The Effect of the Injinhotang Extract on Hepatocarcinogenesis in Rats

인진호탕 추출액의 투여가 흰쥐의 간암 발생에 미치는 효과

  • Yoon, Jung-Sik (Department of Anatomy, College of Oriental Medicine, Dongshin University) ;
  • Kim, Jeong-Sang (Department of Anatomy, College of Oriental Medicine, Dongshin University)
  • 윤중식 (동신대학교 한의과대학 해부학교실) ;
  • 김정상 (동신대학교 한의과대학 해부학교실)
  • Received : 2009.07.27
  • Accepted : 2009.10.23
  • Published : 2009.12.31

Abstract

In order to examine the effect of Injinhotang extract on the liver cancer induced by N-nitrosodiethylamine (NDEA) and carbon tetrachloride ($CCl_4$) in Rats. The animals were divided into three groups. The normal (Nor) group were fed basal diet. Control (Con) group were administered with NDEA (200 mg/kgb.w., i.p.) and $CCl_4$. Injinhotang extract (IJH) group treated with Injinhotang extract (260 mg/kg/day) for 8 weeks after NDEA+$CCl_4$. Enzymic antioxidants, such as superoxide dismutase (SOD) and catalase levels were determined in all the groups of animals. The activities of SOD were significantly increased in the Con, but the activities of catalase were decreased in the Con, but the anti-oxidative enzyme activities of superoxide dismutase and catalase were increased in the IJH. In the immunohistochemistry observation, treatment of Injinhotang extract reduced the rates of p53 immunoreactivity. According to the electron microscopical observation, in the liver cancer cells were increased the smooth endoplasmic reticulum and dilated the rough endoplasmic reticulum in the Con compared with IJH. These results suggest that administration of Injinhotang extract suppress or retard NDEA and $CCl_4$-induced liver cancer.

본 실험은 N-nitrosodiethylamine (NDEA)와 사염화탄소($CCl_4$) 로 유발된 흰쥐의 간암에 대한 인진호탕의 효과를 알아보고자 하였다. 먼저 실험군 설정은 세 군으로 분류하였다. 정상군(Nor)에는 일반적인 식이를 공급하였고, 대조군(Con)에는 NDEA(200 mg/kgb.w., i.p.)와 $CCl_4$를 투여하였고, 인진호탕 추출액 투여군(IJH)에는 인진호탕 추출액(260 mg/kg/day)을 8주 동안 투여하였다. Superoxide dismutase (SOD)와 catalase와 같은 항산화계 효소를 측정한 결과, SOD의 활성은 Con군에서 증가하였으나, catalase의 활성은 Con군에서 감소하였다. 면역조직화학적 관찰에서, 인진호탕 추출액의 투여로 인해 Con군에서보다 p53 면역 반응율을 감소시켰다. 전자현미경 관찰의 경우, 간암 세포들은 Con군에서 과립세포질세망의 팽대와 무과립세포질세망의 증가가 뚜렷하게 관찰되었다. 이상의 결과로 보아 인진호탕 추출액 투여가 NDEA와 사염화탄소로 유도된 흰쥐의 간암을 억제 또는 지연하는 것으로 사료된다.

