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Modeling angiogenesis in the human brain in a tissue-engineered post-capillary venule

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Abstract

Angiogenesis plays an essential role in embryonic development, organ remodeling, wound healing, and is also associated with many human diseases. The process of angiogenesis in the brain during development is well characterized in animal models, but little is known about the process in the mature brain. Here, we use a tissue-engineered post-capillary venule (PCV) model incorporating stem cell derived induced brain microvascular endothelial-like cells (iBMECs) and pericyte-like cells (iPCs) to visualize the dynamics of angiogenesis. We compare angiogenesis under two conditions: in response to perfusion of growth factors and in the presence of an external concentration gradient. We show that both iBMECs and iPCs can serve as tip cells leading angiogenic sprouts. More importantly, the growth rate for iPC-led sprouts is about twofold higher than for iBMEC-led sprouts. Under a concentration gradient, angiogenic sprouts show a small directional bias toward the high growth factor concentration. Overall, pericytes exhibited a broad range of behavior, including maintaining quiescence, co-migrating with endothelial cells in sprouts, or leading sprout growth as tip cells.

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References

  1. Paredes I, Himmels P, de Almodóvar CR (2018) Neurovascular communication during CNS development. Dev Cell 45(1):10–32

    Article  CAS  PubMed  Google Scholar 

  2. Tata M, Ruhrberg C, Fantin A (2015) Vascularisation of the central nervous system. Mech Dev 138:26–36

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Brown CE et al (2007) Extensive turnover of dendritic spines and vascular remodeling in cortical tissues recovering from stroke. J Neurosci 27(15):4101–4109

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Jiang X et al (2018) Blood-brain barrier dysfunction and recovery after ischemic stroke. Prog Neurobiol 163–164:144–171

    Article  PubMed  Google Scholar 

  5. Yao Y (2018) Basement membrane and stroke. J Cereb Blood Flow Metab. https://doi.org/10.1177/0271678X18801467

    Article  PubMed  PubMed Central  Google Scholar 

  6. Croll SD, Wiegand SJ (2001) Vascular growth factors in cerebral ischemia. Mol Neurobiol 23(2–3):121–135

    Article  CAS  PubMed  Google Scholar 

  7. Lange C et al (2016) Vascular endothelial growth factor: a neurovascular target in neurological diseases. Nat Rev Neurol 12(8):439–454

    Article  CAS  PubMed  Google Scholar 

  8. Jain RK et al (2007) Angiogenesis in brain tumours. Nat Rev Neurosci 8(8):610–622

    Article  CAS  PubMed  Google Scholar 

  9. Salehi A, Zhang JH, Obenaus A (2017) Response of the cerebral vasculature following traumatic brain injury. J Cereb Blood Flow Metab 37(7):2320–2339

    Article  PubMed  PubMed Central  Google Scholar 

  10. Shlosberg D et al (2010) Blood-brain barrier breakdown as a therapeutic target in traumatic brain injury. Nat Rev Neurol 6(7):393–403

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Swain RA et al (2003) Prolonged exercise induces angiogenesis and increases cerebral blood volume in primary motor cortex of the rat. Neuroscience 117(4):1037–1046

    Article  CAS  PubMed  Google Scholar 

  12. Pereira, A.C., et al., An in vivo correlate of exercise-induced neurogenesis in the adult dentate gyrus. Proceedings of the National Academy of Sciences, 2007. 104(13): p. 5638-5643.

