The next generation of endothelial differentiation: Tissue-specific ECs

Endothelial cells (ECs) sense and respond to fluid flow regulate immune cell trafficking in all organs. Despite sharing the same mesodermal origin, ECs exhibit heterogeneous tissue-specific characteristics. Human pluripotent stem (hPSCs) can potentially be harnessed capture this heterogeneity further elucidate endothelium behavior satisfy need for increased accuracy breadth of disease models therapeutics. Here, we review current strategies hPSC differentiation blood vascular their maturation into continuous, fenestrated, sinusoidal tissues. We then discuss contribution hPSC-derived recent advances organoid development organ-on-chip approaches. are indispensable. In addition lining lumen vessels transport oxygen nutrients throughout body, play an important role maintaining homeostasis The regenerative ability multilineage potential human facilitate use answer developmental questions, model pathogenesis, develop therapeutics (Williams Wu, 2019Williams I.M. Wu J.C. Generation endothelial from cells.Arterioscler. Thromb. Vasc. Biol. 2019; 39: 1317-1329Crossref PubMed Scopus (16) Google Scholar). Deriving hPSCs provides opportunities decipher regeneration, particularly light importance tissue function. generated (hPSC-ECs) capable enhancing angiogenesis a range mouse vascularizing engineered tissues Although arterial venous different similar each other, among organs is well recognized, with characteristics typically arising capillary networks. Heterogeneous distribution EC markers platelet adhesion molecule (PECAM-1; CD31), cadherin (VE-cad; CD144), von Willebrand factor (vWf) have been reported across heart, lung, liver, kidneys (Marcu et al., 2018Marcu R. Choi Y.J. Xue J. Fortin C.L. Wang Y. Nagao R.J. Xu MacDonald J.W. Bammler T.K. Murry C.E. al.Human organ-specific heterogeneity.iScience. 2018; 4: 20-35Abstract Full Text PDF (68) These differences marker expression continue present when expanded vitro. Different genetic profiles kidney were identified using RNA sequencing Combined endothelial-specific translating ribosome affinity purification (EC-TRAP) high-throughput revealed existence subsets within tissues, allowing understanding diverse functionality (Cleuren 2019Cleuren A.C.A. van der Ent M.A. Jiang H. Hunker K.L. Yee A. Siemieniak D.R. Molema G. Aird W.C. Ganesh S.K. Ginsburg D. vivo translatome highly beds.Proc. Natl. Acad. Sci. U S 116: 23618-23624Crossref (0) Another study demonstrated that healthy or diseased brain, lung gene patterns (Jambusaria 2020Jambusaria Hong Z. Zhang L. Srivastava S. Jana Toth P.T. Dai Malik A.B. Rehman distinct beds during inflammation.eLife. 2020; 9: e51413Crossref (33) Nonetheless, hPSC-EC protocols yet characterization Single-cell on hPSC-ECs four main subtypes clusters. For example, claudin-5 (CLDN5) abundantly expressed one population high metabolism, while another more apelin receptor (APLNR), inflammation-related (Paik 2018Paik D.T. Tian Lee Sayed N. Chen I.Y. Rhee Kim Wirka R.C. Buikema al.Large-scale single-cell RNA-seq reveals molecular signatures populations induced cell-derived cells.Circ. Res. 123: 443-450Crossref (42) This again suggests limitation protocols. Above all, generate tissue- ECs, additional elements incorporated differentiating hPSCs. would ensure obtained through morphology, functionality, metabolic profile as organ under study. believe may offer precise modeling systems