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Endothelial cells expressing low levels of CD143 (ACE) exhibit enhanced sprouting and potency in relieving tissue ischemia

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Abstract

The sprouting of endothelial cells from pre-existing blood vessels represents a critical event in the angiogenesis cascade. However, only a fraction of cultured or transplanted endothelial cells form new vessels. Moreover, it is unclear whether this results from a stochastic process or instead relates to certain endothelial cells having a greater angiogenic potential. This study investigated whether there exists a sub-population of cultured endothelial cells with enhanced angiogenic potency in vitro and in vivo. First, endothelial cells that participated in sprouting, and non-sprouting cells, were separately isolated from a 3D fibrin gel sprouting assay. Interestingly, the sprouting cells, when placed back into the same assay, displayed a sevenfold increase in the number of sprouts, as compared to control cells. Angiotensin-converting enzyme (CD143) was significantly down regulated on sprouting cells, as compared to regular endothelial cells. A subset of endothelial cells with low CD143 expression was then prospectively isolated from an endothelial cell culture. Finally, these cells were found to have greater potency in alleviating local ischemia, and restoring regional blood perfusion when transplanted into ischemic hindlimbs, as compared to unsorted endothelial cells. In summary, this study indicates that low expression of CD143 can be used as a biomarker to identify an endothelial cell sub-population that is more capable to drive neovascularization.

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References

  1. Carmeliet P, Jain R (2011) Molecular mechanisms and clinical applications of angiogenesis. Nature 473(7347):298–307. doi:10.1038/nature10144

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  2. Semenza G (2011) Oxygen sensing, homeostasis, and disease. New Engl J Med 365(6):537–547. doi:10.1056/NEJMra1011165

    Article  CAS  PubMed  Google Scholar 

  3. Potente M, Gerhardt H, Carmeliet P (2011) Basic and therapeutic aspects of angiogenesis. Cell 146(6):873–887. doi:10.1016/j.cell.2011.08.039

    Article  CAS  PubMed  Google Scholar 

  4. Davis G, Senger D (2005) Endothelial extracellular matrix: biosynthesis, remodeling, and functions during vascular morphogenesis and neovessel stabilization. Circ Res 97(11):1093–1107. doi:10.1161/01.RES.0000191547.64391.e3

    Article  CAS  PubMed  Google Scholar 

  5. Ribatti D, Crivellato E (2012) “Sprouting angiogenesis”, a reappraisal. Dev Biol 372(2):157–165. doi:10.1016/j.ydbio.2012.09.018

    Article  CAS  PubMed  Google Scholar 

  6. Gerhardt H, Golding M, Fruttiger M, Ruhrberg C, Lundkvist A, Abramsson A, Jeltsch M, Mitchell C, Alitalo K, Shima D, Betsholtz C (2003) VEGF guides angiogenic sprouting utilizing endothelial tip cell filopodia. J Cell Biol 161(6):1163–1177. doi:10.1083/jcb.200302047

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  7. Blancas AA, Wong LE, Glaser DE, McCloskey KE (2013) Specialized tip/stalk-like and phalanx-like endothelial cells from embryonic stem cells. Stem Cells Dev. doi:10.1089/scd.2012.0376

    PubMed Central  PubMed  Google Scholar 

  8. Carmeliet P, De Smet F, Loges S, Mazzone M (2009) Branching morphogenesis and antiangiogenesis candidates: tip cells lead the way. Nat Rev Clin Oncol 6(6):315–326. doi:10.1038/nrclinonc.2009.64

    Article  CAS  PubMed  Google Scholar 

  9. Staton C, Reed M, Brown N (2009) A critical analysis of current in vitro and in vivo angiogenesis assays. Int J Exp Pathol 90(3):195–221. doi:10.1111/j.1365-2613.2008.00633.x

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  10. Arnaoutova I, George J, Kleinman H, Benton G (2009) The endothelial cell tube formation assay on basement membrane turns 20: state of the science and the art. Angiogenesis 12(3):267–274. doi:10.1007/s10456-009-9146-4

