Abstract
Abstract. The development and characterization of normal hamster tracheal epithelial (HTE) cell system and its initiated subline is described in the present study. Normal HTE cells grew in a monolayer, had a stable diploid karyotype, were anchorage dependent and non‐tumorigenic. The presence of desmosomal attachments and keratin filaments confirmed the epithelial nature of these cells. An initiated subline DTC8 was isolated after treatment of HTE cells with a suboptimal dose of 9,10‐dimethyl‐1,2‐benz[a]anthracene (DMBA). These DTC8 cells grew in a monolayer, had a higher growth rate and saturation density, were weakly anchorage independent and non‐tumorigenic. Treatment of DTC8 cells with 100 ng 12‐O‐tetradecanoyl phorbol‐13‐acetate (TPA), resulted in transformation of these cells which then showed anchorage independent growth on semisolid agar and formed tumours in 85% animals. As DTC8 cells showed heterogeneity in chromosome number, they were further cloned by the limiting dilution method using gamma‐irradiated hamster embryonic fibroblasts as a feeder layer. The clone H71, isolated among these clones had all the properties of initiated cells.
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REFERENCES
- Barrett JC, Tenant RW, eds. (1985) Mammalian cell transformation: mechanism of carcinogenesis and assays for carcinogens. Carcino. Compr. Surv. 9, 90. [PubMed] [Google Scholar]
- Bertolero F, Kaighn ME, Gonda MA, Saffiotti V. (1984) Mouse epidermal keratinocytes clonal proliferation and response to hormones and growth factors in serum free medium. Exp. Cell. Res. 155, 64. [DOI] [PubMed] [Google Scholar]
- Boukamp P, Stanbridge EJ, Yin Foo D, Cerutti PA, Fusenig NE. (1990) c‐Ha‐ras oncogene expression in immortalized human keratinocytes (HaCaT) alters growth potential in vivo but lacks coorelation with malignancy. Cancer Res. 50, 2840. [PubMed] [Google Scholar]
- Breitreuntz D, Hornung J, Pohlmann J et al. (1986) Environmental induction of differentiation‐specific keratins in malignant mouse keratinocyte lines. Eur. J. Cell Biol. 42, 255. [PubMed] [Google Scholar]
- Brinkley BR, Murphy P, Richardson LCJ. (1967) Procedure for embedding in situ selected cells cultured in vitro. J. Cell Biol. 35, 279. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dean PN. (1985) Methods of data analysis in flow cytometry In: Vandilla MA, Dean PN, Laerum OD, Melamed MR, eds. Flow Cytometry, Mechanisms of Diseases. New York : Academic Press, 195. [Google Scholar]
- Dipaolo JA (1983) Relative difficulties in transforming human and animals cells in vitro. J. Natl. Cancer Inst. 70, 3. [PubMed] [Google Scholar]
- Eichner R, Bonitz P, Sun TT. (1984) Classification of epidermal keratin according to their immunoreactivity isoelectrical point and mode of expression. J. Cell Biol. 98, 1388. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Fusenig NE. (1986) Mammalian epidermal cells in culture In: Biology of the Integument, Vol. 2; Vertebrates. Bereiter‐Hahn J, Matoltsy AG, Richards KS, eds. Berlin/Heidelberg : Springer Verlag, 409–442. [Google Scholar]
- Hennings H, Robinson VA, Michael DM, Petit GR, Jung R, Yuspa SH. (1990) Development of an in vitro analogue of initiated mouse epidermis to study tumor promoters and antipromoters. Cancer Res. 50, 4794. [PubMed] [Google Scholar]
- Hennings H, Lowry TD, Robinson VA et al. (1992) Activity of diverse tumor promoters in a keratinocyte co‐culture model of initiated epidermis. Carcinogenesis 13, 2145. [DOI] [PubMed] [Google Scholar]
- Jetten AM, Geoge MA, Smiths HL, Vollberg TM. (1989) Keratin 13 expression is linked to squamous differentiation in rabbit tracheal epithelial cells and down regulated by retinoic acid. Exp. Cell. Res. 182, 622. [DOI] [PubMed] [Google Scholar]
- Katimura H, Fitzgerald DJ, Li LA, Barrett JC, Nettesheim P. (1986) Morphological characterization of transformed colonies of rat tracheal epithelial cells cultures exposed to carcinogen. Cancer Res. 46, 4631. [PubMed] [Google Scholar]
- Laemmli UK. (1970) Clevage of structural proteins during the assembly of the head bacteriophage T4. Nature (London) 227, 680. [DOI] [PubMed] [Google Scholar]
- Macpherson I. (1973) Soft agar techniques In: Kruse PF, Patterson MK, eds. Tissue Culture. New York : Academic Press, 276. [Google Scholar]
- Masui T. (1992) Cell culture and its application. Culture system of normal human epithelial cells and their potential applications in cancer research. Gan To Kagaku Ryoho 19, 921. [PubMed] [Google Scholar]
