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A three-dimensional cell biology model of human hepatocellular carcinoma in vitro

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Tumor Biology

Abstract

We established an in vitro 3-D model of metastatic hepatocellular carcinoma (HCC) by culturing MHCC97H cells on molecular scaffolds within a rotating wall vessel bioreactor. Morphological and biochemical analyses revealed that the 3-D HCC model mirrored many clinical pathological features of HCC in vivo, including cancer cell morphology, tissue ultrastructure, protein production and secretion, glucose metabolism, tissue-specific gene expression, and apoptosis. Xenografts into livers of nude mice resulted in tumorigenesis and distant metastasis. This 3-D HCC spheroid is a promising model for HCC tumor biology, anticancer drug screening, and for the establishment of HCC animal models.

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References

  1. Kim JB. Three-dimensional tissue culture models in cancer biology. Semin Cancer Biol. 2005;15:365–77.

    Article  PubMed  Google Scholar 

  2. Wang R, Xu J, Juliette L, Castilleja A, Love J, Sung SY, et al. Three -dimensional co-culture models to study prostate cancer growth, progression, and metastasis to bone. Semin Cancer Biol. 2005;15:353–64.

    Article  PubMed  CAS  Google Scholar 

  3. Chang TT, Hughes-Fulford M. Monolayer and spheroid culture of human liver hepatocellular carcinoma cell line cells demonstrate distinct global gene expression patterns and functional phenotypes. Tissue Eng Part A. 2009;15:559–67.

    Article  PubMed  CAS  Google Scholar 

  4. Grun B, Benjamin E, Sinclair J, Timms JF, Jacobs IJ, Gayther SA, et al. Three-dimensional in vitro cell biology models of ovarian and endometrial cancer. Cell Prolif. 2009;42:219–28.

    Article  PubMed  CAS  Google Scholar 

  5. Pampaloni F, Reynaud EG, Stelzer EH. The third dimension bridges the gap between cell culture and live tissue. Nat Rev Mol Cell Biol. 2007;8:839–45.

    Article  PubMed  CAS  Google Scholar 

  6. Unsworth BR, Lelkes PI. Growing tissues in microgravity. Nat Med. 1998;4:901–7.

    Article  PubMed  CAS  Google Scholar 

  7. Navran S. The application of low shear modeled microgravity to 3-D cell biology and tissue engineering. Biotechnol Annu Rev. 2008;14:275–96.

    Article  PubMed  CAS  Google Scholar 

  8. Santamaría E, Muñoz J, Fernández-Irigoyen J, Prìeto J, Corrales FJ. Toward the discovery of new biomarkers of hepatocellular carcinoma by proteomics. Liver Int. 2007;27:163–73.

    Article  PubMed  Google Scholar 

  9. Aravalli RN, Steer CJ, Cressman EN. Molecular mechanisms of hepatocellular carcinoma. Hepatology. 2008;48:2047–63.

    Article  PubMed  CAS  Google Scholar 

  10. Blum HE. Hepatocellular carcinoma: therapy and prevention. World J Gastroenterol. 2005;11:7391–400.

    PubMed  CAS  Google Scholar 

  11. Yang Y, Nagano H, Ota H, Morimoto O, Nakamura M, Wada H, et al. Patterns and clinicopathologic features of extrahepatic recurrence of hepatocellular carcinoma after curative resection. Surgery. 2007;141:196–202.

    Article  PubMed  Google Scholar 

  12. Miyoshi A, Kitajima Y, Kido S, Shimonishi T, Matsuyama S, Kitahara K, et al. Snail accelerates cancer invasion by upregulating MMP expression and is associated with poor prognosis of hepatocellular carcinoma. Br J Cancer. 2005;92:252–8.

    PubMed  CAS  Google Scholar 

  13. Tang ZY, Ye SL, Liu YK, Qin LX, Sun HC, Ye QH, et al. A decade’s studies on metastasis of hepatocellular carcinoma. J Cancer Res Clin Oncol. 2004;130:187–96.

    Article  PubMed  Google Scholar 

  14. Li Y, Tian B, Yang J, Zhao L, Wu X, Ye SL, et al. 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. 2004;130:460–8.

    Article  PubMed  CAS  Google Scholar 

  15. Yan L, Zhao-You T, Sheng-Long Y, Yin-Kun L, Jie C, Qiong X, et al. Establishment of cell clones with different metastatic potential from the metastatic hepatocellular carcinoma cell line MHCC97. World J Gastroenterol. 2001;7:630–6.

    Google Scholar 

  16. Cui JF, Liu YK, Zhang LJ, Shen HL, Song HY, Dai Z, et al. Identification of metastasis candidate proteins among HCC cell lines by comparative proteome and biological function analysis of S100A4 in metastasis in vitro. Proteomics. 2006;6:5953–61.

