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Methemoglobin is a supplement for in vitro culture of human nasopharyngeal epithelial cells transformed by human Papillomavirus type 16 DNA

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Summary

NPC-N cells were normal human nasopharyngeal epithelial cells transformed by transfection with human papillomavirus type 16 deoxyribonucleic acid. Bovine pituitary extract (BPE) was one of the indispensable ingredients for in vitro culture of NPC-N cells in a serum-free medium. Chromatographic fractionation of BPE and subsequent immunoblotting analyses identified the hemoglobin growth-stimulating factor. Methemoglobin (metHb) was then synthesized, and also found to be growth stimulating. The growth-stimulating effect of metHb was abolished when NPC-N cells were cultured in a medium that also contained haptoglobin, a molecule that binds to hemoglobin. A defined medium consisting of insulin and metHb was then developed for optimal growth of NPC-N cells. MetHb kept under the conditions identical to those of cell culture released hemin which also enhanced the cell growth. Though all the degradation products of hemin are currently known to be physiologically significant, only ferric iron derived from metHb or hemin could stimulate the growth of NPC-N cells. Abnormal vasculature showing leaky walls and hemorrhage is a common feature of malignant tumors. Hemoglobin orginating from extravasated red blood cells and subsequently oxidized to metHb because of the presence of activated inflammatory cells might contribute to the increased proliferation of cancerous cells.

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References

  • Ascoli, F.; Fanelli, M. R. R.; Antonini, E. Preparation and properties of apohemoglobin and reconstituted hemoglobins. In: Antonini, E.; Rossi-Bernardi, L.; Chiancone, E., ed. Methods in enzymology. Vol. 76. New York: Academic Press; 1981: 72–85.

    Google Scholar 

  • Balla, J.; Jacob, H. S.; Balla, G., et al. Endothelial-cell heme uptake from heme proteins: induction of sensitization and desensitization to oxidant damage. Proc. Natl. Acad. Sci USA 90: 9285–9289; 1993.

    Article  PubMed  CAS  Google Scholar 

  • Beckmann, A. M.; Daling, J. R.; Sherman, K. J., et al. Human papilloma virus infection and anal cancer. Int. J. Cancer 43: 1042–1049; 1989.

    Article  PubMed  CAS  Google Scholar 

  • Boshart, M.; Gissmann, L.; Ikanberg, H., et al. A new type of papillomavirus DNA, its presence in genital cancer biopsies and in cell lines derived from cervical cancer. EMBO J. 3:1151–1157; 1984.

    PubMed  CAS  Google Scholar 

  • Bradford, M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248–254; 1976.

    Article  PubMed  CAS  Google Scholar 

  • Brandsma, J. L.; Abramson, A. L. Association of papillomavirus with cancers of the head and neck. Arch. Otolaryngol. Head Neck Surg. 115: 621–625; 1989.

    PubMed  CAS  Google Scholar 

  • Brandsma, J. L.; Steinberg, B. M.; Abramson, A. L., et al. Presence of human papillomavirus type 16 related sequences in verrucous carcinoma of the larynx. Cancer Res. 46: 2185–2188; 1986.

    PubMed  CAS  Google Scholar 

  • Dallegri, F.; Ballestrero, A.; Frumento, G., et al. Augmentation of neutrophilmediated erythrocyte lysis by cells derived in vitro from human monocytes. Blood 70: 1743–1749; 1987.

    PubMed  CAS  Google Scholar 

  • Debiec-Rychter, M.; Zukowski, K.; Wang, C. Y., et al. Chromosomal characterizations of human nasal and nasopharyngeal cells immortalized by human papillomavirus type 16 DNA. Cancer Genet. Cytogenet. 52: 51–61; 1991.

    Article  PubMed  CAS  Google Scholar 

  • Deeshirst, M. W.; Tso, C. Y.; Secomb, T. W., et al. Morphologic and hemodynamic comparison of tumor and healing normal tissue microvasculature. Int. J. Radiat. Oncol. Biol. Phys. 17: 91–99; 1989.

    Google Scholar 

  • deVilliers, E. M.; Weidauer, H.; Otto, H., et al. Papillomavirus DNA in human togue carcinomas. Int. J. Cancer 36: 575–578; 1985.

    Article  CAS  Google Scholar 

  • Durst, M.; Gissmann, L.; Ikenberg, H., et al. A papillomavirus DNA from a cervical carcinoma and its prevalence in cancer biopsy samples from different geographic regions. Proc. Natl. Acad. Sci. USA 80: 3812–3815; 1983.

