Journal of Physiology & Pathology in Korean Medicine (동의생리병리학회지)

The Physiological Society of Korean Medicine and The Society of Pathology in Korean Medicine (대한동의생리학회·한의병리학회)
권호 표지

Induction of Apoptosis by Samgibopae-tang in Human Non-small-cell Lung Cancer Cells

인체폐암세포 NCI-H460 및 A549의 증식에 미치는 삼기보폐탕의 영향 비교

  • Heo, Man-Kyu (Department of Oriental Medicine, Graduate School, Dong-Eui University) ;
  • Park, Cheol (Department of Oriental Medicine, Graduate School, Dong-Eui University) ;
  • Choi, Young-Hyun (Department of Oriental Medicine, Graduate School, Dong-Eui University) ;
  • Kam, Cheol-Woo (Department of Oriental Medicine, Graduate School, Dong-Eui University) ;
  • Park, Dong-Il (Department of Oriental Medicine, Graduate School, Dong-Eui University)
  • 허만규 (동의대학교 한의과대학 한의학과) ;
  • 박철 (동의대학교 한의과대학 한의학과) ;
  • 최영현 (동의대학교 한의과대학 한의학과) ;
  • 감철우 (동의대학교 한의과대학 한의학과) ;
  • 박동일 (동의대학교 한의과대학 한의학과)
  • Published : 2007.08.25
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Abstract

In the present study, we investigated the antiproliferative activity of the water extract of Samgibopae-tang (SGBPT) in NCI-H460 and A549 non-small-cell lung cancer cell lines. We found that exposure of A549 cells to SGBPT resulted in the growth inhibition in a dose-dependent manner as measured by MTT assay, however SGBPT did not affect the growth of NCI-H460 cells. The antiproliferative effect by SGBPT treatment in A549 cells was associated with morphological changes such as membrane shrinking and cell rounding up. SGBPT treatment did not induce the cell cycle arrest in both cell lines, however the frequency of sub-G1 population was concentration-dependently increased by SGBPT treatment in A549 cells. SGBPT treatment partially induced the expression of tumor suppressor p53 in A549 cells and the expression of cyclin-dependent kinase inhibitor p21(WAF1/CIP1) was markedly increased in both transcriptional and translational levels in A549 cells. The up-regulation of p21 by SGBPT occurred in a similar a concentration dependent manner to that observed with the inhibition of cell viability and induction of sub-G1 population of the cell cycle. However SGBPT treatment did not affect other growth regulation-related genes such as early growth response-1 (Egr-1), nonsteroidal anti-inflammatory drug-activated gene-1 (NAG-1), inducible nitric oxide synthease (iNOS), cyclooxygenases (COXs), telomere-regulatory factors in A549 as well as NCI-H460 cells. Taken together, these findings suggested that SGBPT-induced inhibition of human lung carcinoma A549 cell growth was aoosciated with the induction of p21 and the results provided important new insights into the possible molecular mechanisms of the anti-cancer activity of SGBPT.

