Skip to main content
SpringerLink
Log in

Isolation of a human gallbladder cancer cell clone with high invasive phenotype in vitro and metastatic potential in orthotopic model and inhibition of its invasiveness by heparanase antisense oligodeoxynucleotides

  • Original Paper
  • Published:
Clinical & Experimental Metastasis Aims and scope Submit manuscript

Abstract

The mechanisms involved in gallbladder cancer metastasis still remain unclear to date. The poor understanding is due, in part, to the lack of ideal cell line and animal model for study. In the present study, 21 cell clones were isolated from the human gallbladder carcinoma cells GBC-SD and the cell clone GBC-SDHi with high invasive phenotype was fished out. The invasive phenotype and metastatic potential of GBC-SDHi were confirmed in a novel surgical orthotopic implantation model of gallbladder cancer in nude mice. Heparanase, an endoglycosidase that degrades heparan sulfate, is a critical mediator of tumor metastasis and angiogenesis. RT-PCR, real time RT-PCR and western blot showed that the expression levels of heparanase were significant difference between GBC-SDHi and its parent cells. After treated with antisense oligodeoxynucleotides, the heparanase mRNA and protein expression in GBC-SDHi cells were significantly decreased and its invasive potential in vitro was inhibited in a dose-dependent manner. The study provides a useful cell clone and a clinically relevant orthotopic tumor model for the metastatic study in human gallbladder cancer. The roles of heparanase in gallbladder cancer are also evaluated.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

Abbreviations

SOI:

Surgical orthotopic implantation

HPSE:

Heparanase

HSPGs:

Heparan sulfate proteoglycans

EPM:

Electrophoretic mobility

ASODNs:

Antisense oligodeoxynucleotides

NSODNs:

Nonsense oligodeoxynucleotides

References

  1. Ito H, Matros E, Brooks DC, Osteen RT, Zinner MJ, Swanson RS, Ashley SW, Whang EE (2004) Treatment outcomes associated with surgery for gallbladder cancer: a 20-year experience. J Gastrointest Surg 8:183–190

    Article  PubMed  Google Scholar 

  2. Filder IJ (1978) Tumor heterogeneity and the biology of cancer invasion and metastasis. Cancer Res 38:2651–2660

    Google Scholar 

  3. Farabegoli F, Santini D, Ceccarelli C, Taffurelli M, Marrano D, Baldini N (2001) Clone heterogeneity in diploid and aneuploid breast carcinomas as detected by FISH. Cytometry 46:50–56

    Article  PubMed  CAS  Google Scholar 

  4. Bonsing BA, Corver WE, Fleuren GJ, Cleton-Jansen AM, Devilee P, Cornelisse CJ (2000) Allelotype analysis of flow-sorted breast cancer cells demonstrates genetically related diploid and aneuploid subpopulations in primary tumors and lymph node metastases. Genes Chromosomes Cancer 28:173–183

    Article  PubMed  CAS  Google Scholar 

  5. Fidler IJ (1990) Critical factors in the biology of human cancer metastasis: Twenty eighth G.H.A. Clowes Memorial Award Lecture. Cancer Res 50:6130–6138

    PubMed  CAS  Google Scholar 

  6. Shindo-Okada N, Takeuchi K, Nagamachi Y (2001) Establishment of cell lines with high- and low-metastatic potential from PC-14 human lung adenocarcinoma. Jpn J Cancer Res 92:174–183

    PubMed  CAS  Google Scholar 

  7. Navone NM, Olive M, Ozen M, Davis R, Troncoso P, Tu SM, Johnston D, Pollack A, Pathak S, von Eschenbach AC, Logothetis CJ (1997) Establishment of two human prostate cancer cell lines derived from a single bone metastasis. Clin Cancer Res 3:2493–2500

    PubMed  CAS  Google Scholar 

  8. Li Y, Tian B, Yang J, Zhao L, Wu X, Ye SL, Liu YK, Tang ZY (2004) Stepwise metastatic human hepatocellular carcinoma cell model system with multiple metastatic potentials established through consecutive in vivo selection and studies on metastatic characteristics. J Cancer Res Clin Oncol 130:460–468