    Article  PubMed  CAS  Google Scholar 

  • Garthwaite, J.; Southam, E.; Boulton, C.L., et al. Potent and selective inhibition of nitric oxide-sensitive guanylyl cyclase by 1H-[1,2,4]oxadiazolo[4,3-α]quinoxalin-1-one. Mol. Pharmacol. 48: 184–188; 1995.

    PubMed  CAS  Google Scholar 

  • Glamsta, E. L.; Marklund, A.; Hellman, U., et al. Isolation and characterization of a hemoglobin-derived opioid peptide from the human pituitary gland. Regul. Pept. 34: 169–179; 1991.

    Article  PubMed  CAS  Google Scholar 

  • Glamsta, E. L.; Meyerson, B.; Silberring, J., et al. Isolation of a hemoglobinderived opioid peptide from cerebrospinal fluid of patients with cerebrovascular bleedings. Biochem. Biophys. Res. Commun. 184:1060–1066; 1992.

    Article  PubMed  CAS  Google Scholar 

  • Grunt, T. W.; Lametschwandtner, A.; Staindl, O. The vascular pattern of basal cell tumors: light microscopy and scanning electron microscopic study on vascular corrosion casts. Microvasc. Res. 29: 371–386; 1985.

    Article  PubMed  CAS  Google Scholar 

  • Hadden, J. W.; Galy, A.; Chen, H., et al. A pituitary factor induces thymic epithelial cell proliferation in vitro. Brain Behav. Immun. 3: 149–159; 1989.

    Article  PubMed  CAS  Google Scholar 

  • Halliwell, B.; Gutteridge, J. M. C. Role of free radicals and catalytic metal ions in human disease: an overview. In: Packer, L.; Glazer, A. N., ed. Methods in enzymology. Vol. 186. New York: Academic Press; 1990; 1–85.

    Google Scholar 

  • Hwang, P. K.; Greer, J. Interaction between hemoglobin subunits in the hemoglobin-haptoglobin complex. J. Biol. Chem. 255: 3038–3041; 1980.

    PubMed  CAS  Google Scholar 

  • Ignarro, L. J.; Wood, K. S.; Wolin, M. S. Aotivation of purified soluble guanylate cyclase by protoprohyrin IX. Proc. Natl. Acad. Sci. USA 79: 2870–2873; 1982.

    Article  PubMed  CAS  Google Scholar 

  • Ikenberg, H.; Gissmann, L.; Gross, G., et al. Papillomavirus type 16-related DNA in genital Bowen's disease and Bowenoid papulosis. Int. J. Cancer 32: 563–565; 1983.

    Article  PubMed  CAS  Google Scholar 

  • Ingi, T.; Cheng, J.; Ronnett, G. V. Carbon monoxide: an endogenous modulator of the nitric oxide-cyclic GMP signaling system. Neuron 16:835–842; 1996.

    Article  PubMed  CAS  Google Scholar 

  • Jea, J.-C.; Liu, T.-C.; Wang, S.-Y., et al. Nitric oxide enhances the growth of U937 human leukemic cells through a cyclooxygenase-mediated pathway. J. Leukoc. Biol. 64: 451–458: 1998.

    PubMed  CAS  Google Scholar 

  • Jensenius, J. C.; Andersen, I.; Hau, J., et al. Eggs: conveniently packaged antibodies. Methods for purification of yolk IgG. J. Immunol. Methods 46: 63–68; 1981.

    Article  PubMed  CAS  Google Scholar 

  • Kokko, H.; Kruonen, I.; Karenlampi, S., Rapid production of antibodies in chicken and isolation from eggs. In: Celis, J. E., ed. Cell biology, a laboratory handbook. Vol. 2. New York: Academic Press; 1994; 282–288.

    Google Scholar 

  • Kuroiwa, T.; Moriwaki, K.; Ohta, T., et al. Extradural extension of primitive neuroectodermal tumor-case report. Neurol. Med. Chir. 34:379–381; 1994.

    CAS  Google Scholar 

  • Maines, M. D. The heme oxygenase system: a regulator of second messenger gases. Annu. Rev. Pharmacol. Toxicol. 37: 517–554; 1997.