research therapy. To recapitulate biological events development, researchers endeavored consistent efficient Commonly used approaches include OP-9 coculture, two-dimensional (2D) differentiation, modification (see details by Williams Among these, monolayer-directed supplemented growth factors small molecules widely used. selection supplementary based signaling pathways extracellular matrices supporting (Tables 1 2). method reproducibly 2 weeks. Feeder-free medium mTeSR essential 8 (E8) media rho-associated kinase (ROCK) inhibitor commonly (Chen 2011Chen Gulbranson Hou Bolin J.M. Ruotti V. Probasco M.D. Smuga-Otto K. Howden S.E. Diol N.R. Propson N.E. al.Chemically defined conditions iPSC derivation culture.Nat. Methods. 2011; 8: 424-429Crossref (798) E8 contains components required including ascorbic acid, selenium, insulin, sodium bicarbonate, transferrin, TGF-?1, FGF2. Mesoderm induction usually carried out 6 (E6) medium, which consists formulation without TGF-?1 FGF2.Table 1Growth factors, molecules, transcription mesoderm transductionGrowth factorsMechanismsReferencesBone morphogenetic protein 4 (BMP4)Synergizes (VEGF) induce angiogenesisSriram 2015Sriram Tan J.Y. Islam I. Rufaihah A.J. Cao T. Efficient embryonic feeder- serum-free conditions.Stem Cell Ther. 2015; 6: 261Crossref (58) ScholarFibroblast (FGFs)Promote vesicular formation upregulating glycolytic enzyme hexokinase (HK2) increase glycolysisYu 2017Yu P. Wilhelm Dubrac Tung J.K. Alves T.C. Fang J.S. Xie Zhu De Smet F. al.FGF-dependent control development.Nature. 2017; 545: 224-228Crossref (121) ScholarGlycogen synthase 3 (GSK3) inhibitor: CHIR99021Represses pluripotency promote lineagePatsch 2015Patsch C. Challet-Meylan Thoma E.C. Urich E. Heckel O’Sullivan J.F. Grainger S.J. Kapp F.G. Sun Christensen al.Generation smooth muscle cells.Nat. 17: 994-1003Crossref (237) ScholarE-twenty-six specific sequence (ETS) ETS variant (ETV2)• ETV2 transduction mature leads self-assembled functional revascularization organoids• ETV2-knockout embryos experience embryo lethalityPalikuqi 2020Palikuqi B. Nguyen Li Schreiner Pellegata A.F. Liu Redmond Geng Lin Gómez-Salinero al.Adaptable haemodynamic organogenesis tumorigenesis.Nature. 585: 426-432Crossref (14) Scholar, Oh 2015Oh S.-Y. Park factor, ETV2: master regulator development.Mol. Cells. 38: 1029-1036Crossref Scholar Open table new tab Table 2Extracellular support proliferation, migration, differentiationMatrixMechanismsReferenceCollagen IPromotes morphogenesis inhibition cAMP-dependent A (PKA) actin polymerizationWhelan Senger, 2003Whelan M.C. Senger Collagen I initiates inducing polymerization suppression cyclic AMP Chem. 2003; 278: 327-334Abstract (119) ScholarCollagen IVEnhances angiogenic abilities ECsBahramsoltani 2014Bahramsoltani M. Slosarek Spiegelaere W. Plendl Angiogenesis collagen type IV culture systems.Anat. Histol. Embryol. 2014; 43: 103-115Crossref (27) ScholarFibronectinFibronectin integrin ?5?1 promotes binding VEGF (VEGFR2/KDR), mitogen-activated (MAPK) activation resulting migrationWijelath 2002Wijelath E.S. Murray Rahman Patel Ishida Strand Aziz Cardona Hammond W.P. Savidge G.F. al.Novel domains fibronectin enhance activity.Circ. 2002; 91: 25-31Crossref (203) Zou 2012Zou Kang Huebert Shah V.H. Fibronectin induces migration ?1 Src-dependent phosphorylation fibroblast receptor-1 at tyrosines 653/654 766.J. 2012; 287: 7190-7202Abstract (55) ScholarLaminin• Laminin ?5 barrier function ?3 integrins RhoA expression, stabilizes VE-cadherin• ?4 developing retina responsible tip stalk regulating Dll4/Notch pathwaySong 2017Song X. Buscher Di Russo Lütke-Enking Zarbock Stadtmann al.Endothelial basement membrane laminin 511 contributes junctional tightness thereby inhibits leukocyte transmigration.Cell Rep. 18: 1256-1269Abstract Stenzel 2011Stenzel Franco C.A. Estrach Mettouchi Sauvaget Rosewell Schertel Armer Domogatskaya Rodin limits signalling vivo.EMBO 12: 1135-1143Crossref (100) protocol, CHIR99021 BMP4 conjunction mesoderm. Subsequent VEGF-A forskolin resulted yielding approximately 60% ECs. robustly produced maintained up passage 10. Notably, entire process protocol suspension form, scaled production clinical (Olmer 2018Olmer Engels Usman Menke M.N.H. Pessler Göhring Bornhorst Bolten Abdelilah-Seyfried al.Differentiation scalable culture.Stem Reports. 10: 1657-1672Abstract (25) vitronectin-coated plates maintain hPSCs’ pluripotency. was initiated activate Wnt signaling. cultured media. When - 80% confluent, started replacing E6 CHIR99021. After 48 h, collected seeded I-coated containing TGF-? inhibitor, VEGF-A, ROCK (Y-27632). colonies seen days 5–6 differentiation. able incorporate host following injection ischemic (Cho 2020Cho Macklin B.L. Y.-Y. Zhou Lai M.J. Gerecht Duh E.J. iPSC-derived response hypoxia via SDF1a/CXCR4 axis facilitates incorporation revascularize retina.JCI Insight. 5: e131828Crossref (2) combined small-molecule modulation CHIR99021, chemically modified exogenous mRNA introduced cells. reproducible obtaining lines. authors also rates after only 96 h treatment (Wang 2020Wang R.-Z. Ng A.H. C.N. Neumeyer Ma Pu W.T. al.Robust temporal mRNA.Sci. Adv. eaba7606Crossref (4) colony-forming (ECFCs), subtype progenitor (EPCs), circulate becoming damaged (Paschalaki Randi, 2018Paschalaki K.E. Randi A.M. Recent colony forming toward translation.Front. Med. (Lausanne). 295Crossref (31) differentiate ECFCs three kinds factors: FGF2, activin-A, BMP4. repair limb (Prasain 2014Prasain M.R. Vemula Meador J.L. Yoshimoto Ferkowicz Fett Gupta Rapp B.M. Saadatzadeh cord-blood Biotechnol. 32: 1151-1157Crossref Transmembrane proteins, glycoproteins being identify characterize (Table 3). PECAM-1, known CD31, magnetic-activated sorting, cytometry, immunofluorescence confirming markers, verified vitro examining proinflammatory cytokines shear stress stimuli (Rosa 2019Rosa Praça Pitrez P.R. Gouveia P.J. Aranguren X.L. Ricotti Ferreira L.S. Functional arterial- venous-like cells.Sci. 3826Crossref (21) Mature form vessel-like tube networks 2D Matrigel-coated three-dimensional (3D) hydrogels. Intracellular molecule-1 (ICAM-1) (VCAM-1) associated modulating inflammation-induced adhesion. treating cytokine tumor necrosis (TNF)-?, ICAM-1 VCAM-1 upregulated recruit leukocytes (Kong 2018Kong D.-H. Y.K. Jang J.H. Emerging roles immunological disorders cancer.Int. Mol. 19: 1057Crossref (105) hallmark maturity flow. sensed mechano-sensors membrane, PI3K-Akt activated morphology (Fisher 2001Fisher Chien Barakat A.I. Nerem R.M. cellular altered stress.Am. Physiol. Lung Cell. 2001; 281: L529-L533Crossref Shear plays activating NOTCH1/DLL4 intracellular NOTCH1 cleaved activated, localization transmembrane domain adherence junction enhances (Polacheck 2017Polacheck W.J. Kutys M.L. Yang Eyckmans Vasavada Hirschi K.K. C.S. non-canonical Notch complex regulates adherens junctions function.Nature. 