    Article  PubMed  CAS  Google Scholar 

  9. Flatmark K, GunhildMælandsmo GM, Martinsen M, Rasmussen H, Fodstad ∅ (2004) Twelve colorectal cancer cell lines exhibit highly variable growth and metastatic capacities in an orthotopic model in nude mice. Eur J Cancer 40:1593–1598

    Article  PubMed  Google Scholar 

  10. Fan YZ, Fu JY, Zhao ZM, Chen CQ (2005) Effect of norcantharidin on proliferation and invasion of human gallbladder carcinoma GBC-SD cells. World J Gastroenterol. 11(16):2431–2437

    PubMed  CAS  Google Scholar 

  11. Finkel E (1999) Potential target found for antimetastasis drugs. Science 2:33

    Article  Google Scholar 

  12. Naomoto Y, Takaoka M, Okawa T, Nobuhisa T, Gunduz M, Tanaka N (2005) The role of heparanase in gastrointestinal cancer. Oncol Rep 14:3–8

    PubMed  CAS  Google Scholar 

  13. Xiao Y, Kleeff J, Shi X, Buchler MW, Friess H (2003) Heparanase expression in hepatocellular carcinoma and the cirrhotic liver. Hepatol Res 26:192–198

    Article  PubMed  CAS  Google Scholar 

  14. Dexter DL, Kowalski HM, Blazar BA, Fligiel Z, Vogel R, Heppner GH (1978) Heterogeneity of tumor cells from a single mouse mammary tumor. Cancer Res 38:3174–3181

    PubMed  CAS  Google Scholar 

  15. Omasu F, Nakano Y, Ichiki T (2005) Measurement of the electrophoretic mobility of sheep erythrocytes using microcapillary chips. Electrophoresis 26:1163–1167

    Article  PubMed  CAS  Google Scholar 

  16. Albini A, Iwamoto Y, Kleinman HK, Martin GR, Aaronson SA, Kozlowski JM, McEwan RN (1987) A rapid in vitro assay for quantitating the invasive potential of tumor cells. Cancer Res 47:3239–3245

    PubMed  CAS  Google Scholar 

  17. Zheng JP, Rudra GN, Powell WC et al (1999) Suppression of prostate carcinoma cell invasion by expression of antisense L-plastin. Gene Am J Pathol 155:115–122

    CAS  Google Scholar 

  18. Seabright M (1971) A rapid banding technique for human chromosomes. Lancet 2:971–972

    Article  PubMed  CAS  Google Scholar 

  19. Zhang YL, Fu ZR, Zhang J et al (2003) Inhibition of invasiveness of human mammary carcinoma cell line MDA435 by heparanase antisense oligodeoxynucleotide. Zhonghua Yi Xue Za Zhi 83:204–207

    PubMed  Google Scholar 

  20. Chen AB, Ding YQ, Xie XM, Liu Z, Zhang H (2002) Constructing a high metastasis subline from Lovo cell and studying its biological characteristics. Cancer Res Clin 14:85–88

    Google Scholar 

  21. Nakano T, Tani M, Ishibashi Y, Kimura K, Park YB, Imaizumi N, Tsuda H, Aoyagi K, Sasaki H, Ohwada S, Yokota J (2003) Biological properties and gene expression associated with metastatic potential of human osteosarcoma. Clin Exp Metastasis 20:665–674

    Article  PubMed  CAS  Google Scholar 

  22. Nowell PC (1976) The clonal evolution of tumor cell population. Science 194:23–28

    Article  PubMed  CAS  Google Scholar 

  23. Chang XZ, Wang ZM, Li ZW et al (2004) Establishment of human gallbladder cancer surgical orthotopic implantation model in nude mice. Chin J Hepatobiliary surg 10(7):468–472

    Google Scholar 

  24. Li Y, Tang ZY, Ye SL, Liu YK, Chen J, Xue Q, Chen J, Gao DM, Bao WH (2001) Establishment of cell clones with different metastatic potential from the metastatic hepatocellular carcinoma cell line MHCC97. World J Gastroenterol 7:630–636

    PubMed  CAS  Google Scholar 

  25. Takeuchi S, Nakanishi H, Yoshida K, Yamamoto S, Tonoki H, Tsukamoto T, Fukushima S, Moriuchi T, Kurita K, Tatematsu M (2000) Isolation of differentiated squamous and undifferentiated spindle carcinoma cell lines with differing metastatic potential from a 4-nitroquinoline N-Oxide-induced tongue carcinoma in a F344 rat. Jpn J Cancer Res 91:1211–1221