Keywords

References

  1. 해리슨 내과학 편찬위원회. 해리슨 내과학. 서울, 정담 출판사, pp 1317-1324, 1997
  2. 이은우, 김영철 역. 머크 매뉴얼. 서울, 한우리, p 702, 2667, 2002
  3. 윤창열, 김용진. 난경연구집성. 서울, 주민출판사, p 768, 769, 2002
  4. 許俊. 東醫寶鑑. 서울, 법인문화사, p 1433, 1996
  5. 李梴. 醫學入門. 서울, 남산당, p 212, 213, 1980
  6. 전국한의과대학 폐계내과학교실편저. 동의폐계내과학. 서울, 한문화사, p 87, 526, 2002
  7. 김영환. 분자생물학적 측면에서의 폐암의 발생. 녹십자의보, 27(3):152-157, 1999
  8. 임재형, 김훈, 변미권, 감철우, 박동일. 인체폐암세포의 증식 및 prostaglandin E2 생성에 미치는 청조구폐탕의 영향에 관한 연구. 동의생리병리학회지 20(4):966-972, 2006
  9. 조인주, 감철우, 김기탁, 박동일. 인체폐암세포에서 Bcl-2 발현저하 및 caspase 활성을 통한 청조구폐탕의 apoptosis 유발에 관한 연구. 동의생리병리학회지 21(1):93-97, 2007
  10. 홍수현, 박철, 홍상훈, 최병태, 이용태, 박동일, 최영현. 어성초 메탄올 추출물에 의한 A549 인체 폐암세포 사멸유도에 관한 연구. 동의생리병리학회지 20(6):1584-1592, 2006
  11. 박봉규, 박동일. 천금위경탕의 인체 폐암 세포 증식억제에 관한 연구. 동의생리병리학회지 18(4):1147-1153, 2004
  12. 이성열, 김원일, 박동일. 길경이 인체 폐암세포에 미치는 영향에 대한 실험적 연구. 동의생리병리학회지 17(4):1019-1030, 2003
  13. Evans, V.G. Multiple pathways to apoptosis. Cell Biol. Int 17: 461-476, 1993 https://doi.org/10.1006/cbir.1993.1087
  14. Arends, M.J., Morris, R.G., Wyllie, A.H. Apoptosis. The role of the endonuclease. Am. J. Pathol. 136: 593-608, 1990
  15. Zhan, Q., Fan, S., Bae, I., Guillouf, C., Liebermann, D.A., OConnor, P.M., Fornace, A.J. Jr. Induction of bax by genotoxic stress in human cells correlates with normal p53 status and apoptosis. Oncogene. 9: 3743-3751, 1994
  16. Shi. L., Nishioka, W.K., Th'ng, J., Bradbury, E.M., Litchfield, D.W., Greenberg, A.H. Premature p34cdc2 activation required for apoptosis. Science. 263: 1143-1145, 1994 https://doi.org/10.1126/science.8108732
  17. Chiarugi, V., Magnelli, L., Basi, G. Apoptosis and the cell cycle. Cell. Mol. Biol. Res. 40: 603-612, 1994
  18. Reed, J.C. Bcl-2 family proteins. Oncogene. 17: 3225-3236, 1998 https://doi.org/10.1038/sj.onc.1202591
  19. Miyashita, T., Reed, J.C. Tumor suppressor p53 is a direct transcriptional activator of the human bax gene. Cell. 80: 293-299, 1995 https://doi.org/10.1016/0092-8674(95)90412-3
  20. El-Deiry, W.S., Harper, J.W., O'Connor, P.M., Velculescu, V.E., Canman, C.E., Jackman, J., Pietenpol, J.A., Burrell, M., Hill, D.E., Wang, Y., Wiman, K.G., Mercer, W.E., Kastan, M.B., Kohn, K.W., Elledge, S.J., Kinzler, K.W., Vogelstain, B. WAF1/CIP1 is induced in p53-mediated G1 arrest and apoptosis. Cancer Res. 54: 1169-1174, 1994
  21. Kluck, R.M., Bossy-Wetzel, E., Green, D.R., Newmeyer, D.D. The release of cytochrome c from mitochondria: a primary site for Bcl-2 regulation of apoptosis. Science. 275: 1132-1136, 1997 https://doi.org/10.1126/science.275.5303.1132
  22. Nagata, S. Apoptosis by death factor. Cell. 88: 355-365, 1997 https://doi.org/10.1016/S0092-8674(00)81874-7
  23. Tsujimoto, Y. Role of Bcl-2 family proteins in apoptosis: apoptosomes or mitochondria. Genes Cells. 3: 697-707, 1998 https://doi.org/10.1046/j.1365-2443.1998.00223.x
  24. Cheng, J.Q., Jiang, X., Fraser, M., Li, M., Dan, H.C., Sun, M., Tsang, B.K. Role of X-linked inhibitor of apoptosis protein in chemoresistance in ovarian cancer: possible involvement of the phosphoinositide-3 kinase/Akt pathway. Drug Resist. Update. 5: 131-146, 2002 https://doi.org/10.1016/S1368-7646(02)00003-1
  25. Salvesen, G.S., Duckett, C.S. IAP proteins: blocking the road to death's door. Nat. Rev. Mol. Cell. Biol. 3: 401-410, 2002 https://doi.org/10.1038/nrm830
  26. Holcik, M., Gibson, H., Korneluk, R.G. XIAP: apoptotic brake and promising therapeutic target. Apoptosis. 6: 253-261, 2001 https://doi.org/10.1023/A:1011379307472