  13. Park HY et al (2016) The effects of altitude/hypoxic training on oxygen delivery capacity of the blood and aerobic exercise capacity in elite athletes - a meta-analysis. J Exerc Nutrition Biochem 20(1):15–22

    Article  PubMed  PubMed Central  Google Scholar 

  14. Bogorad MI et al (2019) Cerebrovascular plasticity: processes that lead to changes in the architecture of brain microvessels. J Cereb Blood Flow Metab 39(8):1413–1432

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Lindahl P et al (1997) Pericyte loss and microaneurysm formation in PDGF-B-deficient mice. Science 277(5323):242–245

    Article  CAS  PubMed  Google Scholar 

  16. Daneman R et al (2010) Pericytes are required for blood–brain barrier integrity during embryogenesis. Nature 468(7323):562–566

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Ogunshola OO et al (2000) Neuronal VEGF expression correlates with angiogenesis in postnatal develo** rat brain. Dev Brain Res 119(1):139–153

    Article  CAS  Google Scholar 

  18. Virgintino D et al (2007) An intimate interplay between precocious, migrating pericytes and endothelial cells governs human fetal brain angiogenesis. Angiogenesis 10(1):35–45

    Article  PubMed  Google Scholar 

  19. Wälchli T et al (2015) Quantitative assessment of angiogenesis, perfused blood vessels and endothelial tip cells in the postnatal mouse brain. Nat Protoc 10(1):53–74

    Article  PubMed  Google Scholar 

  20. Carmeliet P, Jain RK (2011) Molecular mechanisms and clinical applications of angiogenesis. Nature 473(7347):298–307

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Gerhardt H, Betsholtz C (2003) Endothelial-pericyte interactions in angiogenesis. Cell Tissue Res 314(1):15–23

    Article  PubMed  Google Scholar 

  22. Benjamin LE, Hemo I, Keshet E (1998) A plasticity window for blood vessel remodelling is defined by pericyte coverage of the preformed endothelial network and is regulated by PDGF-B and VEGF. Development 125(9):1591–1598

    Article  CAS  PubMed  Google Scholar 

  23. Potente M, Gerhardt H, Carmeliet P (2011) Basic and therapeutic aspects of angiogenesis. Cell 146(6):873–887

    Article  CAS  PubMed  Google Scholar 

  24. Conway EM, Collen D, Carmeliet P (2001) Molecular mechanisms of blood vessel growth. Cardiovasc Res 49(3):507–521

    Article  CAS  PubMed  Google Scholar 

  25. Coelho-Santos, V., et al., Imaging the construction of capillary networks in the neonatal mouse brain. Proceedings of the National Academy of Sciences, 2021. 118(26).

  26. Fruttiger M (2002) Development of the mouse retinal vasculature: angiogenesis versus vasculogenesis. Invest Ophthalmol Vis Sci 43(2):522–527

    PubMed  Google Scholar 

  27. Wälchli, T., et al., Nogo-A is a negative regulator of CNS angiogenesis. Proceedings of the National Academy of Sciences, 2013. 110(21): p. E1943-E1952.

  28. Nehls V, Denzer K, Drenckhahn D (1992) Pericyte involvement in capillary sprouting during angiogenesis in situ. Cell Tissue Res 270(3):469–474

    Article  CAS  PubMed  Google Scholar 

  29. Amselgruber W, Schäfer M, Sinowatz F (1999) Angiogenesis in the bovine corpus luteum: an immunocytochemical and ultrastructural study. Anat Histol Embryol 28(3):157–166

    Article  CAS  PubMed  Google Scholar 

  30. DeCicco-Skinner KL et al (2014) Endothelial cell tube formation assay for the in vitro study of angiogenesis. JoVE (Journal of Visualized Experiments) 91:e51312

    Google Scholar 

  31. Nakatsu MN, Davis J, Hughes CC (2007) Optimized fibrin gel bead assay for the study of angiogenesis. JoVE (Journal of Visualized Experiments) 3:e186

    Google Scholar 

  32. Linville RM et al (2020) Three-dimensional induced pluripotent stem-cell models of human brain angiogenesis. Microvasc Res 132:104042

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Nguyen, D.-H.T., et al., Biomimetic model to reconstitute angiogenic sprouting morphogenesis in vitro. Proceedings of the National Academy of Sciences, 2013. 110(17): p. 6712-6717.

