Pericytes in the disease spotlight

Molecular and functional pericyte studies at single-cell resolution are providing new insights into long-standing questions about pericyte heterogeneity.Pericytes are not identified by a single marker but instead by gene expression signatures that show substantial inter-organ differences.Pericytes orchestrate and precede endothelial cell responses during angiogenesis.Pericyte degeneration and dysfunction, that are triggered by the onset of some diseases, contribute to the progression of those diseases in both vascular and non-vascular contexts.The number of diseases with pericyte dysfunction continues to expand, thereby anticipating a promising future for pericyte-focused therapy. Pericytes are classically defined as mural cells (see Glossary) that envelop the endothelium of small caliber blood vessels, the so-called capillaries. Pericytes are embedded within the same basement membrane as endothelial cells (ECs) and interact closely with them [1.Armulik A. et al.Pericytes: developmental, physiological, and pathological perspectives, problems, and promises.Dev. Cell. 2011; 21: 193-215Abstract Full Text Full Text PDF PubMed Scopus (1790) Google Scholar,2.Holm A. et al.Microvascular mural cell organotypic heterogeneity and functional plasticity.Trends Cell Biol. 2018; 28: 302-316Abstract Full Text Full Text PDF PubMed Scopus (63) Google Scholar]. By contrast, vascular smooth muscle cells (vSMCs), the other mural cell type, cover large arteries and veins, and are physically separated from the endothelium by an intimal layer of extracellular matrix (ECM). Of note, lymphatic capillaries lack pericytes under physiological conditions, although collecting lymphatic vessels contain vSMCs [3.Petrova T.V. Koh G.Y. Biological functions of lymphatic vessels.Science. 2020; 369eaax4063Crossref PubMed Scopus (144) Google Scholar]. A fundamental function of mural cells is to regulate the stabilization and function of blood vessels. It is therefore not surprising that pericyte loss and dysfunction were linked to several diseases including cancer and cerebrovascular diseases more than a decade ago [4.Martin J.D. et al.Normalizing function of tumor vessels: progress, opportunities, and challenges.Annu. Rev. Physiol. 2019; 81: 505-534Crossref PubMed Scopus (242) Google Scholar,5.Lendahl U. et al.Emerging links between cerebrovascular and neurodegenerative diseases-a special role for pericytes.EMBO Rep. 2019; 20e48070Crossref PubMed Scopus (71) Google Scholar]. However, pericyte-focused therapies have been poorly explored. Instead, most studies on vascular-directed therapeutic strategies have been on ECs – the central components that build blood vessels. Emerging data are, however, changing the perception of pericytes from mere supporting vascular cells that are recruited at the final stage of vessel formation to essential elements in the early phases of angiogenesis that anticipate and orchestrate EC behavior. In addition, recent research is revealing novel pathological roles for pericytes beyond their implications in the vasculature. Collectively, we believe that these data open exciting avenues for pericyte-focused therapeutic approaches and call for a broader understanding of these cells in disease progression. We provide here a global overview of recent significant advances regarding our understanding of the role of pericytes in different pathobiological scenarios and discuss the field's current paradigms and controversies. First, we address new insights into the functions associated with pericytes during physiological vascular responses. Second, we discuss evidence supporting a role of pericytes in disease, including pericyte cell-autonomous implications beyond the vasculature. For comprehensive details on pericyte biology, function ontology, and specific signaling pathways, we refer the reader to [1.Armulik A. et al.Pericytes: developmental, physiological, and pathological perspectives, problems, and promises.Dev. Cell. 2011; 21: 193-215Abstract Full Text Full Text PDF PubMed Scopus (1790) Google Scholar,2.Holm A. et al.Microvascular mural cell organotypic heterogeneity and functional plasticity.Trends Cell Biol. 2018; 28: 302-316Abstract Full Text Full Text PDF