Tuberculosis and Respiratory Diseases

The Korean Academy of Tuberculosis and Respiratory Diseases (대한결핵및호흡기학회)
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Expression of Peroxiredoxin and Thioredoxin in Human Lung Cancer and Paired Normal Lung

인체의 폐암과 정상 폐조직에서 Peroxiredoxin 및 Thioredoxin의 발현 양상

  • Kim, Young Sun (Department of Pulmonary and Critical Care Medicine, Ajou University School of Medicine) ;
  • Park, Joo Hun (Department of Pulmonary and Critical Care Medicine, Ajou University School of Medicine) ;
  • Lee, Hye Lim (Department of Pulmonary and Critical Care Medicine, Ajou University School of Medicine) ;
  • Shim, Jin Young (Department of Pulmonary and Critical Care Medicine, Ajou University School of Medicine) ;
  • Choi, Young In (Department of Pulmonary and Critical Care Medicine, Ajou University School of Medicine) ;
  • Oh, Yoon Jung (Department of Pulmonary and Critical Care Medicine, Ajou University School of Medicine) ;
  • Shin, Seung Soo (Department of Pulmonary and Critical Care Medicine, Ajou University School of Medicine) ;
  • Choi, Young Hwa (Department of Pulmonary and Critical Care Medicine, Ajou University School of Medicine) ;
  • Park, Kwang Joo (Department of Pulmonary and Critical Care Medicine, Ajou University School of Medicine) ;
  • Park, Rae Woong (Department of Medical Informatics, Ajou University School of Medicine) ;
  • Hwang, Sung Chul (Department of Pulmonary and Critical Care Medicine, Ajou University School of Medicine)
  • 김영선 (아주대학교 의과대학 호흡기 내과학교실) ;
  • 박주헌 (아주대학교 의과대학 호흡기 내과학교실) ;
  • 이혜림 (아주대학교 의과대학 호흡기 내과학교실) ;
  • 심진영 (아주대학교 의과대학 호흡기 내과학교실) ;
  • 최영인 (아주대학교 의과대학 호흡기 내과학교실) ;
  • 오윤정 (아주대학교 의과대학 호흡기 내과학교실) ;
  • 신승수 (아주대학교 의과대학 호흡기 내과학교실) ;
  • 최영화 (아주대학교 의과대학 호흡기 내과학교실) ;
  • 박광주 (아주대학교 의과대학 호흡기 내과학교실) ;
  • 박래웅 (아주대학교 의과대학 의료정보학과) ;
  • 황성철 (아주대학교 의과대학 호흡기 내과학교실)
  • Received : 2005.04.27
  • Accepted : 2005.07.13
  • Published : 2005.08.30
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Abstract

Background : Continuous growth stimulation by various factors, as well as chronic oxidative stress, may co-exist in many solid tumors, such as lung cancer. A new family of antioxidant proteins, the peroxiredoxins (Prxs), have been implicated in the regulation of many cellular processes, including cell proliferation, differentiation and apoptosis. However, a real pathophysiological significance of Prx proteins, especially in lung disease, has not been sufficiently defined. Therefore, this study was conducted to investigate the distribution and expression of various Prx isoforms in lung cancer and other pulmonary conditions. Method : Patients diagnosed with lung cancer, and who underwent surgery at the Ajou Medical Center, were enrolled. The expressions of Prxs, Thioredoxin (Trx) and Thioredoxin reductase (TR) were analyzed using proteomic techniques and the subcellular localization of Prx proteins was studied using immunohistochemistry on normal mouse lung tissue. Result : Immunohistochemical staining has shown the isoforms of Prx I, II, III and V are predominantly expressed in bronchial and alveolar lining epithelia, as well as in the alveolar macrophages of the normal mouse lung. The isoforms of Prx I and III, and thioredoxin were also found to be over-expressed in the lung cancer tissues compared to their paired normal lung controls. There was also an increased amount of the oxidized form of Prx I, as well as a putative truncated form of Prx III, in the lung cancer samples when analyzed using 2-dimensional electrophoresis. In addition, a 43 kDa intermediate molecular weight protein band, and other high molecular weight bands of over 20 kDa, recognized by the anti-Prx I antibody, were present in the tissue extracts of lung cancer patients on 1-Dimensional electrophoresis, which require further investigation. Conclusion : The over-expressions of Prx I and III, and Trx in human lung cancer tissue, as well as their possible chaperoning function, may represent an attempt by tumor cells to adjust to their microenvironment in a manner advantageous to their survival and proliferation, while maintaining their malignant potential.