  34. Shin Y et al (2011) In vitro 3D collective sprouting angiogenesis under orchestrated ANG-1 and VEGF gradients. Lab Chip 11(13):2175–2181

    Article  CAS  PubMed  Google Scholar 

  35. Shirure VS et al (2017) Low levels of physiological interstitial flow eliminate morphogen gradients and guide angiogenesis. Angiogenesis 20(4):493–504

    Article  CAS  PubMed  Google Scholar 

  36. Abaci HE et al (2014) Recapitulating physiological and pathological shear stress and oxygen to model vasculature in health and disease. Sci Rep 4(1):1–9

    Article  Google Scholar 

  37. Lee E et al (2018) A 3D in vitro pericyte-supported microvessel model: visualisation and quantitative characterisation of multistep angiogenesis. J Mater Chem B 6(7):1085–1094

    Article  CAS  PubMed  Google Scholar 

  38. Silvestri VL et al (2020) A tissue-engineered 3D microvessel model reveals the dynamics of mosaic vessel formation in breast cancer. Can Res 80(19):4288–4301

    Article  CAS  Google Scholar 

  39. Price GM et al (2010) Effect of mechanical factors on the function of engineered human blood microvessels in microfluidic collagen gels. Biomaterials 31(24):6182–6189

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Kim S et al (2013) Engineering of functional, perfusable 3D microvascular networks on a chip. Lab Chip 13(8):1489–1500

    Article  CAS  PubMed  Google Scholar 

  41. Lee H et al (2014) A microfluidic platform for quantitative analysis of cancer angiogenesis and intravasation. Biomicrofluidics 8(5):054102

    Article  PubMed  PubMed Central  Google Scholar 

  42. Zervantonakis, I.K., et al., Three-dimensional microfluidic model for tumor cell intravasation and endothelial barrier function. Proceedings of the National Academy of Sciences, 2012. 109(34): pp. 13515-13520.

  43. Morgan JP et al (2013) Formation of microvascular networks in vitro. Nat Protoc 8(9):1820–1836

    Article  PubMed  Google Scholar 

  44. Bischel LL et al (2013) Tubeless microfluidic angiogenesis assay with three-dimensional endothelial-lined microvessels. Biomaterials 34(5):1471–1477

    Article  CAS  PubMed  Google Scholar 

  45. Kim C et al (2015) A quantitative microfluidic angiogenesis screen for studying anti-angiogenic therapeutic drugs. Lab Chip 15(1):301–310

    Article  PubMed  PubMed Central  Google Scholar 

  46. Zheng, Y., et al., In vitro microvessels for the study of angiogenesis and thrombosis. Proceedings of the national academy of sciences, 2012. 109(24): pp. 9342-9347.

  47. Tran KA et al (2022) Oxygen gradients dictate angiogenesis but not barriergenesis in a 3D brain microvascular model. J Cellular Physiol. https://doi.org/10.1002/jcp.30840

    Article  Google Scholar 

  48. Zhao N et al (2022) Engineering the human blood-brain barrier at the capillary scale using a double-templating technique. Adv Functional Mater. https://doi.org/10.1002/adfm.202110289

    Article  Google Scholar 

  49. Linville RM et al (2019) Human iPSC-derived blood-brain barrier microvessels: validation of barrier function and endothelial cell behavior. Biomaterials 190:24–37

    Article  PubMed  Google Scholar 

  50. Lippmann ES et al (2012) Derivation of blood-brain barrier endothelial cells from human pluripotent stem cells. Nat Biotechnol 30(8):783–791

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Lippmann ES et al (2020) Commentary on human pluripotent stem cell-based blood–brain barrier models. Fluids and Barriers of the CNS 17(1):1–6

    Article  Google Scholar 

  52. Linville RM et al (2020) Long-term cryopreservation preserves blood–brain barrier phenotype of iPSC-derived brain microvascular endothelial cells and three-dimensional microvessels. Mol Pharm 17(9):3425–3434

