Skip to main content
PMC home page
Japanese Journal of Cancer Research : Gann logoLink to Japanese Journal of Cancer Research : Gann
. 1998 Sep;89(9):923–932. doi: 10.1111/j.1349-7006.1998.tb00650.x

Selection of Human Ovarian Carcinoma Cells with High Dissemination Potential by Repeated Passage of the Cells in vivo into Nude Mice, and Involvement of Lex‐determinant in the Dissemination Potential

Kazushige Kiguchi 1, Masao Iwamori 2, Yukari Mochizuki 3, Takeshi Kishikawa 3, Katsumi Tsukazaki 3, Masahiko Saga 1, Akira Amemiya 4, Shiro Nozawa 3
PMCID: PMC5921947  PMID: 9818028

Abstract

Cells of the human tumor cell line RMG‐1, derived from a clear‐cell adenocarcinoma of the ovary, were injected intraperitoneally into nude mice, and the cells obtained from the tumor nodules in the mesenterium were found to form a larger number of, and larger‐sized, tumor nodules than the original RMG‐1 cells. The RMG‐1‐h cells, transferred into culture from the tumor nodules after a 4th in vivo passage, showed a dissemination potential as high as that of cells disseminating directly from the tissues, and exceedingly higher than that of RMG‐1 cells. To assess the molecular bases of the different biological properties of RMG‐1 and RMG‐1‐h cells, we compared the content and expression of various carbohydrate antigens in both cells. The chromosomal profile of RMG‐1‐h cells revealed their human origin and was identical to that of the original RMG‐1 cells. In contrast to the broad histogram for the Lex‐bearing cells among RMG‐1 cells in flow cytometry, the weakly and moderately positive cells toward anti‐Lex antibody were found to be eliminated from the histogram for the RMG‐1‐h cells, resulting in the enrichment of cells strongly expressing Lex, which may account for the high dissemination potential. In addition, the adhesion of RMG‐1 cells to mesothelial cells was found to be significantly inhibited by pretreatment of the cells with anti‐Lex antibody, indicating Lex‐mediated cell‐to‐cell interaction between ovarian cancer cells and mesothelial cells. By TLC‐immunostaining, two Lex‐glycolipids, III3Fucα‐nLc4Cer and V3Fucα‐nLc6Cer were detected in both RMG‐1 and RMG‐1‐h cells, and their total concentrations were not significantly different from each other. However, the hydrophobic moieties of Lex‐glycolipids in RMG‐1‐h cells were different from those in RMG‐1 cells, suggesting that a difference in the structure of the hydrophobic moieties of Lex is partly involved in the enhanced reactivity of RMG‐1‐h cells toward anti‐Lex antibody. Thus, the high dissemination potential of ovarian cancer cells was shown to be mediated by the Lex‐determinant and the Lex‐bearing cells are enriched by repeated in vivo passage of the cells into nude mice.

Keywords: key words, Glycolipid, TLC‐immunostaining, Cell adhesion, Flow cytometry

Full Text

The Full Text of this article is available as a PDF (1.0 MB).

Abbreviations used are: GSL, glycosphingolipid; FCS, fetal calf serum; TLC, thin‐layer chromatography; PBS, phosphate‐buffered saline; PVP, polyvinylpyrrolidone; BSA, bovine serum albumin; Hex, hexose; HexNAc, N‐acetylhexosamine; IL‐1, interleukin‐1.

