Production and Characterization of a Transgenic Mouse Model of Human Liver Cancer

형질 전환 기법을 이용한 인체 간암의 마우스 모델 제작 및 특성 규명

  • Li, Zhong-Shu (Department of Animal Science, College of Agriculture, Yanbian University) ;
  • Lee, Jung-Woong (Medical Genomics Research Center, Korea Research Institute of Bioscience & Biotechnology(KRIBB)) ;
  • Hyun, Byung-Hwa (Disease Model Research Center, Korea Research Institute of Bioscience & Biotechnology(KRIBB)) ;
  • Lee, Chul-Ho (Disease Model Research Center, Korea Research Institute of Bioscience & Biotechnology(KRIBB)) ;
  • Jeong, Kyu-Shick (Disease Model Research Center, Korea Research Institute of Bioscience & Biotechnology(KRIBB)) ;
  • Fang, Nan-Zhu (Department of Animal Science, College of Agriculture, Yanbian University) ;
  • Yeom, Young-Il (Medical Genomics Research Center, Korea Research Institute of Bioscience & Biotechnology(KRIBB))
  • 이종숙 (중국길림성 연변대학 농학원) ;
  • 이정웅 (한국생명공학연구원 유전체의학연구센터) ;
  • 현병화 (한국생명공학연구원 질환모델연구센터) ;
  • 이철호 (한국생명공학연구원 질환모델연구센터) ;
  • 정규식 (한국생명공학연구원 질환모델연구센터) ;
  • 방남수 (중국길림성 연변대학 농학원) ;
  • 염영일 (한국생명공학연구원 유전체의학연구센터)
  • Published : 2007.09.30

Abstract

Transgenic mice were generated by microinjecting a plasmid DNA containing the SV40 (simian virus 40) large T antigen (Tag) gene fused with mouse albumin promoter/enhancer sequences into fertilized one-cell mouse embryos. Among eleven founder transgenic animals, four developed hepatocellular carcinoma, two showed kidney cancer and one developed skin and brain tumors. Three stable transgenic lines, #1-2, #1-6 and #1-11 were established. Members of the lines #1-6 and #1-11 reproducibly developed liver tumors by 8 to 10 weeks of age but did not exhibit any phenotypic changes in other tissues. Histological changes loading to liver tumor formation occurred with predictable kinetics and could be classified into three distinct stages; (a) newborn to 3 weeks of age, characterized by hyperplastic hepatocytes with reduced amounts of cytoplasm without any nuclear alterations, (b) between 4 to 8 weeks of age, characterized by diffuse liver cell dysplasia without observable tumor nodules, and (c) 9 weeks of age and thereafter, characterized by hepatocellular carcinomas in the background of extensive liver dysplasia. Metastasis to the lung from a liver carcinoma was observed in #1-11 founder animal. This transgenic mouse system displays similarities with human liver cancers in a number of aspects and provides a useful model for the study of molecular events involved in hepatocarcinogenesis.

본 연구에서는 SV40 Tag을 마우스 albumin 유전자의 promoter/enhancer 조절 하에 발현하도록 설계된 재조합 유전자를 마우스 1세포기 수정란에 미세 주입하여 형질 전환마우스를 제작하고 이들의 인체 간암 모델로써의 적합성을 조사하였다. 형질 전환이 확인된 총 11개체의 founder 생쥐들 중 4개체가 간암을 일으켰고, 두 개체는 신장암을, 한 개체는 피부 및 뇌에서 종양을 각각 일으켰다. 이들로부터 외래 유전자를 계대 유전하는 3가계를 얻었다(#1-2, #1-6, #1-11). 이들 가계의 자소들에서 8주령(#1-2, #1-6)혹은 10주령(#1-11)시부터 간암이 반복적으로 발생되었으며, #1-11 founder개체에서 폐로 암세포가 전이된 것 외에는 다른 조직에서의 형태학적 변이가 발견되지 않았다. 간암 발생은 조직학적 변화에 따라 3단계로 나눌 수 있었다. 즉, 출생에서 3주령까지는 간세포의 과량증식을 보이나 세포핵의 이상은 관찰되지 않았으며, 4주령부터 7주령(#1-2, #1-6) 혹은 9주령(#1-11)까지는 diffuse liver cell dysplasia를 나타내지만 tumor nodule은 발견되지 않았고, 그 이후에는 liver dysplasia를 배경으로 간암이 발생하였다. 본 연구에서 작출한 간암 모델 마우스는 인체 간암과 일부 유사한 소견을 보였는바 인체 간암 기전 연구를 위한 유용한 동물 모델로 이용할 수 있을 것으로 생각된다.

