Melanization of Bomirski hamster amelanotic melanoma cells (Ab line) depends on the type of culture medium
Abstract
Introduction. The effect of melanogenesis intensity on melanoma biology remains an open question, and the biological differences between melanotic and amelanotic melanoma cells have not yet been satisfactorily documented. As a result, the melanization of melanoma cells in in vitro cultures is not considered among experimental procedures. The aim of this study was to investigate the effect of the medium used to culture Bomirski amelanotic Ab melanoma cells on the melanogenesis process.
Material and methods. Amelanotic melanoma cells (Ab) were cultured in two media recommended for in vitro melanoma cell cultures, RPMI and DMEM. The melanization was evaluated by determining the melanin and tyrosinase presence in the cells using spectrophotometrical and western blot methods, respectively. Changes in Ab melanoma cells’ ultrastructure were determined using electron microscopy (EM).
Results. The medium with higher level of tyrosine (DMEM) induced significant melanization of amelanotic
melanoma cells (Ab) after only 24 h, while the RPMI medium, with a lower level of tyrosine, weakly affected melanin production. Melanization of Ab cells was paralleled by an increase in the amount of tyrosinase protein. Induced melanization was easily observed on EM-micrographs in the form of newly formed melanosomes containing melanin pigment. Melanosomes at stages from one (I) to four (IV) were observed.
Conclusions. Culture medium has an important effect on the in vitro biology of amelanotic melanoma cells, since it can affect the rate of cellular melanization. The appropriate medium should be carefully selected, taking into account the known biology of the melanoma cells being used.
Keywords: Bomirski melanomaamelanotic melanomamelanogenesisculture mediamelanintyrosinasewestern blottingelectron microscopy
References
- Koch SE, Lange JR. Amelanotic melanoma: the great masquerader. J Am Acad Dermatol. 2000; 42(5 Pt 1): 731–734.
- Thomas NE, Kricker A, Waxweiler WT, et al. Genes, Environment, and Melanoma (GEM) Study Group. Comparison of clinicopathologic features and survival of histopathologically amelanotic and pigmented melanomas: a population-based study. JAMA Dermatol. 2014; 150(12): 1306–1314.
- Giuliano AE, Cochran AJ, Morton DL. Melanoma from unknown primary site and amelanotic melanoma. Semin Oncol. 1982; 9(4): 442–447.
- Gualandri L, Betti R, Crosti C. Clinical features of 36 cases of amelanotic melanomas and considerations about the relationship between histologic subtypes and diagnostic delay. J Eur Acad Dermatol Venereol. 2009; 23(3): 283–287.
- Kycler W, Grodecka-Gazdecka S, Bręborowicz J, et al. Prognostic value of selected immunohistochemical markers in skin melanoma. Nowotwory. 2002; 52: 487–495.
- Perry MD, Seigler HF, Johnston WW. Diagnosis of metastatic malignant melanoma by fine needle aspiration biopsy: a clinical and pathologic correlation of 298 cases. J Natl Cancer Inst. 1986; 77(5): 1013–1021.
- Kapila K, Kharbanda K, Verma K. Cytomorphology of metastatic melanoma--use of S-100 protein in the diagnosis of amelanotic melanoma. Cytopathology. 1991; 2(5): 229–237.
- Cordoro KM, Gupta D, Frieden IJ, et al. Pediatric melanoma: results of a large cohort study and proposal for modified ABCD detection criteria for children. J Am Acad Dermatol. 2013; 68(6): 913–925.
- Cos S, Garcia-Bolado A, Sánchez-Barceló EJ. Direct antiproliferative effects of melatonin on two metastatic cell sublines of mouse melanoma (B16BL6 and PG19). Melanoma Res. 2001; 11(2): 197–201.
- Potti A, Moazzam N, Langness E, et al. Immunohistochemical determination of HER-2/neu, c-Kit (CD117), and vascular endothelial growth factor (VEGF) overexpression in malignant melanoma. J Cancer Res Clin Oncol. 2004; 130(2): 80–86.
- Vad NM, Yount G, Moridani MY. Biochemical mechanism of acetylsalicylic acid (Aspirin) selective toxicity toward melanoma cell lines. Melanoma Res. 2008; 18(6): 386–399.
- Bomirski A, Słominski A, Bigda J. The natural history of a family of transplantable melanomas in hamsters. Cancer Metastasis Rev. 1988; 7(2): 95–118.
- Cichorek M. Spontaneous and induced apoptosis of two hamster transplantable melanoma lines differing in their biological features. Ann Acad Med Gedan. 2006; 36(Sup.7): 1–149.
- Bomirski A. Biological properties of transplantable melanomas in the Syrian hamster during 16 years of maintenance by several passages. Habilitation thesis. Medical Academy of Gdansk. Gdansk, Poland. ; 1977.