REFERENCES

  • 1. ) Yogeeswaran , G. and Salk , P. L.Metastatic potential is positively correlated with cell surface sialylation of cultured murine tumor cell lines . Science , 212 , 1514 – 1516 ( 1981. ). [DOI] [PubMed] [Google Scholar]
  • 2. ) Dennis , J. W. C. , Timpl , R. and Schirrmmacher , V.Surface sialic acid reduces attachment of metastatic tumor cells to collagen type IV and fibronectin . Nature , 300 , 274 – 276 ( 1982. ). [DOI] [PubMed] [Google Scholar]
  • 3. ) Kiguchi , K. , Nozawa , S. , Iwamori , M. and Nagai , Y.Glycosphingolipids of various human ovarian tumors: a significantly high expression of I3SO3GalCer and Lewis antigen in mucinous cystoadenoma . Cancer Res. , 52 , 416 – 421 ( 1992. ). [PubMed] [Google Scholar]
  • 4. ) Hakomori , S.Glycosphingolipids as differentiation dependent tumor associated markers and as regulators of cell proliferation . Trends Biol. Sci. , 9 , 453 – 458 ( 1984. ). [Google Scholar]
  • 5. ) Kishikawa , T. , Sakamoto , M. , Ino , Y. , Kubushiro , K. , Nozawa , S. and Hirohashi , S.Two distinct patterns of peritoneal involvement shown by in vitro and in vivo ovarian cancer dissemination models . Invasion Metastasis , 15 , 11 – 21 ( 1995. ). [PubMed] [Google Scholar]
  • 6. ) Nozawa , S. , Tsukazaki , K. , Sakayori , M. , Jeng , C. H. and Iizuka , R.Establishment of a human ovarian clear cell carcinoma cell line (RMG‐1) and its single cell cloning — with special reference to the stem cell of the tumor . Hum. Cell , 1 , 426 – 434 ( 1988. ). [PubMed] [Google Scholar]
  • 7. ) Udagawa , Y. , Nozawa , S. , Chin , K. , Sakayori , M. , Mikami , M. , Ohta , K. , Tsukazaki , K. , Kiguchi , K. and Iizuka , R.Establishment and characterization of a human chorionic gonadotropin (hCG) producing cell line (RTSG) from an ovarian epithelial cancer . Hum. Cell , 3 , 70 – 75 ( 1990. ). [PubMed] [Google Scholar]
  • 8. ) Iwamori , M. , Sawada , K. , Hara , Y. , Nishino , M. , Fujikawa , T. , Imura , H. and Nagai , Y.Neutral glycosphingolipids and gangliosides of bovine thyroid . J. Biochem. , 91 , 1875 – 1887 ( 1982. ). [PubMed] [Google Scholar]
  • 9. ) Bartlett , G. R.Phosphorus assay in column chromatography . J. Biol. Chem. , 234 , 466 – 468 ( 1959. ). [PubMed] [Google Scholar]
  • 10. ) Saito , T. and Hakomori , S.Quantitative isolation of total glycosphingolipids from animal cells . J. Lipid Res. , 12 , 257 – 259 ( 1971. ). [PubMed] [Google Scholar]
  • 11. ) Miettinen , T. and Takki‐luukainen , J. T.Use of butyl acetate in determination of sialic acid . Acta Chem. Scand. , 13 , 856 – 858 ( 1959. ). [Google Scholar]
  • 12. ) Iwamori , M. , Sunada , S. , Ishihara , E. , Moki , E. , Fujimoto , S. and Nagai , Y.Differential expression of fucosyl GM1 and disialoganglioside with a NeuAcα2‐6Ga1NAc linkage (GD1e) in various rat ascites hepatoma cells . FEBS Lett. , 198 , 66 – 70 ( 1986. ). [DOI] [PubMed] [Google Scholar]
  • 13. ) Watanabe , M. , Hirohashi , S. , Shimosato , Y. , Ino , Y. , Yamabe , T. , Teshima , S. , Sekine , T. and Abe , O.Carbohydrate antigen defined by monoclonal antibody raised against a gastric cancer xenograft . Jpn. J. Cancer Res. , 76 , 43 – 52 ( 1985. ). [PubMed] [Google Scholar]
  • 14. ) Hirohashi , S. , Shimosato , Y. , Ino , Y. , Tome , Y. , Watanabe , M. , Hirota , T. and Itabashi , M.Distribution of blood group antigens and CA 19‐9 in gastric cancers and non‐neoplastic gastric mucosa . Jpn. J. Cancer Res. , 75 , 540 – 547 ( 1984. ). [PubMed] [Google Scholar]
  • 15. ) Arita , M. , Iwamori , M. , Higuchi , T. and Nagai , Y.Negative ion fast atom bombardment mass spectrometry of gangliosides and asialo‐gangliosides: a useful method for the structural elucidation of gangliosides and related neutral glycosphingolipids . J. Biochem. , 94 , 249 – 256 ( 1983. ). [DOI] [PubMed] [Google Scholar]
  • 16. ) Dennis , J. W. and Laferte , S.Tumor cell surface carbohydrate and the metastatic phenotype . Cancer Metastasis Rev. , 5 , 185 – 204 ( 1987. ). [DOI] [PubMed] [Google Scholar]
  • 17. ) Irimura , T. and Reading , C. L.Surface properties of metastatic tumor cells . Cancer Bull. , 39 , 132 – 141 ( 1987. ). [Google Scholar]
  • 18. ) Irimura , T. , Gonzales , R. and Nicolson , G. L.Effects of tunicamycin on B16 metastatic melanoma cell surface glycoproteins and blood borne arrest and survival properties . Cancer Res. , 41 , 3411 – 3418 ( 1981. ). [PubMed] [Google Scholar]
  • 19. ) Dennis , J. W. and Laffrte , S.Recognition of asparagine‐linked oligosaccharides on murine tumor cells by natural killer cells . Cancer Res. , 45 , 6034 – 6040 ( 1985. ). [PubMed] [Google Scholar]
