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Human breast microvascular endothelial cells retain phenotypic traits in long-term finite life span culture

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Summary

Attempts to study endothelial-epithelial interactions in the human breast have been hampered by lack of protocols for long-term cultivation of breast endothelial cells (BRENCs). The aim of this study was to establish long-term cultures of BRENCs and to compare their phenotypic traits with the tissue of origin. Microvasculature was localized in situ by immunohistochemitry in breast samples. From this tissue, collagen-rich stroma and adipose tissue were dissected mechanically and further disaggregated to release microvessel organoids BRENCs were cultured from these organoids in endothelial specific medium and characterized by staining for endothelial markers. Microvessels were a prominent feature of intralobular tissue as evidenced by immunostaining against endothelial specific markers such as CD31, VE-cadherin, and von Willebrand factor (VWF). Double staining against VE-cadherin and lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1) showed that blood and lymphatic vessels could be distinguished. An antibody against CD31 was used to refine protocols for isolation of microvasculature from reduction mammoplasties. BRENCs retained critical traits even at high passage, including uptake of low-density lipoprotein, and had E-selectin induced upon treatment with tumor necrosis factor-α. The first signs of senescence in passage 14 were accompained by gain of trisomy 11. At passage 18 cells showed chromosomal aberrations and growth arrest as revealed by β-galactosidase staining. We demonstrate here that breast microvasculature may serve as a large-scale source for expansion of BRENCs with molecular and functional traits preserved. These cells will form the basis for studies on the role of endothelial cells in breast morphogenesis.

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References

  • Abbott, N. J. Astrocyte-endothelial interactions and blood-brain barrier permeability. J. Anat. 200:629–638; 2002.

    Article  PubMed  CAS  Google Scholar 

  • Bachetti, T.; Morbidelli, L. Endothelial cells in culture: a model for studying vascular functions. Pharmacol. Res. 42:9–19; 2000.

    Article  PubMed  CAS  Google Scholar 

  • Belloni, P. N.; Nicolson, G. L. Differential expression of cell surface glycoproteins on various organ-derived microvascular endothelia and endothelial cell cultures. J. Cell. Physiol. 136:398–410; 1988.

    Article  PubMed  CAS  Google Scholar 

  • Bissell, M. J.; Radisky, D. C.; Rizki, A.; Weaver, V. M.; Petersen, O. W. The organizing principle: microenvironmental influences in the normal and malignant breast. Differentiation 70:537–546; 2002.

    Article  PubMed  Google Scholar 

  • Boudreau, N.; Myers, C. Breast cancer-induced angiogenesis: multiple mechanisms and the role of the microenvironment. Breast Cancer Res. 5:140–146; 2003.

    Article  PubMed  CAS  Google Scholar 

  • Bouis, D.; Hospers, G. A.; Meijer, C.; Molema, G.; Mulder, N. H. Endothelium in vitro: a review of human vascular endothelial cell lines for blood vessel-related research. Angiogenesis 4:91–102; 2001.

    Article  PubMed  CAS  Google Scholar 

  • Chi, J. T.; Chang, H. Y.; Haraldsen, G., et al.: Endothelial cell diversity revealed by global expression profiling. Proc. Natl. Acad. Sci. USA 100:10623–10628; 2003.

    Article  PubMed  CAS  Google Scholar 

  • Cleaver, O.; Melton, D. A. Endothelial signaling during development. Nat. Med. 9:661–668; 2003.

    Article  PubMed  CAS  Google Scholar 

  • Dabrosin, C. Increase of free insulin-like growth factor-1 in normal human breast in vivo late in the menstrual cycle. Breast Cancer Res. Treat. 80:193–198; 2003a.

    Article  PubMed  CAS  Google Scholar 

  • Dabrosin, C. Variability of vascular endothelial growth factor in normal human breast tissue in vivo during the menstrual cycle. J. Clin. Endocrinol. Metab. 88: 2695–2698; 2003b.

    Article  PubMed  CAS  Google Scholar 

  • Davison, P. M.; Bensch, K.; Karasek, M. A. Isolation and long-term serial cultivation of endothelial cells from the microvessels of the adult human dermis. In Vitro 19:937–945; 1983.

    PubMed  CAS  Google Scholar 

  • Dorovini-Zis, K.; Prameya, R.; Bowman, P. D. Culture and characterization of microvascular endothelial cells derived from human brain. Lab. Invest. 64:425–436; 1991.

    PubMed  CAS  Google Scholar 

  • Folkman, J.; Haudenschild, C. C.; Zetter, B. R. Long-term culture of capillary endothelial cells. Proc. Natl. Acad. Sci. USA 76:5217–5221; 1979.