    Article  PubMed  Google Scholar 

  11. Iruela-Arispe ML, George ED (2009) Cellular and molecular mechanisms of vascular lumen formation. Dev Cell 16. doi:10.1016/j.devcel.2009.01.013

  12. Li X, Claesson-Welsh L (2009) Embryonic stem cell models in vascular biology. JTH 7(Suppl 1):53–56. doi:10.1111/j.1538-7836.2009.03427.x

    CAS  PubMed  Google Scholar 

  13. Nakatsu M, CCW Hughes (2008) An optimized three-dimensional in vitro model for the analysis of angiogenesis. Methods Enzymol 443. doi:10.1016/s0076-6879(08)02004-1

  14. Zeitlin B, Dong Z, Nör J (2012) RAIN-droplet: a novel 3D in vitro angiogenesis model. Lab Invest 92(7):988–998. doi:10.1038/labinvest.2012.77

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  15. Auerbach R, Lewis R, Shinners B, Kubai L, Akhtar N (2003) Angiogenesis assays: a critical overview. Clin Chem 49(1):32–40. doi:10.1373/49.1.32

    Article  CAS  PubMed  Google Scholar 

  16. Jain R, Schlenger K, Höckel M, Yuan F (1997) Quantitative angiogenesis assays: progress and problems. Nat Med 3(11):1203–1208. doi:10.1038/nm1197-1203

    Article  CAS  PubMed  Google Scholar 

  17. Parikh S, Edelman E (2000) Endothelial cell delivery for cardiovascular therapy. Adv Drug Deliv Rev 42(1–2):139–161. doi:10.1016/S0169-409X(00)00058-2

    Article  CAS  PubMed  Google Scholar 

  18. Losordo D, Dimmeler S (2004) Therapeutic angiogenesis and vasculogenesis for ischemic disease: part II: cell-based therapies. Circulation 109(22):2692–2697. doi:10.1161/01.CIR.0000128596.49339.05

    Article  PubMed  Google Scholar 

  19. Dong Z, Neiva K, Jin T, Zhang Z, Hall D, Mooney D, Polverini P, Nör J (2007) Quantification of human angiogenesis in immunodeficient mice using a photon counting-based method. Biotechniques 43(1):73–77

    Article  CAS  PubMed  Google Scholar 

  20. Nör J, Peters M, Christensen J, Sutorik M, Linn S, Khan M, Addison C, Mooney D, Polverini P (2001) Engineering and characterization of functional human microvessels in immunodeficient mice. Lab Invest 81(4):453–463. doi:10.1038/labinvest.3780253

    Article  PubMed  Google Scholar 

  21. Taraboletti G, Giavazzi R (2004) Modelling approaches for angiogenesis. Eur J Cancer 40(6):881–889. doi:10.1016/j.ejca.2004.01.002 Oxford, England: 1990

    Article  CAS  PubMed  Google Scholar 

  22. Newman A, Nakatsu M, Chou W, Gershon P, Hughes C (2011) The requirement for fibroblasts in angiogenesis: fibroblast-derived matrix proteins are essential for endothelial cell lumen formation. Mol Biol Cell 22(20):3791–3800. doi:10.1091/mbc.E11-05-0393

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  23. Nakatsu M, Sainson R, Aoto J, Taylor K, Aitkenhead M, Pérez-del-Pulgar S, Carpenter P, Hughes C (2003) Angiogenic sprouting and capillary lumen formation modeled by human umbilical vein endothelial cells (HUVEC) in fibrin gels: the role of fibroblasts and Angiopoietin-1. Microvasc Res 66(2):102–112. doi:10.1016/s0026-2862(03)00045-1

    Article  CAS  PubMed  Google Scholar 

  24. Couffinhal T, Silver M, Kearney M, Sullivan A, Witzenbichler B, Magner M, Annex B, Peters K, Isner JM (1999) Impaired collateral vessel development associated with reduced expression of vascular endothelial growth factor in ApoE-/- mice. Circulation 99(24):3188–3198