  27. LaCasse, E.C., Baird, S., Korneluk, R.G., MacKenzie, A.E. The inhibitors of apoptosis (IAPs) and their emerging role in cancer. Oncogene. 17: 3247-3259, 1998 https://doi.org/10.1038/sj.onc.1202569
  28. Giercksky, K.E. COX-2 inhibition and prevention of cancer. Best Pract. Res. Clin. Gastroenterol. 15: 821-833, 2001 https://doi.org/10.1053/bega.2001.0237
  29. Turini, M.E., DuBois, R.N. Cyclooxygenase-2: a therapeutic target. Annu. Rev. Med. 53: 35-57, 2002 https://doi.org/10.1146/annurev.med.53.082901.103952
  30. Chiarugi, V., Magnelli, L., Gallo, O. Cox-2, iNOS and p53 as play-makers of tumor angiogenesis. Int. J. Mol. Med. 2: 715-719, 1998
  31. Surh, Y.J., Chun, K.S., Cha, H.H., Han, S.S., Keum, Y.S., Park, K.K., Lee, S.S. Molecular mechanisms underlying chemopreventive activities of anti-inflammatory phytochemicals: down-regulation of COX-2 and iNOS through suppression of NF-$\kappa$B activation. Mutat. Res. pp 243-268, 480-481, 2001
  32. Thun, M.J., Henley, S.J., Patrono, C. Nonsteroidal anti-inflammatory drugs as anticancer agents: mechanistic, pharmacologic, and clinical issues. J. Natl. Cancer Inst. 94: 252-266, 2002 https://doi.org/10.1093/jnci/94.4.252
  33. oole, J.C., Andrews, L.G., Tollefsbol, T.O. Activity, function, and gene regulation of the catalytic subunit of telomerase (hTERT). Gene. 269: 1-12, 2001 https://doi.org/10.1016/S0378-1119(01)00440-1
  34. Kyo, S., Inoue, M. Complex regulatory mechanisms of telomerase activity in normal and cancer cells: How can we apply them for cancer therapy. Oncogene. 21: 688-697, 2002 https://doi.org/10.1038/sj.onc.1205163
  35. Fabian, D., Koppel, J., Maddox-Hyttel, P. Apoptotic processes during mammalian preimplantation development. Theriogenology 64: 221-231, 2005 https://doi.org/10.1016/j.theriogenology.2004.11.022
  36. Baek, S.J., Kim, J.S., Nixon, J.B., DiAugustine, R.P., Eling, T.E. Expression of NAG-1, a transforming growth factor-beta superfamily member, by troglitazone requires the early growth response gene EGR-1. J Biol Chem. 279: 6883-6892, 2004 https://doi.org/10.1074/jbc.M305295200
  37. Liu, C., Rangnekar, V.M., Adamson, E., Mercola, D. Suppression of growth and transformation and induction of apoptosis by EGR-1. Cancer Gene Ther 5: 3-28, 1998
  38. Calogero, A., Arcella, A., de Gregorio, G., Porcellini, A., Mercola, D., Liu, C., Lombari, V., Zani, M., Giannini, G., Gagliardi, F.M., Caruso, R., Gulino, A., Frati, L., Ragona, G. The early growth response gene EGR-1 behaves as a suppressor gene that is down-regulated independent of ARF/Mdm2 but not p53 alterations in fresh human gliomas. Clin Cancer Res 7: 2788-2796, 2001
  39. Huang, R.P., Fan, Y., de Belle, I., Niemeyer, C., Gottardis, M.M., Mercola, D., Adamson, E.D. Decreased Egr-1 expression in human, mouse and rat mammary cells and tissues correlates with tumor formation. Int J Cancer 72: 102-109, 1997 https://doi.org/10.1002/(SICI)1097-0215(19970703)72:1<102::AID-IJC15>3.0.CO;2-L
  40. de Belle, I., Huang, R.P., Fan, Y., Liu, C., Mercola, D., Adamson, E.D. p53 and Egr-1 additively suppress transformed growth in HT1080 cells but Egr-1 counteracts p53-dependent apoptosis. Oncogene 18: 3633-3642, 1999 https://doi.org/10.1038/sj.onc.1202696