    PubMed  CAS  Google Scholar 

  26. Yang YJ, Zhang YL, Li X et al (2003) Contribution of eIF-4E inhibition to the expression and activity of heparanase in human colon adenocarcinoma cell line: LS-174T. World J Gastroenterol 9:1707–1712

    PubMed  CAS  Google Scholar 

  27. Yasoshima T, Denno R, Kawaguchi S, Sato N, Okada Y, Ura H, Kikuchi K, Hirata K (1996) Establishment of human gastric carcinoma lines with high metastatic potential in the liver: changes in integrin expression associated with the ability to metastasize in the liver of nude mice. Jpn J Cancer Res 87:153–160

    PubMed  CAS  Google Scholar 

  28. Samiei M, Waghorne CG (1991) Clonal selection within metastatic SPI mouse mammary tumors is independent of metastatic potential. Int J Cancer 47:771–775

    Article  PubMed  CAS  Google Scholar 

  29. Price JE, Bell C, Frost P (1990) The use of a genotypic marker to demonstrate clonal dominance during the growth and metastasis of a human breast carcinoma in nude mice. In J Cancer 45:968–971

    Article  CAS  Google Scholar 

  30. Hoffman RM (2005) Orthotopic metastatic (MetaMouse) models for discovery and development of novel chemotherapy. Methods Mol Med 111:297–322

    PubMed  CAS  Google Scholar 

  31. Katz MH, Takimoto S, Spivack D, Moossa AR, Hoffman RM, Bouvet M (2004) An imageable highly metastatic orthotopic red fluorescent protein model of pancreatic cancer. Clin Exp Metastasis 21:7–12

    Article  PubMed  CAS  Google Scholar 

  32. Illert B, Otto C, Braendlein S, Thiede A, Timmermann W (2003) Optimization of a metastasizing human gastric cancer model in nude mice. Microsurgery 23:508–512

    Article  PubMed  Google Scholar 

  33. Rashidi B, Gamagami R, Sasson A, Sun FX, Geller J, Moossa AR, Hoffman RM (2000) An orthotopic mouse model of remetastasis of human colon cancer liver metastasis. Clin Cancer Res 6:2556–2561

    PubMed  CAS  Google Scholar 

  34. Horiuchi H, Kawamata H, Fujimori T, Kuroda Y (2003) A MEK inhibitor (U0126) prolongs survival in nude mice bearing human gallbladder cancer cells with K-ras mutation: analysis in a novel orthotopic inoculation model. Int J Oncol 23:957–963

    PubMed  CAS  Google Scholar 

  35. An Z, Jiang P, Wang X, Moossa AR, Hoffman RM (1999) Development of a high metastatic orthotopic model of human renal cell carcinoma in nude mice: benefits of fragment implantation compared to cell-suspension injection. Clin Exp Metastasis 17:265–270

    Article  PubMed  CAS  Google Scholar 

  36. Chen JQ, Zhan WH, He YL et al (2004) Expression of heparanase gene, CD44v6, MMP-7 and nm23 protein and their relationship with the invasion and metastasis of gastric carcinomas. World J Gastroenterol 10:776–782

    PubMed  CAS  Google Scholar 

  37. Sinnappah-Kang ND, Kaiser AJ, Blust BE, Mrak RE, Marchetti D (2005) Heparanase, TrkC and p75NTR: their functional involvement in human medulloblastoma cell invasion. Int J Oncol 27:617–626

    PubMed  CAS  Google Scholar 

  38. Vlodavsky I, Friedmann Y, Elkin M et al (1999) Mammalian heparanase: gene cloning, expression and function in tumor progression and metastasis. Nat Med 5:793–802

    Article  PubMed  CAS  Google Scholar 

  39. Hulett MD, Freeman C, Hamdorf BJ et al (1999) Cloning of mammalian heparanase, an important enzyme in tumor invasion and metastasis. Nat Med 5:803–809

    Article  PubMed  CAS  Google Scholar 

  40. Parish CR, Freeman C, Hulett MD (2001) Heparanase: a key enzyme involved in cell invasion. Biochim Biophys Acta 1471:99–108