    Article  CAS  PubMed  Google Scholar 

  25. Silva EA, Mooney DJ (2010) Effects of VEGF temporal and spatial presentation on angiogenesis. Biomaterials 31(6):1235–1241. doi:10.1016/j.biomaterials.2009.10.052

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  26. Silva EA, Kim ES, Kong HJ, Mooney DJ (2008) Material-based deployment enhances efficacy of endothelial progenitor cells. Proc Natl Acad Sci USA 105(38):14347–14352. doi:10.1073/pnas.0803873105

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  27. Chen RR, Silva EA, Yuen WW, Brock AA, Fischbach C, Lin AS, Guldberg RE, Mooney DJ (2007) Integrated approach to designing growth factor delivery systems. FASEB J 21(14):3896–3903. doi:10.1096/fj.06-7873com

    Article  CAS  PubMed  Google Scholar 

  28. Yuen WW, Du NR, Chan CH, Silva EA, Mooney DJ (2010) Mimicking nature by codelivery of stimulant and inhibitor to create temporally stable and spatially restricted angiogenic zones. Proc Natl Acad Sci USA 107(42):17933–17938. doi:10.1073/pnas.1001192107

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  29. Silva EA, Mooney DJ (2007) Spatiotemporal control of vascular endothelial growth factor delivery from injectable hydrogels enhances angiogenesis. J Thromb Haemost 5(3):590–598. doi:10.1111/j.1538-7836.2007.02386.x

    Article  CAS  PubMed  Google Scholar 

  30. Iruela-Arispe M, Davis G (2009) Cellular and molecular mechanisms of vascular lumen formation. Dev Cell 16(2):222–231. doi:10.1016/j.devcel.2009.01.013

    Article  CAS  PubMed  Google Scholar 

  31. Nakatsu M, Davis J, Hughes C (2007) Optimized fibrin gel bead assay for the study of angiogenesis. J Vis Exp 3:186. doi:10.3791/186

    PubMed  Google Scholar 

  32. Sieminski AL, Hebbel RP, Gooch KJ (2005) Improved microvascular network in vitro by human blood outgrowth endothelial cells relative to vessel-derived endothelial cells. Tissue Eng 11(9–10):1332–1345. doi:10.1089/ten.2005.11.1332

    Article  CAS  PubMed  Google Scholar 

  33. Fantin A, Vieira JM, Plein A, Denti L, Fruttiger M, Pollard JW, Ruhrberg C (2013) NRP1 acts cell autonomously in endothelium to promote tip cell function during sprouting angiogenesis. Blood 121(12):2352–2362. doi:10.1182/blood-2012-05-424713

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  34. Law AY, Wong CK (2013) Stanniocalcin-1 and -2 promote angiogenic sprouting in HUVECs via VEGF/VEGFR2 and angiopoietin signaling pathways. Mol Cell Endocrinol 374(1–2):73–81. doi:10.1016/j.mce.2013.04.024

    Article  CAS  PubMed  Google Scholar 

  35. Hou TC, Lin JJ, Wen HC, Chen LC, Hsu SP, Lee WS (2013) Folic acid inhibits endothelial cell migration through inhibiting the RhoA activity mediated by activating the folic acid receptor/cSrc/p190RhoGAP-signaling pathway. Biochem Pharmacol 85(3):376–384. doi:10.1016/j.bcp.2012.11.011

    Article  CAS  PubMed  Google Scholar 

  36. Spindler V, Schlegel N, Waschke J (2010) Role of GTPases in control of microvascular permeability. Cardiovasc Res 87(2):243–253. doi:10.1093/cvr/cvq086

    Article  CAS  PubMed  Google Scholar 

  37. van Nieuw Amerongen GP, Koolwijk P, Versteilen A, van Hinsbergh VW (2003) Involvement of RhoA/Rho kinase signaling in VEGF-induced endothelial cell migration and angiogenesis in vitro. Arterioscler Thromb Vasc Biol 23(2):211–217