  41. Virolle, T., Adamson, E.D., Baron, V., Birle, D., Mercola, D., Mustelin, T., de Belle, I. The Egr-1 transcription factor directly activates PTEN during irradiation-induced signalling. Nat Cell Biol 3: 1124-1128, 2001 https://doi.org/10.1038/ncb1201-1124
  42. Huang, R.P., Liu, C., Fan, Y., Mercola, D., Adamson, E.D. Egr-1 negatively regulates human tumor cell growth via the DNA-binding domain. Cancer Res 55: 5054-5062, 1995
  43. Eling, T.E., Baek, S.J., Shim, M., Lee, C.H. NSAID activated gene (NAG-1), a modulator of tumorigenesis. J Biochem Mol Biol 39: 649-655, 2006 https://doi.org/10.5483/BMBRep.2006.39.6.649
  44. Baek, S.J., Kim, K.S., Nixon, J.B., Wilson, L.C., Eling, T.E. Cyclooxygenase inhibitors regulate the expression of a TGF-$\beta$ superfamily member that has proapoptotic and antitumorigenic activities. Mol Pharmacol 59: 901-908, 2001 https://doi.org/10.1124/mol.59.4.901
  45. Tan, M., Wang, Y., Guan, K., Sun, Y. PTGF-$\beta$, a type beta transforming growth factor (TGF-$\beta$) superfamily member, is a p53 target gene that inhibits tumor cell growth via TGF-$\beta$ signaling pathway. Proc Natl Acad Sci USA 97: 109-114, 2000
  46. Li, P.X., Wong, J., Ayed, A., Ngo, D., Brade, A.M., Arrowsmith, C., Austin, R.C., Klamut, H.J. Placental transforming growth factor-$\beta$ is a downstream mediator of the growth arrest and apoptotic response of tumor cells to DNA damage and p53 overexpression. J Biol Chem 275: 20127-20135, 2000 https://doi.org/10.1074/jbc.M909580199
  47. Baek, S.J., Wilson, L.C., Eling, T.E. Resveratrol enhances the expression of non-steroidal anti-inflammatory drug-activated gene (NAG-1) by increasing the expression of p53. Carcinogenesis 23: 425-434, 2002 https://doi.org/10.1093/carcin/23.3.425
  48. Koff, A., Giordano, A., Desai, D., Yamashita, K., Harper, J.W., Elledge, S., Nishimoto, T., Morgan, D.O., Franza, B.R., Roberts, J.M. Formation and activation of a cyclin E-cdk2 complex during the G1 phase of the human cell cycle. Science 257: 1689-1694, 1992 https://doi.org/10.1126/science.1388288
  49. Zeng, Y.X., el-Deiry, W.S. Regulation of p21WAF1/CIP1 expression by p53-independent pathways. Oncogene 12: 1557-1564, 1996
  50. Morgan, D.O. Principles of CDK regulation. Nature 374: 131-134, 1995
  51. Harper, J.W., Adami, G.R., Wei, N., Keyomarsi, K., Elledge, S.J. The p21 Cdk-interacting protein Cip1 is a potent inhibitor of G1 cyclin-dependent kinases. Cell 75: 805-816, 1993 https://doi.org/10.1016/0092-8674(93)90499-G
  52. Xiong, Y., Hannon, G., Zhang, H., Casso, D., Kobayashi, R., Beach, D. p21 is a universal inhibitor of cyclin kinases. Nature 366: 701-704, 1993 https://doi.org/10.1038/366701a0
  53. Vainio, H. Is COX-2 inhibition a panacea for cancer prevention? Int J Cancer 94: 613-614, 2001 https://doi.org/10.1002/ijc.1518
  54. Dempke, W., Rie, C., Grothey, A., Schmoll, H.J. Cyclooxygenase-2: a novel target for cancer chemotherapy? J Cancer Res Clin Oncol 127: 411-417, 2001 https://doi.org/10.1007/s004320000225
  55. Sawaoka, H., Tsuji, S., Tsujii, M., Gunawan, E.S., Sasaki, Y., Kawano, S., Hori, M. Cyclooxygenase inhibitors suppress angiogenesis and reduce tumor growth in vivo. Lab Invest 79: 1469-1477, 1999
  56. Yamamoto, Y., Gaynor, R.B. Therapeutic potential of inhibition of the NF-${\kappa}B$ pathway in the treatment of inflammation and cancer. J Clin Invest 107: 135-142, 2001 https://doi.org/10.1172/JCI11914
  57. Cerni, C. Telomeres, telomerase, and myc. An update. Mutat Res 462: 31-47, 2000 https://doi.org/10.1016/S1383-5742(99)00091-5