- Mondal S, Brankow DW, Heidelberger C. (1976) Two stage chemical oncogenesis in culture of C3H/10T1/2 cells. Cancer Res. 36, 2254. [PubMed] [Google Scholar]
- Nakazawa H, Aguelon AM, Yamasaki H. (1992) Identification and quantification of carcinogen‐induced molecular initiation event in cell transformation. Oncogene 7, 2295. [PubMed] [Google Scholar]
- Namba M, Kimoto T. (1979) An attempt for improving plating efficiency of human diploid fibroblasts. Gann 70, 21. [PubMed] [Google Scholar]
- Nettesheim P, Marchok AC. (1983) Neoplastic development in air way epithelium. Adv. Cancer Res. 39, 1. [DOI] [PubMed] [Google Scholar]
- Nettesheim P, Terraghi M, Klein Szanto APJ. (1982) In: Harris CC, ed. Mechanisms of Chemical Carcinogenesis. New York : Alan R Liss Inc., 473–489. [Google Scholar]
- O'Farrell PH. (1975) High resolution two dimensional electrophoresis for protein. J. Biol. Chem. 250, 4007. [PMC free article] [PubMed] [Google Scholar]
- Pai SB, Steel VE, Nettesheim P. (1983) Neoplastic transformation of primary tracheal epithelial cell cultures. Carcinogenesis, 4, 369. [DOI] [PubMed] [Google Scholar]
- Reddel RR, Ke Y, Gerwin BI et al. (1988) Transformation of human bronchial epithelial cells by infection with SV‐40 or adenovirus‐12‐SV40 hybrid virus, or transfection via strontiumphosphate coprecipitation with a plasmid containing SV40 early region genes. Cancer Res. 48, 1904. [PubMed] [Google Scholar]
- Reddel RR, DE Silva R, Duncan EL et al. (1995) SV 40‐induced immortalization and ras‐transformation of human bronchial epithelial cells. Int. J. Cancer 61, 199. [DOI] [PubMed] [Google Scholar]
- Rheinwald JG, Green H. (1977). Epidermal growth factor and the multiplication of culture human epidermal keratinocytes. Nature 265, 421. [DOI] [PubMed] [Google Scholar]
- Rhim JS, Jay G, Arnstein P, Price FM, Sanford KK, Aaronson SA. (1985) Neoplastic transformation of human epidermal keratinocytes by Ad12‐SV40 and Kirsten sarcoma viruses. Science 227, 1250. [DOI] [PubMed] [Google Scholar]
- Sanford KK, Evans VJ. (1982) A quest for the mechanism of ‘Spontaneous’ malignant transformation in culture with associated advances in culture technology. J. Natl. Cancer Inst. 68, 895. [PubMed] [Google Scholar]
- Sanford KK, Earle WR, Evan VJ, Waltz HK, Shannon JE. (1951) Measurement of proliferation in tissue culture by enumeration of cell nuclei. J. Natl. Cancer Inst. 11, 147. [PubMed] [Google Scholar]
- Sun NC, Chu EHY, Chang CC. (1973) Staining method for the banding pattern of tumor mitotic chromosomes. Mam. Chr. New. Lett. 14, 26. [Google Scholar]
- Thomassen DG, Saffiotti U, Kaighn ME. (1986). Clonal proliferation of rat tracheal epithelial cells in serum free medium and their responses to hormones, growth factors and carcinogens. Carcinogenesis 7, 2033. [DOI] [PubMed] [Google Scholar]
- Towbin H, Staehelin T, Gordon J. (1979) Electrophoretic transfer of protein from polyacrylamide gel to nitrocellulose sheets. Procedure and some application. Proc. Natl. Acad. Sci. USA 76, 4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tsuchiya T, Umeda M. (1997) Relationship between exposure to TPA and appearance of transformed cells in MNNG‐initiated transformation of BALB/c 3T3 cells. Int. J. Cancer 73, 271. [DOI] [PubMed] [Google Scholar]
- Vaidya MM, Borges AM, Pradhan SA, Rajpal RM, Bhisey AN. (1989) Altered keratin expression in buccal mucosal squamous cell carcinoma. J. Oral Path. Med. 18, 282. [DOI] [PubMed] [Google Scholar]
- Walker C, Nettesheim P. (1989) In vitro neoplastic progression of rat tracheal epithelial cells transformation by diverse carcinogens. Cancer Res. 49, 4427. [PubMed] [Google Scholar]
- Wasilenko W, Marchok AC. (1986) Utilization of pyruvate and pyruvate precursors by normal and carcinogen altered rat tracheal epithelial cells in culture. J. Cell Physiol. 126, 69. [DOI] [PubMed] [Google Scholar]
- Wasilenko WJ, Marchok AC. (1984) Pyruvate regulation of growth and differentiation in primary cultures of rat tracheal epithelial cells. Exp. Cell. Res. 155, 507. [DOI] [PubMed] [Google Scholar]
- Weinberg RA. (1989) Oncogenes, antioncogenes and molecular bases of multistep carcinogenesis. Cancer Res. 49, 3713. [PubMed] [Google Scholar]
- Yunis JJ. (1981) New chromosome technique in the study of human neoplasm. Human Pathol. 12, 540. [DOI] [PubMed] [Google Scholar]
- Yuspa SH, Poirier MC. (1988) Chemical carcinogenesis: from animal models to molecular models in one decade. Adv. Cancer Res. 50, 25. [DOI] [PubMed] [Google Scholar]