PubMed Scopus (63) Google Scholar,5.Lendahl U. et al.Emerging links between cerebrovascular and neurodegenerative diseases-a special role for pericytes.EMBO Rep. 2019; 20e48070Crossref PubMed Scopus (71) Google Scholar]. Of importance, some of the emerging concepts in pericyte biology described in the following sections have only been studied in one specific tissue. To avoid confusion about the generalizability of pericyte properties, we frame each function by considering the relevant organ of study. Pericytes exhibit significant inter- and intra-tissue molecular differences and exert tissue-specific functions [2.Holm A. et al.Microvascular mural cell organotypic heterogeneity and functional plasticity.Trends Cell Biol. 2018; 28: 302-316Abstract Full Text Full Text PDF PubMed Scopus (63) Google Scholar]. Their molecular, morphological, and functional heterogeneity is inextricably linked to their diverse developmental origins, modes of vessel recruitment, and specific anatomical localization. For example, pericytes of the central nervous system (CNS) microvasculature are firmly and continuously invested around the endothelium to support vascular barrier properties, whereas liver pericytes, commonly referred to as hepatic stellate cells (HSCs), reside in the perisinusoidal space, are loosely and discontinuously associated to ECs, and hold a unique vitamin A storage capacity [2.Holm A. et al.Microvascular mural cell organotypic heterogeneity and functional plasticity.Trends Cell Biol. 2018; 28: 302-316Abstract Full Text Full Text PDF PubMed Scopus (63) Google Scholar]. To meet tissue-specific demands, pericyte distribution and density are variable among organs and vascular beds, with the CNS microvasculature showing the greatest pericyte-to-EC abundance. From a molecular standpoint there is no single molecular marker that can exclusively identify pericytes (Box 1), albeit the emergence of single-cell techniques is shedding light on tissue-specific pericyte molecular markers and functions. For example, the first molecular atlas of vascular cell types in the brain of adult mice by single-cell RNA sequencing (scRNA-seq) revealed that mural cells follow a gradient of transitional phenotypes. This gradient occurs at the interface of precapillary arterioles, capillaries, and postcapillary venules, and does not follow a single continuum along the arteriovenous axis (Figure 1 and Box 1) [6.Vanlandewijck M. et al.A molecular atlas of cell types and zonation in the brain vasculature.Nature. 2018; 554: 475-480Crossref PubMed Scopus (876) Google Scholar]. Whether this gradient of transitional phenotypes is specifically restricted to the brain vasculature or is also present in other vascular beds remains to be determined. Indeed, pericytes exhibit many organotypic differences in the expression of molecular markers (Figure 2 illustrates three top-ranked pericyte markers with enriched expression per organ), of which the expression of transporters and components of the contractile machinery exhibit the greatest differences between organs [7.Muhl L. et al.Single-cell analysis uncovers fibroblast heterogeneity and criteria for fibroblast and mural cell identification and discrimination.Nat. Commun. 2020; 11: 3953Crossref PubMed Scopus (187) Google Scholar]. Another intriguing observation is that pericytes exhibit more cross-organ heterogeneity than vSMCs [7.Muhl L. et al.Single-cell analysis uncovers fibroblast heterogeneity and criteria for fibroblast and mural cell identification and discrimination.Nat. Commun. 2020; 11: 3953Crossref PubMed Scopus (187) Google Scholar,8.Muhl L. et al.A single-cell transcriptomic inventory of murine smooth muscle cells.Dev. Cell. 2022; 57: 2426-2443Abstract Full Text Full Text PDF PubMed Google Scholar]. Currently, the inter-tissue differences in the behavior of the two main mural cell types are not completely understood. However, this may be because pericytes exhibit a greater cell-intrinsic plasticity to adapt their molecular portfolio and function to tissue-specific demands, whereas vSMCs fulfill a more universal function across tissues. In contrast to the tissue-specific transcriptomic differences, the expression of transcription factors appears to be relatively conserved in mural cells across organs, thereby suggesting that mural cell subtypes are defined by epigenetic mechanisms [7.Muhl L. et al.Single-cell analysis uncovers fibroblast heterogeneity and criteria for fibroblast and mural cell identification and discrimination.Nat. Commun. 