular vesicles in driving either protective or pathogenic responses. The Nobel Prize in Physiology or Medicine 2021 recognised the importance of the molecular basis of mechanosensing and here we review breakthroughs in cardiovascular sensing of mechanical force. We also summarise discoveries in the field of atherosclerosis including the role of clonal haematopoiesis of indeterminate potential, and new mechanisms of cross-talk between hyperglycemia, lipid mediators and inflammation. The past 12 months also witnessed major advances in the field of cardiac arrhythmia including new mechanisms of fibrillation. We also focus on inducible pluripotent stem cell (iPSC) technology which has demonstrated disease causality for several genetic polymorphisms in long QT syndrome and aortic valve disease, paving the way for personalized medicine approaches. Finally, the cardiovascular community has continued to better understand COVID-19 with significant advancement in our knowledge of cardiovascular tropism, molecular markers, the mechanism of vaccine-induced thrombotic complications and new anti-viral therapies that protect the cardiovascular system.

  View details for DOI 10.1093/cvr/cvac114

  View details for PubMedID 35899362

• Single-cell RNA-seq profiling of mouse endothelial cells in response to pulmonary arterial hypertension. Cardiovascular research Rodor, J., Chen, S., Scanlon, J. P., Monteiro, J. P., Caudrillier, A., Sweta, S., Stewart, K. R., Shmakova, A., Dobie, R., Henderson, B. E., Stewart, K., Hadoke, P. W., Southwood, M., Moore, S. D., Upton, P. D., Morrell, N. W., Li, Z., Chan, S. Y., Handen, A., Lafyatis, R., de Rooij, L. P., Henderson, N. C., Carmeliet, P., Spiroski, A., Brittan, M., Baker, A. H. 2021

  Abstract

  AIMS: Endothelial cell dysfunction drives the initiation and pathogenesis of pulmonary arterial hypertension (PAH). We aimed to characterise endothelial cell (EC) dynamics in PAH at single-cell resolution.METHODS AND RESULTS: We carried out single-cell RNA sequencing (scRNA-seq) of lung ECs isolated from an EC lineage-tracing mouse model in Control and SU5416/Hypoxia-induced PAH conditions. EC populations corresponding to distinct lung vessel types, including two discrete capillary populations, were identified in both Control and PAH mice. Differential gene expression analysis revealed global PAH-induced EC changes that were confirmed by bulk RNA-seq. This included upregulation of the major histocompatibility complex class II pathway, supporting a role for ECs in the inflammatory response in PAH. We also identified a PAH response specific to the second capillary EC population including upregulation of genes involved in cell death, cell motility and angiogenesis. Interestingly, four genes with genetic variants associated with PAH were dysregulated in mouse ECs in PAH. To compare relevance across PAH models and species, we performed a detailed analysis of EC heterogeneity and response to PAH in rats and humans through whole-lung PAH scRNA-seq datasets, revealing that 51% of up-regulated mouse genes were also up-regulated in rat or human PAH. We identified promising new candidates to target endothelial dysfunction including CD74, the knockdown of which regulates EC proliferation and barrier integrity in vitro. Finally, with an in silico cell ordering approach, we identified zonation-dependent changes across the arteriovenous axis in mouse PAH and showed upregulation of the Serine/threonine-protein kinase Sgk1 at the junction between the macro- and micro-vasculature.CONCLUSIONS: This study uncovers PAH-induced EC transcriptomic changes at a high resolution, revealing novel targets for potential therapeutic candidate development.