552: 258-262Crossref (108) xenograft assays. includes injecting alone together scaffolds zebrafish immunocompromised mice. Integration injected often observed 2–3 (Orlova 2014Orlova V.V. Drabsch Freund Petrus-Reurer den Hil F.E. Muenthaisong Dijke Mummery Functionality pericytes plexus xenografts.Arterioscler. 34: 177-186Crossref (122) Scholar; 2016Zhang Nör Cucco Shi J.E. Wnt/?-catenin determines vasculogenic fate postnatal mesenchymal cells.Stem 2016; 1576-1587Crossref methods classify not fully developed. Therefore, it imperative understand involved organogenesis.Table 3Characterization ECsMethodEC CharacteristicsReferenceMarker expressionPECAM-1 (CD31), VE-cad (CD144), Ulex europaeus agglutinin (UEA-l), vWf, eNOS, VEGFR2/KDR, CD34, CD105, Tie2, NOS3Wang Wimmer 2019Wimmer R.A. Leopoldi Aichinger Wick Hantusch Novatchkova Taubenschmid Hämmerle Esk Bagley J.A. vessel organoids diabetic vasculopathy.Nature. 565: 505-510Crossref (150) de Haan 2020de Øie Benkheil Dheedene Vinckier Coppiello Beerens Jaekers Topal al.Unraveling transcriptional determinants liver specialization.Am. Gastrointest. Liver 318: G803-G815Crossref Orlova Olmer ScholarIn tests• 2D: cord formation• 3D: formation, spheroid sprouting• Low-density lipoprotein (LDL) uptake• Nitric oxide production• stress: alignment direction flow, Proinflammatory challenge: expressionWang Cho Kraehling 2016Kraehling J.R. Chidlow Rajagopal Sugiyama M.G. Fowler M.Y. Ramírez C.M. Tao al.Genome-wide RNAi screen ALK1 mediates LDL uptake transcytosis Commun. 7: 13516Crossref 2017Zhang Chu L.-F. Schwartz M.P. Hacker Vickerman Swanson Leng B.K. Elwell al.Functional cells.Proc. 114: E6072-E6078Crossref Kong test: assays• Mouse: wound healing, retinopathy (OIR), ischemia, subcutaneous graft injection• Zebrafish: (blastula) injection, post-fertilization Cuvier duct injectionWang Vasculogenesis starts mesoderm-derived multipotent progenitors angioblasts plexus. During angiogenesis, yields capillaries, veins, arteries (Kume, 2010Kume Specification arterial, venous, lymphatic development.Histol. Histopathol. 2010; 25: 637-646PubMed Whereas veins direct heart respectively, capillaries closely surrounding deliver absorb nutrients. general regardless type, specification relies NOTCH (Slukvin Uenishi, 2019Slukvin I.I. Uenishi G.I. Arterial identity hemogenic endothelium: key unlock definitive hematopoietic commitment cultures.Exp. Hematol. 71: 3-12Abstract (11) Scholar), dependent COUP-TFII, EphB2, BMP/SMAD-mediated EphB4 ALK3/BMPR1A 2014Kim J.D. H.W. Jin S.W. Diversity my veins: bone illustrates 1838-1845Crossref Neal 2019aNeal Nornes Payne Wallace Fritzsche Louphrasitthiphol Wilkinson R.N. Chouliaras K.M. Plant al.Venous requires BMP ALK3.Nat. 453Crossref Recapitulation these arteriovenous has captured (Arora 2019Arora Yim E.K.F. Toh Y.C. Environmental derived subtypes.Front. Bioeng. 143Crossref (6) Starting precursors exhibiting CD31+/KDR+/VE-cad+/EphB2?/COUP-TF? phenotype, successfully modulated varying levels concentrations On basis level analyses, arterial-like cells, differentiated elevated concentration, umbilical artery (HUAECs), they NOTCH4, JAG1, HEY2, EFNB1, DLL4, CXCR4. contrast, low resemble vein (HUVECs) positive NR2F2, EphB4, COUP-TFII physiological stressors such proven influence (Huang 2019Huang Che Zhan Q. Ji Fan endothelium-oriented mouse-induced Cells Int. 2019: 1847098Crossref (3) Adams 2013Adams Cloutier Kuchimanchi Newton Sehrawat Mayadas T.N. Luscinskas F.W.