2020; 11: 3953Crossref PubMed Scopus (187) Google Scholar]. Accordingly, DNA hypermethylation was recently found to control alpha smooth muscle actin (αSMA) expression in renal mural cells after ischemia [9.Chou Y.H. et al.Methylation in pericytes after acute injury promotes chronic kidney disease.J. Clin. Invest. 2020; 130: 4845-4857Crossref PubMed Scopus (18) Google Scholar]. This indicates that methods such as assay for transposase-accessible chromatin sequencing (ATAC-seq) will be instrumental to further understand mural cell phenotypes.Box 1Unraveling the identity of pericytesThe identification of pericytes remains a challenging task. Despite ongoing efforts, there is no consensus regarding unambiguous criteria for pericyte identification. To date no single molecular marker can exclusively identify all pericytes or distinguish pericytes from other cell types, although scRNA-seq is now providing new opportunities to discern pericyte marker heterogeneity and tissue specificity [6.Vanlandewijck M. et al.A molecular atlas of cell types and zonation in the brain vasculature.Nature. 2018; 554: 475-480Crossref PubMed Scopus (876) Google Scholar,8.Muhl L. et al.A single-cell transcriptomic inventory of murine smooth muscle cells.Dev. Cell. 2022; 57: 2426-2443Abstract Full Text Full Text PDF PubMed Google Scholar,71.Teuwen L.A. et al.Tumor vessel co-option probed by single-cell analysis.Cell Rep. 2021; 35109253Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar,93.Baek S.H. et al.Single cell transcriptomic analysis reveals organ specific pericyte markers and identities.Front Cardiovasc. Med. 2022; 9876591Crossref Scopus (9) Google Scholar]. The use of transgenic reporter mouse models has been instrumental to label, trace, and locate different mural cell populations in vivo. A combination of multiple reporter lines is often necessary to properly identify and discriminate pericytes from endothelial cells (ECs) and other perivascular cells [6.Vanlandewijck M. et al.A molecular atlas of cell types and zonation in the brain vasculature.Nature. 2018; 554: 475-480Crossref PubMed Scopus (876) Google Scholar, 7.Muhl L. et al.Single-cell analysis uncovers fibroblast heterogeneity and criteria for fibroblast and mural cell identification and discrimination.Nat. Commun. 2020; 11: 3953Crossref PubMed Scopus (187) Google Scholar, 8.Muhl L. et al.A single-cell transcriptomic inventory of murine smooth muscle cells.Dev. Cell. 2022; 57: 2426-2443Abstract Full Text Full Text PDF PubMed Google Scholar]. Mural cells are highly plastic cells; phenotypic zonation of mouse brain mural cells has revealed that these cells do not follow a single continuum along the arteriovenous axis (see Figure 1A,B in main text) [6.Vanlandewijck M. et al.A molecular atlas of cell types and zonation in the brain vasculature.Nature. 2018; 554: 475-480Crossref PubMed Scopus (876) Google Scholar]. From a transcriptional point of view, there are two distinct continuums of mural cells: (i) capillary pericytes and venous smooth muscle cells (SMCs), where pericytes gradually transition to a venous SMC phenotype, and (ii) arterial SMCs which transition in an distinct pattern towards arteriole SMCs. The transcriptional resemblance between mouse brain pericytes and venular mural cells [6.Vanlandewijck M. et al.A molecular atlas of cell types and zonation in the brain vasculature.Nature. 2018; 554: 475-480Crossref PubMed Scopus (876) Google Scholar], as well as the lack of classic pericytes in several organs [7.Muhl L. et al.Single-cell analysis uncovers fibroblast heterogeneity and criteria for fibroblast and mural cell identification and discrimination.Nat. Commun. 