    Article  PubMed  CAS  Google Scholar 

  • Gasparini, G.; Harris, A. L. Clinical importance of the determination of tumor angiogenesis in breast carcinoma: much more than a new prognostic tool. J. Clin. Oncol. 13:765–782; 1995.

    PubMed  CAS  Google Scholar 

  • Haraldsen, G.; Rugtveit, J.; Kvale, D.; Scholz, T.; Muller, W. A.; Hovig, T.; Brandtzaeg, P. Isolation and longterm culture of human intestinal microvascular endothelial cells. Gut 37:225–234; 1995.

    PubMed  CAS  Google Scholar 

  • Hewett, P. W.; Murray, J. C. Human lung microvessel endothelial cells: isolation, culture, and characterization. Microvasc. Res. 46:89–102; 1993a.

    Article  PubMed  CAS  Google Scholar 

  • Hewett, P. W.; Murray, J. C. Human microvessel endothelial cells: isolation, culture and characterization. In Vitro Cell. Dev. Biol. 29A:823–830; 1993b.

    Article  CAS  Google Scholar 

  • Hewett, P.; Murray, J.; Price, E.; Watts, M.; Woodcock, M. Isolation and characterization of microvessel endothelial cells from human mammary adipose tissue. In Vitro Cell. Dev. Biol. 29A:325–331; 1992.

    Google Scholar 

  • Hewett, P. W.; Murray, J. C.; Price, E. A.; Watts, M. E.; Woodcock, M. Isolation and characterization of microvessel endothelial cells from human mammary adipose tissue. In Vitro Cell. Dev. Biol. 29A:325–331; 1993.

    Article  CAS  Google Scholar 

  • Jackson, C. J.; Garbett, P. K.; Nissen, B.; Schrieber, L. Binding of human endothelium to Ulex europaeus 1-coated Dynabeads: application to the isolation of microvascular endothelium. J. Cell Sci. 96(Pt 2): 257–262; 1990.

    PubMed  Google Scholar 

  • Jackson, C. J.; Nguyen, M. Human microvascular endothelial cells differ from macrovascular endothelial cells in their expression of matrix metalloproteinases. Int. J. Biochem. Cell Biol. 29:1167–1177; 1997.

    Article  PubMed  CAS  Google Scholar 

  • Johnson, T. F.; Umbenhauer, D. R.; Hill, R., Bradt, C.; Mueller, S. N.; Levine, E. M.; Nichols, W. W. Karyotypic and phenotypic changes during in vitro aging of human endothelial cells. J. Cell Physiol. 150:17–27; 1992.

    Article  PubMed  CAS  Google Scholar 

  • Lammert, E.; Cleaver, O.; Melton, D. Induction of pancreatic differentiation by signals from blood vessels. Science 294:564–567; 2001.

    Article  PubMed  CAS  Google Scholar 

  • Lammert, E.; Cleaver, O.; Melton, D. Role of endothelial cells in early pancreas and liver development. Mech. Dev. 120:59–64; 2003.

    Article  PubMed  CAS  Google Scholar 

  • Lamszus, K.; Schmidt, N. O.; Ergun, S.; Westphal, M. Isolation and culture of human neuromicrovascular endothelial cells for the study of angiogenesis in vitro. J. Neurosci. Res. 55:370–381; 1999.

    Article  PubMed  CAS  Google Scholar 

  • Manconi, F.; Markham, R.; Fraser, I. S. Culturing endothelial cells of microvascular origin. Methods Cell Sci. 22:89–99; 2000.

    Article  PubMed  CAS  Google Scholar 

  • Matsumoto, K.; Yoshitomi, H.; Rossant, J.; Zaret, K. S. Liver organogenesis promoted by endothelial cells prior to vascular function. Science 294:559–563; 2001.

    Article  PubMed  CAS  Google Scholar 

  • McCarthy, S. A.; Kuzu, I.; Gatter, K. C.; Bicknell, R. Heterogeneity of the endothelial cell and its role in organ preference of tumour metastasis. Trends Pharmacol. Sci. 12:462–467; 1991.

    Article  PubMed  CAS  Google Scholar 

  • Mitelman, F. An international system for human cytogenetic nomenclature. ISCN, S. Karger, Basel, 1995.

    Google Scholar 

  • Naccarato, A. G.; Viacava, P.; Bocci, G.; Fanelli, G.; Aretini, P.; Lonobile, A.; Montruccoli, G.; Bevilacqua, G. Dehinition of the microvascular pattern of the normal human adult mammary gland. J. Anat. 203:599–603; 2003.