    Article  PubMed  CAS  Google Scholar 

  • Mancuso, C.; Pistritto, G.; Tringali, G., et al. Evidence that carbon monoxide stimulates prostaglandin endoperoxide synthase activity in rat hypothalamic explants and in primary cultures of rat hypothalamic astrocytes. Mol. Brain Res. 45: 294–300; 1997.

    Article  PubMed  CAS  Google Scholar 

  • Mancuso, C.; Tringali, G.; Grossman, A., et al. The generation of nitric oxide and carbon monoxide produces opposite effects on the release of immunoreactive interleukin-1β from the rat hypothalamus in vitro: evidence for the involvement of different signaling pathways. Endocrinology 139: 1031–1037; 1998.

    Article  PubMed  CAS  Google Scholar 

  • McCance, D. J.; Kopan, R.; Fuchs, E., et al. Human papillomavirus type 16 alters human epithelial cell differentiation in vitro. Proc. Natl. Acad. Sci. USA 85: 7169–7173; 1988.

    Article  PubMed  CAS  Google Scholar 

  • Parisi, L.; Creek, K. E.; Doniger, J., et al. Continuous cell lines with altered growth and differentiation properties originate after transfection of human keratinocytes with human papillomavirus type 16 DNA. Carcinogenesis 9: 1573–1579; 1988.

    Article  Google Scholar 

  • Pakowski, A.; Sikorski, A. F. Effect of hemin on growth and DNA synthesis of HL-60 cells. In Vitro Cell. Dev. Biol. 29A: 679–682; 1993.

    Google Scholar 

  • Pauling, L.; Coryell, C. D., et al. The magnetic properties and structure of hemoglobin, oxyhemoglobin and carbonmonosyhemoglobin. Proc. Natl. Acad. Sci. USA 22: 210–216, 1936.

    Article  PubMed  CAS  Google Scholar 

  • Pittelkow, M. R.; Shipley, G. D. Serum-free culture of normal melanocytes: growth kinetics and growth factor requirements. J. cell. Physiol. 140: 565–576; 1989.

    Article  PubMed  CAS  Google Scholar 

  • Ramos, K. S.; Lin, H.; McCrath, J. J., Modulation of cyclic guanosine monophosphate levels in cultured aortic smooth muscle cells by carbon monoxide. Biochem. Pharmacol. 38: 1368–1370; 1989.

    Article  PubMed  CAS  Google Scholar 

  • Savci, G.; Kilicturgay, S.; Sivri, Z., et al. Solid and papillary epithelial neoplasm of the pancreas: CT and MR findings. Eur. Radiol. 6: 86–88; 1996.

    Article  PubMed  CAS  Google Scholar 

  • Shah-Yukich, A. A.; Nelson, A. C., Charaterization of solid tumor microvasculature: a three-dimensional analysis using the polymer casting technique. Lab. Invest. 58: 236–244; 1988.

    PubMed  CAS  Google Scholar 

  • Spoto, G. P.; Press, G. A.; Hesselink, J. R., et al. Intracranial apendymone and subependymoma: MR manifestations. Am. J. Neuroad. 11: 83–91; 1990.

    CAS  Google Scholar 

  • Stenzel, K. H.; Rubin, A. L.; Novogrodsky, A. Mitogenic and co-mitogenic properties of hemin. J. Immunol. 127: 2469–2473; 1981.

    PubMed  CAS  Google Scholar 

  • Stocker, R.; Yamamoto, Y; McDonagh, A. F., et al. Bilirubin is an antioxidant of possible physiological importance. Science 235: 1043–1047; 1987.

    Article  PubMed  CAS  Google Scholar 

  • Sussman, H. H. Iron and cancer. Pathobiology 60: 2–9; 1992.

    Article  PubMed  CAS  Google Scholar 

  • Syrjanen, S. M.; Happonen, R. P.; Virolainen, E., et al. Detection of human papillomavirus (HPV) structural antigens and DNA types in inverted papillomas and squamous cell carcinomas of the nasal cavities and paranasal innuses. Acta Otolaryngol. 104: 334–341; 1987.

    PubMed  CAS  Google Scholar 

  • Syrjanen, S. M.; Syrjanen, K. J.; Lamberg M. A. Detection of human papillomavirus DNA in oral mucosal lesions using in situ DNA hybridization applied on paraffin sections. Oral Pathol. 62: 660–667; 1986.