2020; 11: 3953Crossref PubMed Scopus (187) Google Scholar,8.Muhl L. et al.A single-cell transcriptomic inventory of murine smooth muscle cells.Dev. Cell. 2022; 57: 2426-2443Abstract Full Text Full Text PDF PubMed Google Scholar], have led to the hypothesis that capillary pericytes are transcriptionally and morphologically similar to venous SMCs in some tissues. Human brain mural cells recapitulate the mouse zonation pattern, although human pericytes are evenly distributed over capillaries and veins [50.Yang A.C. et al.A human brain vascular atlas reveals diverse mediators of Alzheimer’s risk.Nature. 2022; 603: 885-892Crossref PubMed Scopus (117) Google Scholar,94.Garcia F.J. et al.Single-cell dissection of the human brain vasculature.Nature. 2022; 603: 893-899Crossref PubMed Scopus (53) Google Scholar]. Unlike the anatomical separation of pericytes and venous SMCs in the mouse brain, subtypes of human pericytes are discerned by functionality marked by solute transport and extracellular matrix (ECM) organization [50.Yang A.C. et al.A human brain vascular atlas reveals diverse mediators of Alzheimer’s risk.Nature. 2022; 603: 885-892Crossref PubMed Scopus (117) Google Scholar]. Unfortunately, the ability of mouse markers to predict the presence of human pericytes remains limited, and only a select few retain adequate specificity. The use of zebrafish models may provide a better alternative to study conserved pericyte genes [95.Shih Y.H. et al.Integrated molecular analysis identifies a conserved pericyte gene signature in zebrafish.Development. 2021; 148dev200189Crossref PubMed Scopus (4) Google Scholar]. We believe that RGS5, NDUFA4L2, KNCJ8, HIGD1B, ABCC9, NOTCH3, and PDGFRB are currently the most organ and species conserved pericyte markers, although detailed intra-tissue characterization remains necessary when studying pericytes (see Figure 2 in main text).Figure 2Organotypic heterogeneity of pericyte markers.Show full captionThis figure summarizes top-ranked pericyte markers in the brain, heart, lung, kidney, and colon of mouse (upper row) and human (lower row). Pericyte markers were chosen based on a stringent evaluation of transcriptional abundance, specificity, and homogeneity utilizing information provided by single-cell RNA sequencing (scRNA-seq) data [6.Vanlandewijck M. et al.A molecular atlas of cell types and zonation in the brain vasculature.Nature. 2018; 554: 475-480Crossref PubMed Scopus (876) Google Scholar, 7.Muhl L. et al.Single-cell analysis uncovers fibroblast heterogeneity and criteria for fibroblast and mural cell identification and discrimination.Nat. Commun. 2020; 11: 3953Crossref PubMed Scopus (187) Google Scholar, 8.Muhl L. et al.A single-cell transcriptomic inventory of murine smooth muscle cells.Dev. Cell. 2022; 57: 2426-2443Abstract Full Text Full Text PDF PubMed Google Scholar,50.Yang A.C. et al.A human brain vascular atlas reveals diverse mediators of Alzheimer’s risk.Nature. 2022; 603: 885-892Crossref PubMed Scopus (117) Google Scholar,82.Muhl L. et al.The SARS-CoV-2 receptor ACE2 is expressed in mouse pericytes but not endothelial cells: implications for COVID-19 vascular research.Stem Cell Rep. 2022; 17: 1089-1104Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar,84.Dobie R. et al.Single-cell transcriptomics uncovers zonation of function in the mesenchyme during liver fibrosis.Cell Rep. 2019; 29: 1832-1847Abstract Full Text Full Text PDF PubMed Scopus (164) Google Scholar,85.Kuppe C. et al.Decoding myofibroblast origins in human kidney fibrosis.Nature. 2021; 589: 281-286Crossref PubMed Scopus (225) Google Scholar,95.Shih Y.H. et al.Integrated molecular analysis identifies a conserved pericyte gene signature in zebrafish.Development. 2021; 148dev200189Crossref PubMed Scopus (4) Google Scholar,100.Winkler E.A. et al.A single-cell atlas of the normal and malformed human brain vasculature.Science. 2022; 375: eabi7377Crossref PubMed Scopus (5) Google Scholar, 101.Travaglini K.J. et al.A molecular cell atlas of the human lung from single-cell RNA sequencing.Nature. 2020; 587: 619-625Crossref PubMed Scopus (470) Google Scholar, 102.Kinchen J. et al.Structural remodeling of the human colonic mesenchyme in inflammatory bowel disease.Cell. 2018; 175: 372-386Abstract Full Text Full Text PDF PubMed Scopus (313) Google Scholar]. Validation of the selected markers by in situ analysis was used as a second criteria for their selection.View Large Image Figure ViewerDownload Hi-res image Download (PPT) The identification of pericytes remains a challenging task. Despite ongoing efforts, there is no consensus regarding unambiguous criteria for pericyte identification. To date no single molecular marker can exclusively identify all pericytes or distinguish pericytes from other cell types, although scRNA-seq is now providing new opportunities to discern pericyte marker heterogeneity and tissue specificity [6.Vanlandewijck M. et al.A molecular atlas of cell types and zonation in the brain vasculature.Nature. 