    Article  PubMed  CAS  Google Scholar 

  17. Tian B, Li Y, Ji XN, Chen J, Xue Q, Ye SL, et al. Basement membrane proteins play an active role in the invasive process of human hepatocellular carcinoma cells with high metastasis potential. J Cancer Res Clin Oncol. 2005;131:80–6.

    Article  PubMed  CAS  Google Scholar 

  18. Li WC, Ye SL, Sun RX, Liu YK, Tang ZY, Kim Y, et al. Inhibition of growth and metastasis of human hepatocellular carcinoma by antisense oligonucleotide targeting signal transducer and activator of transcription3. Clin Cancer Res. 2006;12:7140–8.

    Article  PubMed  CAS  Google Scholar 

  19. Mueller-Klieser W. Multicellular spheroids. A review on cellular aggregates in cancer research. J Cancer Res Clin Oncol. 1987;113:101–22.

    Article  PubMed  CAS  Google Scholar 

  20. Lopez JB, Balasegaram M, Thambyrajah V, Timor J. The value of liver function tests in hepatocellular carcinoma. Malays J Pathol. 1996;18:95–9.

    PubMed  CAS  Google Scholar 

  21. Tsutsumi M, Sakamuro D, Takada A, Zang SC, Furukawa T, Taniguchi N. Detection of a unique gamma-glutamyl transpeptidase messenger RNA species closely related to the development of hepatocellular carcinoma in humans: a new candidate for early diagnosis of hepatocellular carcinoma. Hepatology. 1996;23:1093–7.

    PubMed  CAS  Google Scholar 

  22. Weber G. Enzymology of cancer cells (second of two parts). N Engl J Med. 1977;296:541–51.

    Article  PubMed  CAS  Google Scholar 

  23. Sun FX, Tang ZY, Lui KD, Ye SL, Xue Q, Gao DM, et al. Establishment of a metastatic model of human hepatocellular carcinoma in nude mice via orthotopic implantation of histologically intact tissues. Int J Cancer. 1996;66:239–43.

    Article  PubMed  CAS  Google Scholar 

  24. Tian J, Tang ZY, Ye SL, Liu YK, Lin ZY, Chen J, et al. New human hepatocellular carcinoma (HCC) cell line with highly metastatic potential (MHCC97) and its expressions of the factors associated with metastasis. Br J Cancer. 1999;81:814–21.

    Article  PubMed  CAS  Google Scholar 

  25. Kenny PA, Lee GY, Myers CA, Neve RM, Semeiks JR, Spellman PT, et al. The morphologies of breast cancer cell lines in three-dimensional assays correlate with their profiles of gene expression. Mol Oncol. 2007;1:84–96.

    Article  PubMed  CAS  Google Scholar 

  26. Feder-Mengus C, Ghosh S, Reschner A, Martin I, Spagnoli GC. New dimensions in tumor immunology: what does 3D culture reveal? Trends Mol Med. 2008;14:333–40.

    Article  PubMed  CAS  Google Scholar 

  27. Bissell MJ, Radisky DC, Rizki A, Weaver VM, Petersen OW. The organizing principle: microenvironmental influences in the normal and malignant breast. Differentiation. 2002;70:537–46.

    Article  PubMed  Google Scholar 

  28. Kim JB, Stein R, O’Hare MJ. Three-dimensional in vitro tissue culture models of breast cancer—a review. Breast Cancer Res Treat. 2004;85:281–91.

    Article  PubMed  Google Scholar 

  29. Yoffe B, Darlington GJ, Soriano HE, Krishnan B, Risin D, Pellis NR, et al. Cultures of human liver cells in simulated microgravity environment. Adv Space Res. 1999;24:829–36.

    Article  PubMed  CAS  Google Scholar 

  30. Khaoustov VI, Darlington GJ, Soriano HE, Krishnan B, Risin D, Pellis NR, et al. Induction of three-dimensional assembly of human liver cells by simulated microgravity. In Vitro Cell Dev Biol Anim. 1999;35:501–9.

    Article  PubMed  CAS  Google Scholar 

  31. Miyazawa M, Torii T, Toshimitsu Y, Koyama I. Effect of mechanical stress imposition on co-culture of hepatic parenchymal and nonparenchymal cells: possibility of stimulating production of regenerating factor. Transplant Proc. 2005;37:2398–401.

    Article  PubMed  CAS  Google Scholar 

  32. Miyazawa M, Torii T, Toshimitsu Y, Okada K, Koyama I. Hepatocyte dynamics in a three-dimensional rotating bioreactor. J Gastroenterol Hepatol. 2007;22:1959–64.