    Article  CAS  Google Scholar 

  • Utz, J.; Ullrich, V. Carbon monoxide relaxes ilial smooth muscle through activation of guanylate cyclase. Biochem. Pharmacol. 41: 1195–2001; 1991.

    Article  PubMed  CAS  Google Scholar 

  • Valette, I.; Waks, M.; Wejman, J. C., et al. Haptoglobin heavy and light chains. Structural properties, reassembly, and formation of minicomplex with hemoglobin. J. Biol. Chem. 256: 672–679; 1981.

    PubMed  CAS  Google Scholar 

  • Vercellotti, G. M.; van Asbeck, B. S.; Jacob, H. S. Oxygen radical-induced erythrocyte hemolysis by neutrophils: critical role of iron and lactoferrin. J. Clin. Invest. 76: 956–962; 1985.

    Article  PubMed  CAS  Google Scholar 

  • Verger, C.; Sassa, S.; Kappas, A. Growth-promoting effects of iron-and cobaltprotoporphyrins on cultured embryonic cells. J Cell. Physiol 116: 135–141; 1983.

    Article  PubMed  CAS  Google Scholar 

  • Waldman, S. A.; Sinacore, M. S.; Lweicki, J. A., et al. Selective activation of particulate guanylate cyclase by a specific class of porphyrins. J. Biol. Chem. 259: 4038–4042; 1984.

    PubMed  CAS  Google Scholar 

  • Weinberg, E. D. Iron and neoplasia. Biol. Trace Elem. Res. 3: 55–80: 1981.

    CAS  Google Scholar 

  • Weinberg, E. D. Roles of iron in neoplasia, promotion, prevention and therapy. Biol. Trace Elem. Res. 34: 923–940; 1992.

    Google Scholar 

  • Weiss, S. J. Neutrophil-mediated methemoglobin formation in the erythrocyte The role of superoxide and hydrogen peroxide. J. Biol. Chem. 257: 2947–2953; 1982.

    PubMed  CAS  Google Scholar 

  • Wen, W.; Liu, C.; Chang, M., et al. Continual growth of two newly established human hepatoma cell lines in basal media supplemnted with no further exogenous growth ingredients. In: Murakami, H., ed. Trends in animal cell culture technology. Tokyo: Kondasha; 1990: 75–80.

    Google Scholar 

  • Wille Jr., J. J.; Pittelkow, M. R.; Shipley, G. D., et al. Integrated control of growth and differentiation of normal human prokeratinocytes cultured fin serum-free medium: clonal analyses, growth kinetics, and cell cycle studies. J Cell. Physiol. 121: 31–44: 1984.

    Article  PubMed  CAS  Google Scholar 

  • Willey, J. C.; Laveck, M. A.; Mcclendon, I. A., et al. Relationship of ornithine decarbosylase activity and cAMP metabolism to proliferation of normal human bronchial epithelial cells. J. Cell. Physiol. 124:207–212; 1985.

    Article  PubMed  CAS  Google Scholar 

  • Wolin, M. S.; Wood, K. S.; Ignarro, L. J. Guanylate cyclase from bovine lung. A kinetic analysis of the regulation of the purified soluble enzyme by protoporphyrin IX, heme, and nitrosyl-heme. J. Biol. Chem. 257:13,312–13,320; 1982.

    CAS  Google Scholar 

  • Woodworth, C. D.; Bowden, P. E.; Doniger, J., et al. Characterization of normal human exocervical epithelial cells immortalized in vitro by papillomavirus types 16 and 16 DNA. Cancer Res. 48:4620–4628; 1988.

    PubMed  CAS  Google Scholar 

  • Zimmerman, R. A; Bilaniuk, L. T.; Hackney, D. B., et al. Magnetic resonance imaging of dural venous sinus invasion, occlusion and thrombosis. Acta Radiol. Suppl. 369:110–112; 1986.

    PubMed  CAS  Google Scholar 

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  1. Institute of Biochemistry, College of Medicine, National Taiwan University, 100, Taipei, Taiwan

    Wu-Nan Wen

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Wen, WN. Methemoglobin is a supplement for in vitro culture of human nasopharyngeal epithelial cells transformed by human Papillomavirus type 16 DNA. In Vitro Cell.Dev.Biol.-Animal 37, 668–675 (2001). https://doi.org/10.1290/1071-2690(2001)037<0668:MIASFI>2.0.CO;2

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  • DOI: https://doi.org/10.1290/1071-2690(2001)037<0668:MIASFI>2.0.CO;2

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