2018; 554: 475-480Crossref PubMed Scopus (876) Google Scholar,8.Muhl L. et al.A single-cell transcriptomic inventory of murine smooth muscle cells.Dev. Cell. 2022; 57: 2426-2443Abstract Full Text Full Text PDF PubMed Google Scholar,71.Teuwen L.A. et al.Tumor vessel co-option probed by single-cell analysis.Cell Rep. 2021; 35109253Abstract Full Text Full Text PDF PubMed Scopus (35) Google Scholar,93.Baek S.H. et al.Single cell transcriptomic analysis reveals organ specific pericyte markers and identities.Front Cardiovasc. Med. 2022; 9876591Crossref Scopus (9) Google Scholar]. The use of transgenic reporter mouse models has been instrumental to label, trace, and locate different mural cell populations in vivo. A combination of multiple reporter lines is often necessary to properly identify and discriminate pericytes from endothelial cells (ECs) and other perivascular cells [6.Vanlandewijck M. et al.A molecular atlas of cell types and zonation in the brain vasculature.Nature. 2018; 554: 475-480Crossref PubMed Scopus (876) Google Scholar, 7.Muhl L. et al.Single-cell analysis uncovers fibroblast heterogeneity and criteria for fibroblast and mural cell identification and discrimination.Nat. Commun. 2020; 11: 3953Crossref PubMed Scopus (187) Google Scholar, 8.Muhl L. et al.A single-cell transcriptomic inventory of murine smooth muscle cells.Dev. Cell. 2022; 57: 2426-2443Abstract Full Text Full Text PDF PubMed Google Scholar]. Mural cells are highly plastic cells; phenotypic zonation of mouse brain mural cells has revealed that these cells do not follow a single continuum along the arteriovenous axis (see Figure 1A,B in main text) [6.Vanlandewijck M. et al.A molecular atlas of cell types and zonation in the brain vasculature.Nature. 2018; 554: 475-480Crossref PubMed Scopus (876) Google Scholar]. From a transcriptional point of view, there are two distinct continuums of mural cells: (i) capillary pericytes and venous smooth muscle cells (SMCs), where pericytes gradually transition to a venous SMC phenotype, and (ii) arterial SMCs which transition in an distinct pattern towards arteriole SMCs. The transcriptional resemblance between mouse brain pericytes and venular mural cells [6.Vanlandewijck M. et al.A molecular atlas of cell types and zonation in the brain vasculature.Nature. 2018; 554: 475-480Crossref PubMed Scopus (876) Google Scholar], as well as the lack of classic pericytes in several organs [7.Muhl L. et al.Single-cell analysis uncovers fibroblast heterogeneity and criteria for fibroblast and mural cell identification and discrimination.Nat. Commun. 2020; 11: 3953Crossref PubMed Scopus (187) Google Scholar,8.Muhl L. et al.A single-cell transcriptomic inventory of murine smooth muscle cells.Dev. Cell. 2022; 57: 2426-2443Abstract Full Text Full Text PDF PubMed Google Scholar], have led to the hypothesis that capillary pericytes are transcriptionally and morphologically similar to venous SMCs in some tissues. Human brain mural cells recapitulate the mouse zonation pattern, although human pericytes are evenly distributed over capillaries and veins [50.Yang A.C. et al.A human brain vascular atlas reveals diverse mediators of Alzheimer’s risk.Nature. 2022; 603: 885-892Crossref PubMed Scopus (117) Google Scholar,94.Garcia F.J. et al.Single-cell dissection of the human brain vasculature.Nature. 2022; 603: 893-899Crossref PubMed Scopus (53) Google Scholar]. Unlike the anatomical separation of pericytes and venous SMCs in the mouse brain, subtypes of human pericytes are discerned by functionality marked by solute transport and extracellular matrix (ECM) organization [50.Yang A.C. et al.A human brain vascular atlas reveals diverse mediators of Alzheimer’s risk.Nature. 