    Article  PubMed  CAS  Google Scholar 

  33. Dayong Xu, Yun W, Meifu F. Studies on HepG2 growth under simulated microgravity: to establish a method for three-dimensional cultivation in vitro as a research model. Prog Biochem Biophys. 2007;34:146–53.

    Google Scholar 

  34. Arii S, Mise M, Harada T, Furutani M, Ishigami S, Niwano M, et al. Overexpression of matrix metalloproteinase 9 gene in hepatocellular carcinoma with invasive potential. Hepatology. 1996;24:316–22.

    Article  PubMed  CAS  Google Scholar 

  35. Sun JJ, Zhou XD, Liu YK, Tang ZY, Feng JX, Zhou G, et al. Invasion and metastasis of liver cancer: expression of intercellular adhesion molecule 1. J Cancer Res Clin Oncol. 1999;125:28–34.

    Article  PubMed  CAS  Google Scholar 

  36. Hirohashi K, Yamamoto T, Uenishi T, Ogawa M, Sakabe K, Takemura S, et al. CD44 and VEGF expression in extrahepatic metastasis of human hepatocellular carcinoma. Hepatogastroenterology. 2004;51:1121–3.

    PubMed  Google Scholar 

  37. Osada T, Sakamoto M, Ino Y, Iwamatsu A, Matsuno Y, Muto T, et al. E-cadherin is involved in the intrahepatic metastasis of hepatocellular carcinoma. Hepatology. 1996;24:1460–7.

    Article  PubMed  CAS  Google Scholar 

  38. Liu LX, Jiang HC, Liu ZH, Zhou J, Zhang WH, Zhu AL, et al. Integrin gene expression profiles of human hepatocellular carcinoma. World J Gastroenterol. 2002;8:631–7.

    PubMed  CAS  Google Scholar 

  39. Hayasaka A, Suzuki N, Fujimoto N, Iwama S, Fukuyama E, Kanda Y, et al. Elevated plasma levels of matrix metalloproteinase-9 (92-kd type IV collagenase/gelatinase B) in hepatocellular carcinoma. Hepatology. 1996;24:1058–62.

    Article  PubMed  CAS  Google Scholar 

  40. Lin RZ, Chou LF, Chien CC, Chang HY. Dynamic analysis of hepatoma spheroid formation: roles of E-cadherin and beta1-integrin. Cell Tissue Res. 2006;324:411–22.

    Article  PubMed  CAS  Google Scholar 

  41. Islam S, Carey TE, Wolf GT, Wheelock MJ, Johnson KR. Expression of N-cadherin by human squamous carcinoma cells induces a scattered fibroblastic phenotype with disrupted cell–cell adhesion. J Cell Biol. 1996;135:1643–54.

    Article  PubMed  CAS  Google Scholar 

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Acknowledgement

The authors sincerely thank their colleagues Jun Chen, Ruixia Sun, and Qiong Xue for the kind help in animal experiments. This study was sponsored by grants from National Natural Science Foundation of China (No.30772062 and No.81071902), China National High-Tech Research and Development Program (2006AA02A-308), China National Key Projects for Infectious Disease (2008ZX10002-021 and 2008ZX 10002-017), Shanghai Pujiang Program (No.08PJ140300), and Shanghai Natural Science Foundation (09ZR1406400).

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Authors and Affiliations

  1. Liver Cancer Institute, Zhongshan Hospital, Fudan University and Key Laboratory of Carcinogenesis and Cancer Invasion, Ministry of Education, 136 Xue Yuan Road, Shanghai, 200032, People’s Republic of China

    Jianhua Tang, Jiefeng Cui, Rongxin Chen, Kun Guo, Yan Li, Dongmei Gao, Jie Chen, Zhaoyou Tang & Yinkun Liu

  2. Research Center for Cancer, Institute of Biomedical Science, Fudan University, Shanghai, 200032, People’s Republic of China

    Xiaonan Kang & Lu Sun

  3. Department of Nuclear Medicine, the Fifth People’s Hospital of Shanghai, Fudan University, Shanghai, 200240, People’s Republic of China

    Changde Xu

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  1. Jianhua Tang
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  2. Jiefeng Cui
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Correspondence to Yinkun Liu.

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Jianhua Tang and Jiefeng Cui contributed equally to this work.

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Tang, J., Cui, J., Chen, R. et al. A three-dimensional cell biology model of human hepatocellular carcinoma in vitro. Tumor Biol. 32, 469–479 (2011). https://doi.org/10.1007/s13277-010-0140-7

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  • DOI: https://doi.org/10.1007/s13277-010-0140-7

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