Keywords

References

  1. Bosch, FX, Munoz, N (1989): Epidemiology of in hepatocellular carcinoma. In: Bannasch P, Keppler D, Weber G (eds). Liver cell carcinoma. 1st ed. Kluwer Academic Publishers, The Netherlands, pp 3-14
  2. Cady B (1983): Natural history of primary and secondary tumors of the liver. Semin Oneal 10:127-134
  3. Dragan YP, Sargent L, Xu YD, Xy YH, Pitot HC (1993): The initiation-promotion-progression model of rat hepatocarcinogenesis. Proc Soc Exp Biol Med 202:16-24
  4. Dubois N, Bennoun M, Allemand I, Molina T, Grimber G, Daudet-Monsac M, Abelanet R, Briand P (1991): Time course development of differentiated hepatocarcinoma and lung metastasis in transgenic mice. J Hepatol 13:227-239 https://doi.org/10.1016/0168-8278(91)90819-W
  5. Fanning E, Knippers R (1992): Structure and function of simian virus 40 large tumor antigen. Annu Rev Biochem 61:55-85 https://doi.org/10.1146/annurev.bi.61.070192.000415
  6. Farber E, Chen Z- Y, Harris L, Lee G, Rinaudo JS, Roomi WM, Rotstein J, Semple E (1989): The biochemical-molecular pathology of the stepwise development of liver cancer: new insights and problems. In: Bannasch P, Keppler D, Weber G (eds). Liver cell carcinoma. 1st ed. Kluwer Academic Publishers, The Netherlands, pp 273-291
  7. Gordon JW (1994): Transgenic mouse models of hepatocellular carcinoma. Hepatology 19:538-539 https://doi.org/10.1002/hep.1840190236
  8. Gorski K, Carneiro M, Schibler U (1986): Tissuespecific in vitro transcription from the mouse albumin promoter. Cell 47:767-776 https://doi.org/10.1016/0092-8674(86)90519-2
  9. Grisham JW (1996): Interspecies comparison of liver carcinogenesis: implications for cancer risk assessment. Carcinogenesis 18:59-81 https://doi.org/10.1093/carcin/18.1.59
  10. Hahn WC, Counter CM, Lundberg AS, Beijersbergen RL, Brooks MW, Weinberg RA (1999): Creation of human tumour cells with defined genetic elements. Nature, 400:464-468 https://doi.org/10.1038/22780
  11. Harbst RS, Friedman N, Darnell JE Jr, Babiss LE (1989): Positive and negative regulatory elements in the mouse albumin enhancer. Proc Natl Acad Sci USA 86:1553-1557
  12. Hino O, Kitagawa T, Nomura K, Ohtake K, Cui L, Furuta Y, Aizawa S (1991): Hepatocarcinogenesis in transgenic mice carrying albumin-promoted SV40 T antigen gene. Jpn J Cancer Res 82:1226-1233 https://doi.org/10.1111/j.1349-7006.1991.tb01785.x
  13. Jhappan C, Stahle C, Harkins RN, Fausto N, Smith GH, Merlino GT (1990): TGF $\alpha$ overexpression in transgenic mice induces liver neoplasia and abnormal development of the mammary gland and pancreas. Cell 61:1137-1146 https://doi.org/10.1016/0092-8674(90)90076-Q
  14. Kanzler S, Galle PR (2000): Apoptosis and the liver. Cancer BioI 10:173-184 https://doi.org/10.1006/scbi.2000.0318
  15. Kokura K, Nakadai T, Kishimoto T, Makino Y, Muramatsu M, Tamura TA (1998): Gene expression in hepatomas. J Gastroenterol Hepatol 13:S132-141 https://doi.org/10.1111/jgh.1998.13.s1.132
  16. Moore MA and Kitagawa T (1986): Hepatocarcinogenesis in the rat: the effect of promoters and carcinogens in vivo and in vitro. Int Rev Cytol 101: 125-173 https://doi.org/10.1016/S0074-7696(08)60248-X
  17. Preston-Martin S, Pike MC, Ross RK, Jones PK, and Henderson BE (1990): Increased cell division as a cause of human cancer. Cancer Res 50:7425-7421
  18. Sandgren EP, Palmiter RD, Heckel JL, Brinster RL (1992): Hepatocellular carcinogenesis in albuminplasmminogen activator trangenic mice. Proc Natl Acad Sci USA 89:11523-11527
  19. Sandgren EP, Quaife CJ, Pinkert CA, Palmiter RD, Brinster RL (1989): Oncogene-induced liver neoplasia in transgenic mice. Oncogene 4:715-724
  20. Schafer DF, Sorrell MF (1999): Hepatocellular carcinoma. The Lancet 353:1253-1257 https://doi.org/10.1016/S0140-6736(98)09148-X
  21. Sepulveda AS, Finegold MJ, Smith B, Slagle BL, DeMayo JL, Shen RF, Woo SLC, Butel JS (1989): Development of a transgenic mouse system for the lysis of stages in liver carcinogenesis using tissue-specific expression of SV40 T-Antigen controlled by regulatory elements of the human $\alpha$-1-antitrypsin gene. Cancer Research 49:6108-6117
  22. Skamoto M Hirohashi S, Shimosato Y (1991): Early stages of multistep hepatocarcinogenesis: adenomatous hyperplasia and early hepatocellular carcinoma. Hum Pathol 22:172-178 https://doi.org/10.1016/0046-8177(91)90039-R
  23. Stuver SO (1998): Towards global control of liver cancer? Cancer BioI 8:299-306 https://doi.org/10.1006/scbi.1998.0079
  24. Thorgeirsson SS, Eric SR (1996): Transgenic mouse models in carcinogenesis: interaction of c-myc with transforming growth factor a and hepatocyte growth factor in hepatocarcinogenesis. Br J Clin Pharmacol 42:43-52 https://doi.org/10.1046/j.1365-2125.1996.03748.x
  25. Tzeng YJ, Zimmermann C, Guhll E, Berg B, Avantaggiati ML, Graessmann A (1998): SV40T/t-antigen induces premature mammary gland involution by apoptosis and selects for p53 missense mutation in mammary tumors. Oncogene 16:2103-2114 https://doi.org/10.1038/sj.onc.1201733
  26. 김창민 (1996): 발암유전자와 간암. 생명공학동향 3:57-63