2022; 603: 885-892Crossref PubMed Scopus (117) Google Scholar]. Unfortunately, the ability of mouse markers to predict the presence of human pericytes remains limited, and only a select few retain adequate specificity. The use of zebrafish models may provide a better alternative to study conserved pericyte genes [95.Shih Y.H. et al.Integrated molecular analysis identifies a conserved pericyte gene signature in zebrafish.Development. 2021; 148dev200189Crossref PubMed Scopus (4) Google Scholar]. We believe that RGS5, NDUFA4L2, KNCJ8, HIGD1B, ABCC9, NOTCH3, and PDGFRB are currently the most organ and species conserved pericyte markers, although detailed intra-tissue characterization remains necessary when studying pericytes (see Figure 2 in main text). This figure summarizes top-ranked pericyte markers in the brain, heart, lung, kidney, and colon of mouse (upper row) and human (lower row). Pericyte markers were chosen based on a stringent evaluation of transcriptional abundance, specificity, and homogeneity utilizing information provided by single-cell RNA sequencing (scRNA-seq) data [6.Vanlandewijck M. et al.A molecular atlas of cell types and zonation in the brain vasculature.Nature. 2018; 554: 475-480Crossref PubMed Scopus (876) Google Scholar, 7.Muhl L. et al.Single-cell analysis uncovers fibroblast heterogeneity and criteria for fibroblast and mural cell identification and discrimination.Nat. Commun. 2020; 11: 3953Crossref PubMed Scopus (187) Google Scholar, 8.Muhl L. et al.A single-cell transcriptomic inventory of murine smooth muscle cells.Dev. Cell. 2022; 57: 2426-2443Abstract Full Text Full Text PDF PubMed Google Scholar,50.Yang A.C. et al.A human brain vascular atlas reveals diverse mediators of Alzheimer’s risk.Nature. 2022; 603: 885-892Crossref PubMed Scopus (117) Google Scholar,82.Muhl L. et al.The SARS-CoV-2 receptor ACE2 is expressed in mouse pericytes but not endothelial cells: implications for COVID-19 vascular research.Stem Cell Rep. 2022; 17: 1089-1104Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar,84.Dobie R. et al.Single-cell transcriptomics uncovers zonation of function in the mesenchyme during liver fibrosis.Cell Rep. 2019; 29: 1832-1847Abstract Full Text Full Text PDF PubMed Scopus (164) Google Scholar,85.Kuppe C. et al.Decoding myofibroblast origins in human kidney fibrosis.Nature. 2021; 589: 281-286Crossref PubMed Scopus (225) Google Scholar,95.Shih Y.H. et al.Integrated molecular analysis identifies a conserved pericyte gene signature in zebrafish.Development. 2021; 148dev200189Crossref PubMed Scopus (4) Google Scholar,100.Winkler E.A. et al.A single-cell atlas of the normal and malformed human brain vasculature.Science. 2022; 375: eabi7377Crossref PubMed Scopus (5) Google Scholar, 101.Travaglini K.J. et al.A molecular cell atlas of the human lung from single-cell RNA sequencing.Nature. 2020; 587: 619-625Crossref PubMed Scopus (470) Google Scholar, 102.Kinchen J. et al.Structural remodeling of the human colonic mesenchyme in inflammatory bowel disease.Cell. 2018; 175: 372-386Abstract Full Text Full Text PDF PubMed Scopus (313) Google Scholar]. Validation of the selected markers by in situ analysis was used as a second criteria for their selection. Many studies have documented that pericytes contribute to angiogenesis [10.Potente M. et al.Basic and therapeutic aspects of angiogenesis.Cell. 2011; 146: 873-887Abstract Full Text Full Text PDF PubMed Scopus (1978) Google Scholar]. The historical view proposes that pericytes mainly contribute to the late stages of vessel formation [2.Holm A. et al.Microvascular mural cell organotypic heterogeneity and functional plasticity.Trends Cell Biol. 2018; 28: 302-316Abstract Full Text Full Text PDF PubMed Scopus (63) Google Scholar,10.Potente M. et al.Basic and therapeutic aspects of angiogenesis.Cell. 2011; 146: 873-887Abstract Full Text Full Text PDF PubMed Scopus (1978) Google Scholar]. By taking advantage of the mouse retina as a paradigmatic experimental model of developmental angiogenesis, this concept has been challenged [11.Park D.Y. et al.Plastic roles of pericytes in the blood-retinal barrier.Nat. Commun. 2017; 8: 15296Crossref PubMed Scopus (175) Google Scholar, 12.Figueiredo A.M. et al.Phosphoinositide 3-kinase-regulated pericyte maturation governs vascular remodeling.Circulation. 