    Article  PubMed  Google Scholar 

  38. Wacker A, Gerhardt H (2011) Endothelial development taking shape. Curr Opin Cell Biol 23(6):676–685. doi:10.1016/j.ceb.2011.10.002

    CAS  PubMed  Google Scholar 

  39. Blanco R, Gerhardt H (2013) VEGF and Notch in tip and stalk cell selection. Cold Spring Harb Perspect Med 3(1). doi:10.1101/cshperspect.a006569

  40. Kume T (2012) Ligand-dependent notch signaling in vascular formation. Adv Exp Med Biol 727:210–222. doi:10.1007/978-1-4614-0899-4_16

    Article  CAS  PubMed  Google Scholar 

  41. del Toro R, Prahst C, Mathivet T, Siegfried G, Kaminker JS, Larrivee B, Breant C, Duarte A, Takakura N, Fukamizu A, Penninger J, Eichmann A (2010) Identification and functional analysis of endothelial tip cell-enriched genes. Blood 116(19):4025–4033. doi:10.1182/blood-2010-02-270819

    Article  PubMed  Google Scholar 

  42. Lei Y, Zouani OF, Rami L, Chanseau C, Durrieu MC (2012) Modulation of lumen formation by microgeometrical bioactive cues and migration mode of actin machinery. Small. doi:10.1002/smll.201202410

    Google Scholar 

  43. Napoli C, Hayashi T, Cacciatore F, Casamassimi A, Casini C, Al-Omran M, Ignarro LJ (2011) Endothelial progenitor cells as therapeutic agents in the microcirculation: an update. Atherosclerosis 215(1):9–22. doi:10.1016/j.atherosclerosis.2010.10.039

    Article  CAS  PubMed  Google Scholar 

  44. Gerhardt H (2003) VEGF guides angiogenic sprouting utilizing endothelial tip cell filopodia. J Cell Biol 161. doi:10.1083/jcb.200302047

  45. Hercus TR, Broughton SE, Ekert PG, Ramshaw HS, Perugini M, Grimbaldeston M, Woodcock JM, Thomas D, Pitson S, Hughes T, D’Andrea RJ, Parker MW, Lopez AF (2012) The GM-CSF receptor family: mechanism of activation and implications for disease. Growth Factors 30(2):63–75. doi:10.3109/08977194.2011.649919

    Article  CAS  PubMed  Google Scholar 

  46. Stamenkovic I (1995) The L-selectin adhesion system. Curr Opin Hematol 2(1):68–75

    Article  CAS  PubMed  Google Scholar 

  47. Shah IM, Macrae IM, Di Napoli M (2009) Neuroinflammation and neuroprotective strategies in acute ischaemic stroke—from bench to bedside. Curr Mol Med 9(3):336–354

    Article  CAS  PubMed  Google Scholar 

  48. Suffee N, Richard B, Hlawaty H, Oudar O, Charnaux N, Sutton A (2011) Angiogenic properties of the chemokine RANTES/CCL5. Biochem Soc Trans 39(6):1649–1653. doi:10.1042/BST20110651

    Article  CAS  PubMed  Google Scholar 

  49. Bourghardt Peebo B, Fagerholm P, Traneus-Rockert C, Lagali N (2011) Time-lapse in vivo imaging of corneal angiogenesis: the role of inflammatory cells in capillary sprouting. Invest Ophthalmol Vis Sci 52(6):3060–3068. doi:10.1167/iovs.10-6101

    Article  PubMed  Google Scholar 

  50. Arroyo AG, Iruela-Arispe ML (2010) Extracellular matrix, inflammation, and the angiogenic response. Cardiovasc Res 86(2):226–235. doi:10.1093/cvr/cvq049

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  51. Crackower MA, Sarao R, Oudit GY, Yagil C, Kozieradzki I, Scanga SE, Oliveira-dos-Santos AJ, da Costa J, Zhang L, Pei Y, Scholey J, Ferrario CM, Manoukian AS, Chappell MC, Backx PH, Yagil Y, Penninger JM (2002) Angiotensin-converting enzyme 2 is an essential regulator of heart function. Nature 417(6891):822–828. doi:10.1038/nature00786