  • 20. ) Altevogt , P. , Fogel , M. , Chein Song‐popov , R. , Dennis , J. and Schirrmmacher , V.Related high and low metastatic tumor lines show different patterns of lectin binding and cell surface sialylation . Cancer Res. , 43 , 5138 – 5144 ( 1983. ). [PubMed] [Google Scholar]
  • 21. ) Codington , J. F. , Klein , G. , Lee , N. , Brown , R. W. and Jeanloz , R. W.Further studies on the relationship between large glycoprotein molecules and allotransplantability in the TA3 tumor of the mouse: studies on segregating TA‐3 Ha hybrids . J. Natl. Cancer Inst. , 60 , 811 – 818 ( 1978. ). [DOI] [PubMed] [Google Scholar]
  • 22. ) Sherblom , A. P. and Moody , C. E.Cell surface sialomucin and resistance to natural cell mediated cytotoxicity of rat mammary tumor ascites cells . Cancer Res. , 46 , 4543 – 4546 , ( 1986. ). [PubMed] [Google Scholar]
  • 23. ) Pearlstein , E. , Salk , P. L. , Yogeeswaran , G. and Karpakin , S.Correlation between spontaneous metastatic‐variant derivatives of rat renal sarcoma cell line . Proc. Natl. Acad. Sci. USA , 77 , 4336 – 4339 ( 1980. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24. ) Kijima‐Suda , I. , Miyamoto , Y. , Toyoshima , S. , Itoh , M. and Osawa , T.Inhibition of experimental pulmonary metastasis of mouse colon adenocarcinoma 26 subline by a sialic acid nucleoside conjugate having sialyltransferase inhibiting activity . Cancer Res. , 46 , 858 – 862 ( 1986. ). [PubMed] [Google Scholar]
  • 25. ) Blosher , J. M. , Schallier , D. C. C. , Van Rooy , H. , Storme , G. A. and Smets , L. A.Modification of cell surface carbohydrates and invasive behavior by an alkyl lysophospholipid . Cancer Res. , 48 , 977 – 982 ( 1988. ). [PubMed] [Google Scholar]
  • 26. ) Kannagi , R. , Fukushii , Y. , Tachikawa , T. , Noda , A. , Shin , S. , Shigeta , K. , Hirakawa N. , Fukuda Y. , Inamoto , T. , Hakomori , S. and Irimura , H.Quantitative and qualitative characterization of human cancer‐associated serum glycoprotein antigens expressing fucosyl or sialyl‐fucosyl type 2 chain poly lactosamine . Cancer Res. , 46 , 2619 – 2626 ( 1986. ). [PubMed] [Google Scholar]
  • 27. ) Itai , S. , Arii , S. , Tobe , R. , Kitahara , A. , Kim , Y. C. , Yamabe , H. , Ohhashi , H. , Kurihara , Y. , Shigeta , K. and Kannagi , R.Significance of 2‐3 and 2‐6 sialylation of Lewis antigen in pancreas cancer . Cancer , 61 , 775 – 787 ( 1988. ). [DOI] [PubMed] [Google Scholar]
  • 28. ) Itai , S. , Nishikawa , J. , Yoneda , T. , Ohmori , K. , Tsunekawa , S. , Hiraiwa , N. , Yamabe , H. , Arii , S. , Tobe , T. and Kannagi , R.Tissue distribution of sialyl 2‐3 and 2‐6 Lewis a antigen and the significance of serum 2‐3/2‐6 sialyl Lewis a antigen ratio for the differential diagnosis of malignant and benign disorders of the digestive tract . Cancer , 67 , 1576 – 1587 ( 1991. ). [DOI] [PubMed] [Google Scholar]
  • 29. ) Takada , A. , Ohmori , K. , Takahashi , N. , Tsuyuoka , K. , Yago , A. , Zenita , K. , Hasegawa , A. and Kannagi , R.Adhesion of human cancer cells to vascular endothelium mediated by a carbohydrate antigen, sialyl Lewis‐A . Biochem. Biophys. Res. Commun. , 179 , 713 – 719 ( 1991. ). [DOI] [PubMed] [Google Scholar]
  • 30. ) Takada , A. , Ohmori , K. , Yoneda , T. , Tsuyuoka , K. , Hasegawa , A. , Iso , M. and Kannagi , R.Contribution of carbohydrate antigens sialyl Lewis A and sialyl Lewis X to adhesion of human cancer cells to vascular endothelium . Cancer Res. , 53 , 354 – 361 ( 1993. ). [PubMed] [Google Scholar]
  • 31. ) Bevilacqua , M. P. , Pober , J. S. , Menderick , D. L. , Cotran , R. S. and Gimgrone , M. A. , Jr.Identification of an inducible endothelial‐leukocyte adhesion molecule . Proc. Natl. Acad. Sci. USA , 84 , 9238 – 9242 ( 1987. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 32. ) Bevilacqua , M. P. , Stengelin , S. , Gimbrone , M. A. , Jr. and Seed , B.Endothelial leukocyte adhesion molecule 1: an inducible receptor for neutrophils related to complement regulatory proteins and lectins . Science , 243 , 1160 – 1165 ( 1989. ). [DOI] [PubMed] [Google Scholar]
  • 33. ) Phillips , M. L. , Nudelman , E. , Gaeta , F. C. A. , Perez , M. , Singhal , A. K. , Hakomori , S. and Paulson , J. C.ELAM‐1 mediates cell adhesion by regression of a carbohydrate ligand, sialyl‐Lex . Science , 250 , 1130 – 1132 ( 1990. ). [DOI] [PubMed] [Google Scholar]

Articles from Japanese Journal of Cancer Research : Gann are provided here courtesy of Wiley

RESOURCES