2020; 142: 688-704Crossref PubMed Scopus (25) Google Scholar, 13.Orlich M.M. et al.Mural cell SRF controls pericyte migration, vessel patterning and blood flow.Circ. Res. 2022; 131: 308-327Crossref PubMed Scopus (0) Google Scholar, 14.Dieguez-Hurtado R. et al.Loss of the transcription factor RBPJ induces disease-promoting properties in brain pericytes.Nat. Commun. 2019; 10: 2817Crossref PubMed Scopus (0) Google Scholar, 15.Teichert M. et al.Pericyte-expressed Tie2 controls angiogenesis and vessel maturation.Nat. Commun. 2017; 8: 16106Crossref PubMed Scopus (174) Google Scholar, 16.Eilken H.M. et al.Pericytes regulate VEGF-induced endothelial sprouting through VEGFR1.Nat. Commun. 2017; 8: 1574Crossref PubMed Scopus (134) Google Scholar]. Indeed, these studies showed that, during the early phases of developmental angiogenesis, pericytes, which have not yet achieved the maturity seen in formed vessels, are permissive to cell-cycle progression, morphological adaptation, and migration [12.Figueiredo A.M. et al.Phosphoinositide 3-kinase-regulated pericyte maturation governs vascular remodeling.Circulation. 2020; 142: 688-704Crossref PubMed Scopus (25) Google Scholar,13.Orlich M.M. et al.Mural cell SRF controls pericyte migration, vessel patterning and blood flow.Circ. Res. 2022; 131: 308-327Crossref PubMed Scopus (0) Google Scholar]. In this setting, pericyte growth precedes the expansion of ECs, although it is still unclear why. One possibility is that, by expanding rapidly, pericytes ensure the production of sufficient EC growth signals, a hypothesis which is coherent with the observation that inhibition of pericyte activation blocks EC proliferation [12.Figueiredo A.M. et al.Phosphoinositide 3-kinase-regulated pericyte maturation governs vascular remodeling.Circulation. 2020; 142: 688-704Crossref PubMed Scopus (25) Google Scholar] and induces nuclear translocation of FOXO1 [11.Park D.Y. et al.Plastic roles of pericytes in the blood-retinal barrier.Nat. Commun. 2017; 8: 15296Crossref PubMed Scopus (175) Google Scholar], the master regulator of EC quiescence [17.Kobialka P. Graupera M. Revisiting PI3-kinase signalling in angiogenesis.Vasc. Biol. 2019; 1: H125-H134Crossref PubMed Google Scholar]. Another study that examined the brain vasculature showed that, when pericytes are absent, ECs become angiogenic but are not able to proliferate [18.Mae M.A. et al.Single-cell analysis of blood–brain barrier response to pericyte loss.Circ. Res. 2021; 128: e46-e62Crossref PubMed Scopus (0) Google Scholar], thereby supporting a model in which ECs require the presence of pericytes to expand. Nonetheless, it is fair to acknowledge that other studies have shown that reduced pericyte coverage leads to increased EC proliferation [19.Dave J.M. et al.Pericyte ALK5/TIMP3 axis contributes to endothelial morphogenesis in the developing brain.Dev. Cell. 2018; 47: 388-389Abstract Full Text Full Text PDF PubMed Scopus (8) Google Scholar]. Although these discrepancies highlight that pericyte–EC interactions are complex, they may be explained by the differences between the animal models and genetic strategies used to interfere with pericytes. Importantly, pericyte behaviors during angiogenesis have been mostly described in tissues belonging to the CNS. Hence, given the high abundance of pericytes in the CNS, it is possible that angiogenic pericytes fulfill different roles in tissues where ECs substantially outnumber them. Another interesting observation is that, during angiogenesis, immature pericytes remain in close contact with ECs, although they do not cover them in their entirety [12.Figueiredo A.M. et al.Phosphoinositide 3-kinase-regulated pericyte maturation governs vascular remodeling.Circulation. 2020; 142: 688-704Crossref PubMed Scopus (25) Google Scholar,20.Crouch E.E. et al.Ensembles of endothelial and mural cells promote angiogenesis in prenatal human brain.Cell. 2022; 185: 3753-3769Abstract Full Text Full Text PDF PubMed Scopus (11) Google Scholar]. This suggests that pericyte–EC communication during angiogenesis relies on both paracrine and juxtracrine signaling, and may explain why pericyte loss [11.Park D.Y. et al.Plastic roles of pericytes in the blood-retinal barrier.Nat. Commun. 2017; 8: 15296Crossref Pu