    Article  PubMed  Google Scholar 

  • Nichols, W. W.; Buynak, E. B.; Bradt, C.; Hill, R.; Aronson, M.; Jarrell, B. E.; Mueller, S. N.; Levine, E. M. Cyrogenetic evaluation of human endothelial cell cultures. J. Cell. Physiol. 132:453–462; 1987.

    Article  PubMed  CAS  Google Scholar 

  • O'Hare, M. J.; Bond, J.; Clarke, C., et al. Conditional immortalization of freshly isolated human mammary fibroblasts and endothelial cells. Proc. Natl. Acad. Sci. USA 98:646–651; 2001.

    Article  PubMed  Google Scholar 

  • Pasqualini, R.; Arap, W. Prohling the molecular diversity of blood vessels. Cold Spring Harbor Symp. Quant. Biol. 67:223–225; 2002.

    Article  PubMed  CAS  Google Scholar 

  • Pasqualini, R.; Arap, W.; McDonald, D. M. Probing the structural and molecular diversity of tumor vasculature. Trends Mol. Med. 8:563–71; 2002.

    Article  PubMed  CAS  Google Scholar 

  • Petersen, O. W.; van Deurs, B. Growth factor control of myoepithelial-cell differentiation in cultures of human mammary gland. Differentiation 39:197–215; 1988.

    Article  PubMed  CAS  Google Scholar 

  • Richard, L.; Velasco, P.; Detmar, M. A simple immunomagnetic protocol for the selective isolation and long-term culture of human dermal microvascular endothelial cells. Exp. Cell Res. 240:1–6; 1998.

    Article  PubMed  CAS  Google Scholar 

  • Rønnov-Jessen, L.; Petersen, O. W. Induction of alpha-smooth muscle actin by transforming growth factor-beta 1 in quiescent human breast gland fibroblasts. Implications for myofibroblast generation in breast neoplasia. Lab. Invest. 68:696–707; 1993.

    PubMed  Google Scholar 

  • Rønnov-Jessen, L.; Petersen, O. W.; Bissell, M. J., Cellular changes involved in conversion of normal to malignant breast: importance of the stromal reaction. Physiol. Rev. 76:69–125; 1996.

    PubMed  Google Scholar 

  • Shekhar, M. P.; Werdell, J.; Tait, L. Interaction with endothelial cells is a prerequisite for branching ductal-alveolar morphogenesis and hyperplasia of preneoplastic human breast epithelial cells regulation by estrogen. Cancer Res. 60:439–449; 2000.

    PubMed  CAS  Google Scholar 

  • Shen, Q.; Goderie, S. K.; Jin, L.; Karanth, N.; Sun, Y.; Abramova, N.; Vincent, P.; Pumiglia, K.; Temple, S. Endothelial cells stimulate self-renewal and expand neurogenesis of neural stem cells. Science 304:1338–1340; 2004.

    Article  PubMed  CAS  Google Scholar 

  • Wagner, E. F.; Risau, W. Oncogenes in the study of endothelial cell growth and differentiation. Semin. Cancer Biol. 5:137–145; 1994.

    PubMed  CAS  Google Scholar 

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Authors and Affiliations

  1. Faculty of Medicine, University of Iceland, Revkjavik, Iceland

    Valgardur Sigurdsson, Helga M. Ogmundsdottir & Thorarinn Gudjonsson

  2. Molecular and Cell Biology Research Laboratory, Icelandic Cancer Society, Revkjavik, Iceland

    Valgardur Sigurdsson, Agla J. B. Fridriksdottir, Helga M. Ogmundsdottir & Thorarinn Gudjonsson

  3. Structural Cell Biology Unit, Department of Medical Anatomy-A, The Panum Institute, University of Copenhagen, Denmark

    Agla J. B. Fridriksdottir & Ole William Petersen

  4. St. Josefs Hospital, Iceland

    Jens Kjartansson

  5. Department of Pathology, University Hospital of Iceland, Iceland

    John G. Jonasson

  6. Chromosome Laboratory, Department of Genetics and Molecular Medicine, University Hospital of Iceland, Iceland

    Margret Steinarsdottir

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  1. Valgardur Sigurdsson
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  2. Agla J. B. Fridriksdottir
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Sigurdsson, V., Fridriksdottir, A.J.B., Kjartansson, J. et al. Human breast microvascular endothelial cells retain phenotypic traits in long-term finite life span culture. In Vitro Cell.Dev.Biol.-Animal 42, 332–340 (2006). https://doi.org/10.1290/0602017.1

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  • DOI: https://doi.org/10.1290/0602017.1

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