- Cichorek M, Kozłowska K, Bryl E. Mitochondrial transmembrane potential in spontaneous and camptothecin-induced apoptosis of melanotic and amelanotic melanoma cells. Neoplasma. 2007; 54(1): 29–36.
- Cichorek M, Kozłowska K, Bryl E. The activity of caspases in spontaneous and camptothecin-induced death of melanotic and amelanotic melanoma cell. Cancer Biol Ther. 2007; 6(3): 346–353.
- Sidor-Kaczmarek J, Cichorek M, Spodnik JH, et al. Proteasome inhibitors against amelanotic melanoma. Cell Biol Toxicol. 2017; 33(6): 557–573.
- Dębowski D, Cichorek M, Lubos M, et al. Noncovalent inhibitors of human 20S and 26S proteasome based on trypsin inhibitor SFTI-1. Biopolymers. 2016; 106(5): 685–696.
- Halaban R, Cheng E, Zhang Y, et al. Aberrant retention of tyrosinase in the endoplasmic reticulum mediates accelerated degradation of the enzyme and contributes to the dedifferentiated phenotype of amelanotic melanoma cells. Proc Natl Acad Sci U S A. 1997; 94(12): 6210–6215.
- Słomiński A. Some properties of Bomirski Ab amelanotic melanoma cells, which underwent spontaneous melanization in primary cell culture. Growth kinetics, cell morphology, melanin content and tumorigenicity. J Cancer Res Clin Oncol. 1985; 109(1): 29–37.
- Słomiński A, Słomiński A. Rapid melanization of Bomirski amelanotic melanoma cells in cell culture. Biosci Rep. 1983; 3(2): 189–194.
- Prota G. The Chemistry of Melanins and Melanogenesis. Fortschritte der Chemie organischer Naturstoffe / Progress in the Chemistry of Organic Natural Products. 1995: 93–148.
- Siegrist W, Eberle AN. In situ melanin assay for MSH using mouse B16 melanoma cells in culture. Anal Biochem. 1986; 159(1): 191–197.
- Chen YM, Chavin W. Melanogenesis in human melanomas. Cancer Res. 1975; 35(3): 606–612.
- Prezioso JA, Wang N, Duty L, et al. Enhancement of pulmonary metastasis formation and gamma-glutamyltranspeptidase activity in B16 melanoma induced by differentiation in vitro. Clin Exp Metastasis. 1993; 11(3): 263–274.
- Ancans J, Thody AJ. Activation of melanogenesis by vacuolar type H(+)-ATPase inhibitors in amelanotic, tyrosinase positive human and mouse melanoma cells. FEBS Lett. 2000; 478(1-2): 57–60.
- Cohen-Solal KA, Crespo-Carbone SM, Namkoong J, et al. Progressive appearance of pigmentation in amelanotic melanoma lesions. Pigment Cell Res. 2002; 15(4): 282–289.
- Slominski A, Ermak G, Wortsman J. Modification of melanogenesis in cultured human melanoma cells. In Vitro Cell Dev Biol Anim. 1999; 35(10): 564–565.
- Slominski A, Paus R, Mihm MC. Inhibition of melanogenesis as an adjuvant strategy in the treatment of melanotic melanomas: selective review and hypothesis. Anticancer Res. 1998; 18(5B): 3709–3715.
- Brozyna AA, VanMiddlesworth L, Slominski AT. Inhibition of melanogenesis as a radiation sensitizer for melanoma therapy. Int J Cancer. 2008; 123(6): 1448–1456.
- Slominski A, Zbytek B, Slominski R. Inhibitors of melanogenesis increase toxicity of cyclophosphamide and lymphocytes against melanoma cells. Int J Cancer. 2009; 124(6): 1470–1477.
- Wasiewicz T, Szyszka P, Cichorek M, et al. Antitumor effects of vitamin D analogs on hamster and mouse melanoma cell lines in relation to melanin pigmentation. Int J Mol Sci. 2015; 16(4): 6645–6667.
- Słomiński A, Bomirski A, Scisłowski PW, et al. Tyrosinase activity in primary cell culture of amelanotic melanoma cells. Biosci Rep. 1983; 3(11): 1027–1034.
- Słominski A, Moellmann G, Kuklinska E, et al. Positive regulation of melanin pigmentation by two key substrates of the melanogenic pathway, L-tyrosine and L-dopa. J Cell Sci. 1988; 89 ( Pt 3): 287–296.
- Slominski A, Costantino R. L-tyrosine induces tyrosinase expression via a posttranscriptional mechanism. Experientia. 1991; 47(7): 721–724.
- Slominski A, Costantino R. Molecular mechanism of tyrosinase regulation by L-dopa in hamster melanoma cells. Life Sci. 1991; 48(21): 2075–2079.
- Slominski A, Zmijewski MA, Pawelek J. L-tyrosine and L-dihydroxyphenylalanine as hormone-like regulators of melanocyte functions. Pigment Cell Melanoma Res. 2012; 25(1): 14–27.
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