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Wang K et al (2020) Robust differentiation of human pluripotent stem cells into endothelial cells via temporal modulation of ETV2 with modified mRNA. Sci Adv 6(30):7606

    Article  Google Scholar 

  54. Stebbins MJ et al (2019) Human pluripotent stem cell–derived brain pericyte–like cells induce blood-brain barrier properties. Sci Adv 5(3):eaau7375

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Jamieson JJ et al (2019) Role of iPSC-derived pericytes on barrier function of iPSC-derived brain microvascular endothelial cells in 2D and 3D. Fluids and Barriers of the CNS 16(1):1–16

    Article  CAS  Google Scholar 

  56. Wong AD et al (2013) The blood-brain barrier: an engineering perspective. Front Neuroeng 6:7

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Huxley VH, Curry FE, Adamson RH (1987) Quantitative fluorescence microscopy on single capillaries - alpha-lactalbumin transport. Am J Physiol 252(1):H188–H197

    CAS  PubMed  Google Scholar 

  58. Red-Horse K et al (2010) Coronary arteries form by developmental reprogramming of venous cells. Nature 464(7288):549–553

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Bussmann J, Wolfe SA, Siekmann AF (2011) Arterial-venous network formation during brain vascularization involves hemodynamic regulation of chemokine signaling. Development 138(9):1717–1726

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. Xu C et al (2014) Arteries are formed by vein-derived endothelial tip cells. Nat Commun 5(1):1–11

    Article  Google Scholar 

  61. Lee H-W et al (2021) Role of venous endothelial cells in developmental and pathologic angiogenesis. Circulation 144(16):1308–1322

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  62. Abbott NJ et al (2010) Structure and function of the blood-brain barrier. Neurobiol Dis 37(1):13–25

    Article  CAS  PubMed  Google Scholar 

  63. Sweeney MD et al (2019) Blood-brain barrier: from physiology to disease and back. Physiol Rev 99(1):21–78

    Article  CAS  PubMed  Google Scholar 

  64. Lippmann ES et al (2020) Commentary on human pluripotent stem cell-based blood-brain barrier models. Fluids Barriers CNS 17(1):64

    Article  PubMed  PubMed Central  Google Scholar 

  65. Linville RM, Searson PC (2021) Next-generation in vitro blood-brain barrier models: benchmarking and improving model accuracy. Fluids Barriers CNS 18(1):56

    Article  PubMed  PubMed Central  Google Scholar 

  66. Wu H et al (1996) VEGF induces NO-dependent hyperpermeability in coronary venules. Am J Physiology-Heart Circulatory Physiol 271(6):H2735–H2739

    Article  CAS  Google Scholar 

  67. Bates D, Curry F (1996) Vascular endothelial growth factor increases hydraulic conductivity of isolated perfused microvessels. Am J Physiol-Heart Circulatory Physiol 271(6):H2520–H2528

    Article  CAS  Google Scholar 

  68. Weis S et al (2004) Src blockade stabilizes a Flk/cadherin complex, reducing edema and tissue injury following myocardial infarction. J Clin Investig 113(6):885–894

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  69. Roberts WG, Palade GE (1995) Increased microvascular permeability and endothelial fenestration induced by vascular endothelial growth factor. J Cell Sci 108(6):2369–2379

    Article  CAS  PubMed  Google Scholar 

  70. Ghajar CM et al (2006) Mesenchymal stem cells enhance angiogenesis in mechanically viable prevascularized tissues via early matrix metalloproteinase upregulation. Tissue Eng 12(10):2875–2888

    Article  CAS  PubMed  Google Scholar 

  71. Galie PA et al (2014) Fluid shear stress threshold regulates angiogenic sprouting. Proc Natl Acad Sci 111(22):7968–7973

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  72. Phng L-K, Gerhardt H (2009) Angiogenesis: a team effort coordinated by notch. Dev Cell 16(2):196–208