    Google Scholar 

  41. Kuniyasu H, Chihara Y, Kubozoe T, Takahashi T (2002) Co-expression of CD44v3 and heparanase is correlated with metastasis of human colon cancer. Int J Mol Med 10:333–337

    PubMed  Google Scholar 

  42. Kim AW, Xu X, Hollinger EF et al (2002) Human heparanase-1 gene expression in pancreatic adenocarcinoma. J Gastrointest Surg 6:167–172

    Article  PubMed  Google Scholar 

  43. Rohloff J, Zinke J, Schoppmeyer K et al (2002) Heparanase expression is a prognostic indicator for postoperative survival in pancreatic adenocarcinoma. Br J Cancer 86:1270–1275

    Article  PubMed  CAS  Google Scholar 

  44. Uno F, Fujiwara T, Takata Y et al (2001) Antisense-mediated suppression of human heparanase gene expression inhibits pleural dissemination of human cancer cells. Cancer Res 61:7855–7860

    PubMed  CAS  Google Scholar 

  45. Chen KS, Zhang L, Tang L, Zhang YH, Gao DL, Yan L, Zhang L (2005) Expression of heparanase mRNA in anti-sense oligonucleotide-transfected human esophageal cancer EC9706 cells. World J Gastroenterol 11:4916–4919

    PubMed  CAS  Google Scholar 

  46. Miao HQ, Elkin M, Aingorn E et al (1999) Inhibition of heparanase activity and tumor metastasis by laminarin sulfate and synthetic phosphorothioate oligodeoxynucleotides. Int J Cancer 83:424–431

    Article  PubMed  CAS  Google Scholar 

  47. Parish CR, Freeman C, Brown KJ et al (1999) Identification of sulfated oligosaccharide-based inhibitors of tumor growth and metastasis using novel in vitro assays for angiogenesis and heparanase activity. Cancer Res 59:3433–3441

    PubMed  CAS  Google Scholar 

Download references

Acknowledgements

We gratefully acknowledge Xiao-Peng Wu, Zhi-Lun Zhao, Dao-Xin Ma, Bo Liu, Lei Wang and Lu-Bin Tang for their excellent technical assistance in cell culture, cell electrophoresis, western blot, in vitro invasion and migration assays, respectively. This research was supported in part by the Grant from Shandong Science and Technology Committee (No. 003130107 and No. 963000053).

Author information

Authors and Affiliations

  1. Central Laboratory, Cancer Hospital/Cancer Institute, Fudan University, Shanghai, 200032, P. R. China

    Xin-Zhong Chang, Wei Jin, Lian-Fang Li, Xiao-Feng Liu & Zhi-Min Shao

  2. Qilu Hospital, Shandong University, Jinan, 250012, P. R. China

    Zhan-Min Wang & Zhi-Wei Li

  3. Shandong Cancer Hospital and Institute, Jinan, 250117, P. R. China

    Jin-Ming Yu & Yi Zhang

  4. Shanxi Cancer Hospital, Taiyuan, 030000, P. R. China

    Fu-Guo Tian

  5. Weifang University, Weifang, 261041, P. R. China

    Jie Yu

  6. Department of Breast Surgery, Breast Cancer Institute, Cancer Hospital/Cancer Institute, Fudan University, 270 Dong’An Road , Shanghai, 200032, P. R. China

    Zhi-Min Shao

Authors
  1. Xin-Zhong Chang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Zhan-Min Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jin-Ming Yu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Fu-Guo Tian
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Wei Jin
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Yi Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Jie Yu
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Lian-Fang Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Xiao-Feng Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Zhi-Wei Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Zhi-Min Shao
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Zhi-Min Shao.

Additional information

This work was supported in part by the Grant from Shandong Science and Technology Committee (No. 003130107 and No. 963000053)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Chang, XZ., Wang, ZM., Yu, JM. et al. Isolation of a human gallbladder cancer cell clone with high invasive phenotype in vitro and metastatic potential in orthotopic model and inhibition of its invasiveness by heparanase antisense oligodeoxynucleotides. Clin Exp Metastasis 24, 25–38 (2007). https://doi.org/10.1007/s10585-006-9053-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10585-006-9053-7

Keywords

Search

Navigation