    Article  CAS  PubMed  Google Scholar 

  52. English WR, Corvol P, Murphy G (2012) LPS activates ADAM9 dependent shedding of ACE from endothelial cells. Biochem Biophys Res Commun 421(1):70–75. doi:10.1016/j.bbrc.2012.03.113

    Article  CAS  PubMed  Google Scholar 

  53. Corvol P, Michaud A, Soubrier F, Williams TA (1995) Recent advances in knowledge of the structure and function of the angiotensin I converting enzyme. J Hypertens Suppl 13(3):S3–10

    Article  CAS  PubMed  Google Scholar 

  54. Niu T, Chen X, Xu X (2002) Angiotensin converting enzyme gene insertion/deletion polymorphism and cardiovascular disease: therapeutic implications. Drugs 62(7):977–993

    Article  CAS  PubMed  Google Scholar 

  55. Kohlstedt K, Trouvain C, Boettger T, Shi L, Fisslthaler B, Fleming I (2013) The AMP-activated protein kinase regulates endothelial cell angiotensin-converting enzyme expression via p53 and the post-transcriptional regulation of microRNA-143/145. Circ Res. doi:10.1161/CIRCRESAHA.113.301282

    PubMed  Google Scholar 

  56. Paulis L, Unger T (2010) Novel therapeutic targets for hypertension. Nat Rev Cardiol 7(8):431–441. doi:10.1038/nrcardio.2010.85

    Article  CAS  PubMed  Google Scholar 

  57. Hamdi HK, Castellon R (2003) ACE inhibition actively promotes cell survival by altering gene expression. Biochem Biophy Res Commun 310(4):1227–1235

    Article  CAS  Google Scholar 

  58. Rubin H (1997) Cell aging in vivo and in vitro. Mech Ageing Dev 98(1):1–35

    CAS  PubMed  Google Scholar 

  59. Ljungberg LU, Persson K (2008) Effect of nicotine and nicotine metabolites on angiotensin-converting enzyme in human endothelial cells. Endothelium 15(5–6):239–245. doi:10.1080/10623320802487627

    Article  CAS  PubMed  Google Scholar 

  60. Mima Y, Fukumoto S, Koyama H, Okada M, Tanaka S, Shoji T, Emoto M, Furuzono T, Nishizawa Y, Inaba M (2012) Enhancement of cell-based therapeutic angiogenesis using a novel type of injectable scaffolds of hydroxyapatite-polymer nanocomposite microspheres. PLoS ONE 7(4):e35199. doi:10.1371/journal.pone.0035199

    Article  CAS  PubMed Central  PubMed  Google Scholar 

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Acknowledgments

The authors thank Brian Tilton, Patricia Rogers (FAS Center for Systems Biology), Catia Verbeke and Cristiana Cunha for all the help and support with the flow cytometry. Financial support of this research was provided by the NIH (R01 HL069957), and we thank the National Cancer Institute for providing VEGF165.

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Authors and Affiliations

  1. Wyss Institute for Biologically Inspired Engineering, Harvard University, Cambridge, MA, USA

    Eduardo A. Silva & David J. Mooney

  2. School of Engineering and Applied Sciences, Harvard University, Cambridge, MA, USA

    Eduardo A. Silva, Chikezie Eseonu & David J. Mooney

  3. School of Medicine, Yale University, New Haven, CT, USA

    Chikezie Eseonu

  4. Department of Biomedical Engineering, University of California, Davis, CA, USA

    Eduardo A. Silva

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  1. Eduardo A. Silva
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Correspondence to David J. Mooney.

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Silva, E.A., Eseonu, C. & Mooney, D.J. Endothelial cells expressing low levels of CD143 (ACE) exhibit enhanced sprouting and potency in relieving tissue ischemia. Angiogenesis 17, 617–630 (2014). https://doi.org/10.1007/s10456-014-9414-9

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