Keywords

References

  1. Bourdon JC, D'Errico A, Paterlini P, Grigioni W, May E, Debuire B: p53 protein accumulation in European hepatocellular carcinoma is not always dependent on p53 gene mutation. Gastroenterology 108 : 1176-1182, 1995 https://doi.org/10.1016/0016-5085(95)90217-1
  2. Chu CY, Tseng TH, Hwang JM, Chou FP, Wang CJ: Protective effects of capillarisin on tertbutylhydro-peroxide-induced oxidative damage in rat primary hepatocytes. Arch Toxicol 73 :263-268, 1999 https://doi.org/10.1007/s002040050615
  3. Collier JD, Carpenter M, Burt AD, Bassendine MF: Expression of mutant p53 protein in hepatocellular carcinoma. Gut 35 : 98-100, 1994 https://doi.org/10.1136/gut.35.1.98
  4. Deng HL, Zhang JT: Anti-lipid peroxilative effect of ginsenoside Rb1 and Rg1. Chin Med J 104(5) : 395-398, 1991
  5. Dragan YP, Hully JR, Nakamura J, Mass MJ, Swenberg JA, Pitot HC: Biochemical events during initiation of rat hepatocarcinogenesis. Carcinogenesis 15 : 1451-1458, 1994 https://doi.org/10.1093/carcin/15.7.1451
  6. Eliyahu D, Raz A, Gruss P, Givol D, Oren M: Participation of p53 cellular tumour antigen in transformation of normal embryonic cells. Nature 312 : 646-649, 1984 https://doi.org/10.1038/312646a0
  7. Farber E: The multistep nature of cancer development. Cancer Res 44 : 4217-4223, 1984
  8. Greenblatt MS, Hollstein M, Harris CC: Mutations in the p53 tumor suppressor gene: clues to cancer etiology and molecular pathogenesis. Cancer Res 54 : 4855-4878, 1994
  9. Gutteridge JM, Beard AP, Quinlan GJ: Superoxide-dependent lipid peroxidation. Problems with the use of catalase as a specific probe for fenton-derived hydroxyl radicals. Biochem Biophys Res Commun 117 : 901-907, 1983 https://doi.org/10.1016/0006-291X(83)91681-9
  10. Han KH, Jeon YJ, Athukorala Y, Choi KD, Kim CJ, Cho JK, Sekikawa M, Fukushima M, Lee CH: A water extract of Artemisia capillaris prevents 2,2'-azobis(2-amidinopropane) dihydrochloride-induced liver damage in rats. J Med Food 9 : 342-347, 2006 https://doi.org/10.1089/jmf.2006.9.342
  11. Hinds PW, Finlay CA, Quartin RS, Baker SJ, Fearon ER, Vogelstein B, Levine AJ: Mutant p53 DNA clones from human colon carcinomas cooperate with ras in transforming primary rat cells: a comparison of the 'hot spot' mutant phenotypes. Cell Growth Differ 1 : 571-580, 1990
  12. Hu YQ, Tan RX, Chu MY, Zhou J: Apoptosis in human hepatoma cell line SMMC-7721 induced by water-soluble macromolecular components of Artemisia capillaris Thunberg. Jpn J Cancer Res 91 : 113-117, 2000 https://doi.org/10.1111/j.1349-7006.2000.tb00867.x
  13. Jin H, Sakaida I, Tsuchiya M, Okita K: Herbal medicine Rhei rhizome prevents liver fibrosis in rat liver cirrhosis induced by a choline-deficient L-amino acid-defined diet. Life Sci 76 : 2805-2816, 2005 https://doi.org/10.1016/j.lfs.2004.09.041
  14. Kimura Y, Okuda H, Arichi S: Effects of geniposide isolated from Gardenia jasminoides on metabolic alterations in high sugar diet-fed rats. Chem Pharm Bull 30 : 4444-4447, 1982 https://doi.org/10.1248/cpb.30.4444
  15. Kiso Y, Ogasawara S, Hirota K, Watanabe N, Oshima Y, Konno C, Hikino H: Antihepatotoxic principles of Artemisia capillaris buds. Planta Med 50 : 81-85, 1984 https://doi.org/10.1055/s-2007-969627
  16. Kovalszky I, Szeberenyi S, Zalatnai A, Vincze I, Lapis K, Jeney A: Modification of DENA-induced hepatocarcinogenesis by $CCl_4$ cirrhosis. Comparison of the marker enzyme patterns. Carcinogenesis 13(5) : 773-778, 1992 https://doi.org/10.1093/carcin/13.5.773
  17. Kus I, Ogeturk M, Oner H, Sahin S, Yekeler H, Sarsilmaz M: Protective effects of melatonin against carbon tetrachloride-induced hepatotoxicity in rats: a light microscopic and biochemical study. Cell Biochem Funct 23 : 169-174, 2005 https://doi.org/10.1002/cbf.1136
  18. Lee J, Chae K, Ha J, Park BY, Lee HS, Jeong S, Kim MY, Yoon M: Regulation of obesity and lipid disorders by herbal extracts from Morus alba, Melissa officinalis, and Artemisia capillaris in high-fat diet-induced obese mice. J Ethnopharmacol 115 :263-270, 2008 https://doi.org/10.1016/j.jep.2007.09.029
  19. Legros Y, Meyer A, Ory K, Soussi T: Mutations in p53 produce a common conformational effect that can be detected with a panel of monoclonal antibodies directed toward the central part of the p53 protein. Oncogene 9 : 3689-3694, 1994