    Article  CAS  PubMed  Google Scholar 

  73. Mathiisen TM et al (2010) The perivascular astroglial sheath provides a complete covering of the brain microvessels: an electron microscopic 3D reconstruction. Glia 58(9):1094–1103

    Article  PubMed  Google Scholar 

  74. Frank RN et al (1983) Galactose-induced retinal capillary basement membrane thickening: prevention by Sorbinil. Invest Ophthalmol Vis Sci 24(11):1519–1524

    CAS  PubMed  Google Scholar 

  75. Crouch EE, Doetsch F (2018) FACS isolation of endothelial cells and pericytes from mouse brain microregions. Nat Protoc 13(4):738–751

    Article  CAS  PubMed  Google Scholar 

  76. Chen J et al (2017) CD146 coordinates brain endothelial cell–pericyte communication for blood–brain barrier development. Proc Natl Acad Sci 114(36):E7622–E7631

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  77. Kang T-Y et al (2019) Pericytes enable effective angiogenesis in the presence of proinflammatory signals. Proc Natl Acad Sci 116(47):23551–23561

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  78. Kim J et al (2015) Engineering of a biomimetic pericyte-covered 3D microvascular network. PLoS ONE 10(7):e0133880

    Article  PubMed  PubMed Central  Google Scholar 

  79. Campisi M et al (2018) 3D self-organized microvascular model of the human blood-brain barrier with endothelial cells, pericytes and astrocytes. Biomaterials 180:117–129

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  80. Gerhardt H et al (2003) VEGF guides angiogenic sprouting utilizing endothelial tip cell filopodia. J Cell Biol 161(6):1163–1177

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  81. Linville RM et al (2022) Three-dimensional microenvironment regulates gene expression, function, and tight junction dynamics of iPSC-derived blood–brain barrier microvessels. Fluids Barriers of the CNS 19(1):1–18

    Article  Google Scholar 

  82. Weis SM, Cheresh DA (2005) Pathophysiological consequences of VEGF-induced vascular permeability. Nature 437(7058):497–504

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

The authors would like to thank Mr. Grant Kitchen, Dr. Dinh-Tuan Phan, and Dr. Yi Lu for help with the design and fabrication of the mold. This work was supported by NIH (NINDS R01NS106008 and NHLBI R61HL154252).

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Authors and Affiliations

  1. Institute for Nanobiotechnology, Johns Hopkins University, Baltimore, MD, 21218, USA

    Nan Zhao, Raleigh M. Linville, Tracy D. Chung, Zhaobin Guo, John J. Jamieson, Lily Liang, Alexander F. Pessell & Peter Searson

  2. Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD, 21218, USA

    Sarah Kulkarni, Raleigh M. Linville, Tracy D. Chung, Lily Liang, Alexander F. Pessell & Peter Searson

  3. Department of Biology, Johns Hopkins University, Baltimore, MD, 21218, USA

    Sophia Zhang

  4. Department of Chemical and Biomolecular Engineering, Johns Hopkins University, Baltimore, MD, 21218, USA

    John J. Jamieson & Danielle Norman

  5. Department of Materials Science and Engineering, Johns Hopkins University, Baltimore, MD, 21218, USA

    Peter Searson

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  1. Nan Zhao
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  2. Sarah Kulkarni
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  3. Sophia Zhang
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  4. Raleigh M. Linville
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  8. Danielle Norman
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  9. Lily Liang
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  10. Alexander F. Pessell
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  11. Peter Searson
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Contributions

NZ and PS conceived the original idea. NZ wrote the manuscript. NZ, SK, SZ, RL, TC, ZG, JJ, DN, LL, and AP contributed to the data acquisition, analysis, and interpretation. PS supervised all work.

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Correspondence to Peter Searson.

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Zhao, N., Kulkarni, S., Zhang, S. et al. Modeling angiogenesis in the human brain in a tissue-engineered post-capillary venule. Angiogenesis 26, 203–216 (2023). https://doi.org/10.1007/s10456-023-09868-7

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