  View details for DOI 10.1093/cvr/cvab296

  View details for PubMedID 34528097

• MIR503HG Loss Promotes Endothelial-to-Mesenchymal Transition in Vascular Disease. Circulation research Monteiro, J. P., Rodor, J., Caudrillier, A., Scanlon, J. P., Spiroski, A., Dudnakova, T., Pfluger-Muller, B., Shmakova, A., von Kriegsheim, A., Deng, L., Taylor, R. S., Wilson-Kanamori, J. R., Chen, S., Stewart, K., Thomson, A., Mitic, T., McClure, J. D., Iyinikkel, J., Hadoke, P. W., Denby, L., Bradshaw, A. C., Caruso, P., Morrell, N. W., Kovacic, J. C., Ulitsky, I., Henderson, N. C., Caporali, A., Leisegang, M. S., Brandes, R. P., Baker, A. H. 2021

  Abstract

  Rationale: Endothelial-to-mesenchymal transition (EndMT) is a dynamic biological process involved in pathological vascular remodelling. However, the molecular mechanisms that govern this transition remain largely unknown, including the contribution of long non-coding RNAs (lncRNAs). Objective: To investigate the role of lncRNAs in EndMT and their relevance to vascular remodelling. Methods and Results: To study EndMT in vitro, primary endothelial cells (EC) were treated with transforming growth factor-beta2 and interleukin-1beta. Single-cell and bulk RNA-sequencing were performed to investigate the transcriptional architecture of EndMT and identify regulated lncRNAs. The functional contribution of seven lncRNAs during EndMT was investigated based on a DsiRNA screening assay. The loss of lncRNA MIR503HG was identified as a common signature across multiple human EC types undergoing EndMT in vitro. MIR503HG depletion induced a spontaneous EndMT phenotype, while its overexpression repressed hallmark EndMT changes, regulating 29% of its transcriptome signature. Importantly, the phenotypic changes induced by MIR503HG were independent of miR-424 and miR-503, which overlap the lncRNA locus. The pathological relevance of MIR503HG down-regulation was confirmed in vivo using Sugen/Hypoxia (SuHx)-induced pulmonary hypertension (PH) in mouse, as well as in human clinical samples, in lung sections and blood outgrowth endothelial cells (BOECs) from pulmonary arterial hypertension (PAH) patients. Overexpression of human MIR503HG in SuHx mice led to reduced mesenchymal marker expression, suggesting MIR503HG therapeutic potential. We also revealed that MIR503HG interacts with the Polypyrimidine Tract Binding Protein 1 (PTB1) and regulates its protein level. PTBP1 regulation of EndMT markers suggests that the role of MIR503HG in EndMT might be mediated in part by PTBP1. Conclusions: This study reports a novel lncRNA transcriptional profile associated with EndMT and reveals the crucial role of the loss of MIR503HG in EndMT and its relevance to pulmonary hypertension.

  View details for DOI 10.1161/CIRCRESAHA.120.318124

  View details for PubMedID 33703914

• Endothelial function and dysfunction in the cardiovascular system: the long non-coding road CARDIOVASCULAR RESEARCH Monteiro, J. P., Bennett, M., Rodor, J., Caudrillier, A., Ulitsky, I., Baker, A. H. 2019; 115 (12): 1692-1704

  Abstract

  Present throughout the vasculature, endothelial cells (ECs) are essential for blood vessel function and play a central role in the pathogenesis of diverse cardiovascular diseases. Understanding the intricate molecular determinants governing endothelial function and dysfunction is essential to develop novel clinical breakthroughs and improve knowledge. An increasing body of evidence demonstrates that long non-coding RNAs (lncRNAs) are active regulators of the endothelial transcriptome and function, providing emerging insights into core questions surrounding EC contributions to pathology, and perhaps the emergence of novel therapeutic opportunities. In this review, we discuss this class of non-coding transcripts and their role in endothelial biology during cardiovascular development, homeostasis, and disease, highlighting challenges during discovery and characterization and how these have been overcome to date. We further discuss the translational therapeutic implications and the challenges within the field, highlighting lncRNA that support endothelial phenotypes prevalent in cardiovascular disease.

  View details for DOI 10.1093/cvr/cvz154

  View details for Web of Science ID 000491246600010

  View details for PubMedID 31214683

  View details for PubMedCentralID PMC6755355

• Loss of the Long Non-Coding RNA MIR503HG Promotes Endothelial-to-Mesenchymal Transition Caudrillier, A., Monteiro, J., Rodor, J., Shmakova, A., Baker, A. H. KARGER. 2019: 100-101

  View details for Web of Science ID 000463529300219