  20. Li X, Zhou XP, Guan YS, Wang YX: Magnetic resonance imaging of hepatocellular carcinoma induced by diethylnitrosamine in Sprague-Dawley rats. Hepatobiliary Pancreat Dis Int 4 : 427-432, 2005
  21. Luo H, Lin S, Ren F, Wu L, Chen L, Sun Y: Antioxidant and antimicrobial capacity of Chinese medicinal herb extracts in raw sheep meat. J Food Prot 70 : 1440-1445, 2007 https://doi.org/10.4315/0362-028X-70.6.1440
  22. Matsuda H, Tomohiro N, Hiraba K, Harima S, Ko S, Matsuo K, Yoshikawa M, Kubo M: Study on anti-Oketsu activity of rhubarb II. Anti-allergic effects of stilbene components from Rhei undulati Rhizoma. Biol Pharm Bull 24 : 264-267, 2001 https://doi.org/10.1248/bpb.24.264
  23. Mori H, Xu QA, Sakamoto O, Uesugi Y, Ono Y, Koda A, Nishioka I: Immunological mechanisms of antitumor activity of some kinds of Chinese herbs: Meth A-induced delayed type hypersensitivity. Jpn J Pharmacol 48 : 37-46, 1988 https://doi.org/10.1254/jjp.48.37
  24. Mulcahy LS, Gans JH: The fidelity of mouse liver mitochondrial DNA polymerase following long-term administration of carbon tetrachloride, diethylnitrosamine, or phenobarbital. Mol Pharmacol 24 : 329-335, 1983
  25. Onishi M, Sokuza Y, Nishikawa T, Mori C, Uwataki K, Honoki K Tsujiuchi T: Different mutation patterns of mitochondrial DNA displacement-loop in hepatocellular carcinomas induced by Nnitrosodiethylamine and a choline-deficient l-amino acid-defined diet in rats. Biochem Biophys Res Commun 362 : 183-187, 2007 https://doi.org/10.1016/j.bbrc.2007.07.175
  26. Peng CH, Huang CN, Wang CJ: The anti-tumor effect and mechanisms of action of penta-acetyl geniposide. Curr Cancer Drug Targets 5 : 299-305, 2005 https://doi.org/10.2174/1568009054064633
  27. Scherer E, Emmelot P: Kinetics of induction and growth of precancerous liver-cell foci, and liver tumor formation by diethylnitrosamine in the rat. Eur J Cancer 11 : 689-696, 1975 https://doi.org/10.1016/0014-2964(75)90042-0
  28. Seol HC, Suzuki M, Ohnishi-Kameyama M, Oh MJ, Kim HR, Kim JH, Nagata T: Extraction and identification of antioxidant components from Artemisia capillaris herba. Plant Foods for Human Nutrition 58 : 1-12, 2003
  29. Tamano S, Merlino GT, Ward JM: Rapid development of hepatic tumors in transforming growth factor alpha transgenic mice associated with increased cell proliferation in precancerous hepatocellular lesions initiated by N-nitrosodiethylamine and promoted by phenobarbital. Carcinogenesis 15(9) : 1791-1798, 1994 https://doi.org/10.1093/carcin/15.9.1791
  30. Tan RX, Zheng WF, Tang HQ: Biologically active substances from the genus Artemisia. Planta Med 64 : 295-302, 1998 https://doi.org/10.1055/s-2006-957438
  31. Tessitore L, Pani P, Dianzani MU: Mechanisms of the enhanced liver carcinogenesis by choline in female rats: delay in liver growth after partial hepatectomy and stimulation of 2-AAF mitoinhibition. Carcinogenesis 13(10) : 1929-1932, 1992 https://doi.org/10.1093/carcin/13.10.1929
  32. Volkmann M, Hofmann WJ, M$\ddot{u}$ller M, R$\ddot{a}$th U, Otto G, Zentgraf H, Galle PR: p53 overexpression is frequent in European hepatocellular carcinoma and largely independent of the codon 249 hot spot mutation. Oncogene 9(1) : 195-204, 1994
  33. Watanabe S, Horie Y, Suzuki A: Hepatocyte-specific Pten-deficient mice as a novel model for nonalcoholic steatohepatitis and hepatocellular carcinoma. Hepatol Res 33 : 161-166, 2005 https://doi.org/10.1016/j.hepres.2005.09.026
  34. Yamahara J, Kobayashi G, Matsuda H, Katayama T, Fujimura H: The effect of scoparone, a coumarin derivative isolated from the Chinese crude drug Artemisiae capillaris flos, on the heart. Chem Pharm Bull 37 : 1297-1299, 1989 https://doi.org/10.1248/cpb.37.1297
  35. Yamahara J, Yamada T, Kimura H, Sawada T, Fujimura H: Biologically active principles of crude drugs. II. Anti-allergic principles in 'Shoseiryu-To' anti inflammatory properties of paeoniflorin and its derivatives. J Pharmacobiodyn 5 : 921-929, 1982 https://doi.org/10.1248/bpb1978.5.921
  36. Yoshikawa T, Murakami M, Yoshida N, Seto O, Kondo M: Effects of superoxide dismutase and catalase on disseminated intravascular coagulation in rats. Thromb Haemost 50 : 869-872, 1983