연구배경 : Peroxiredoxin (Prx) 은 최근에 알려진 항산화제로 세포의 증식과 분화, 세포사멸이나 발암과정에 관여하는 것으로 알려져 있다. 하지만, 현재까지 폐암을 비롯한 각종 질병에서의 이들 Prx단백의 역할은 잘 규명되어 있지 않다. 이에 본 연구는 폐암 조직과 정상폐조직에서 Prx 단백의 발현 양상과 분포를 연구하여 이들의 병태 생리학적인 의미를 찾아보고자 하였다. 방 법 : 아주대학교 병원에서 폐암으로 진단된 환자의 폐암조직과, 동일 환자의 정상 폐조직에서, 1 차원전기영동 (reducing 조건과 non-reducing 조건에서의 SDSPAGE) 혹은 2차원전기영동을 시행한 후 Western blot으로 Prx, Trx 및 TR의 발현 양상을 분석 하였으며, 백서의 정상 폐조직과, 환자의 폐암 조직에서 anti-Prx rabbit polyclonal 항체로 하여 면역조직화학염색법을 통해, Prx 단백의 분포를 관찰하였다. 결 과 : 면역조직화학염색법 결과 백서의 정상 폐조직에서는 Prx I, II, III 및 V 유형이, 주로 기관지상피세포, 폐포상피세포 및 폐포대식세포에서 발현되고 있음을 관찰하였다. 인체 폐암조직에서는, 정상 폐조직 부위에 비해서 Prx I 과 Prx III 유형 및 Trx단백의 발현이 선택적으로 증가되어 있고, 특히, 2차원 전기영동을 통한 프로티옴 분석에서 산화된 형태의 Prx I 과 Prx II가 증가 한 것을 비롯하여, 분자량과 등전점(pI)이 약간 변화된 형태의 Prx III가 폐암조직에 존재함을 알 수 있었다. 한편, 폐암 조직의 non-reducing 전기영동 후 Western blot에서는, monomer와 dimer 사이의 중간 크기에 해당하는 약 40 kDa과 200 kDa 이상 크기의, 항 Prx 항체와 반응하는 단백 띠 (reactive bands)가 관찰되었다. 결 론 : 폐암 조직에서 관찰되는 Prx I 과 Prx III 유형 및 Trx의 과발현 양상은, 종양 세포들이 주변의 미세 환경으로부터 겪는 여러 스트레스에 대하여 단백질을 보호하고 세포의 생명력을 유지하는데 있어 주요한 역할을 하는 것으로 사료된다.

Keywords

References

  1. Lavrovsky Y, Chatterjee B, Clark RA, Roy AK. Role of redox-regulated transcription factors in inflammation, aging and age-related diseases. Exp Gerontol 2000;35:521-32 https://doi.org/10.1016/S0531-5565(00)00118-2
  2. Abraham E, Arcaroli J, Carmody A, Wang H, Tracey KJ. HMG-1 as a mediator of acute lung inflammation. J Immunol 2000;165:2950-4 https://doi.org/10.4049/jimmunol.165.6.2950
  3. Abraham E, Carmody A, Shenkar R, Arcaroli J. Neutrophils as early immunologic effectors in hemorrhage- or endotoxemia-induced acute lung injury. Am J Physiol Lung Cell Mol Physiol 2000;279:L1137-45 https://doi.org/10.1152/ajplung.2000.279.6.L1137
  4. Abraham E. Nuclear factor-kappaB and its role in sepsis-associated organ failure. J Infect Dis 2003;187(Suppl 2):S364-9 https://doi.org/10.1086/367707
  5. Jones RD, Hancock JT, Morice AH. NADPH oxidase: a universal oxygen sensor? Free Radic Biol Med 2000;29:416-24 https://doi.org/10.1016/S0891-5849(00)00320-8
  6. Dhaunsi GS, Paintlia MK, Kaur J, Turner RB. NADPH oxidase in human lung fibroblasts. J Biomed Sci 2004;11:617-22 https://doi.org/10.1007/BF02256127
  7. Marshall C, Mamary AJ, Verhoeven AJ, Marshall BE. Pulmonary artery NADPH-oxidase is activated in hypoxic pulmonary vasoconstriction. Am J Respir Cell Mol Biol 1996;15:633-44 https://doi.org/10.1165/ajrcmb.15.5.8918370
  8. Griendling KK. Novel NAD(P)H oxidases in the cardiovascular system. Heart 2004;90:491-3 https://doi.org/10.1136/hrt.2003.029397
  9. Stone JR. An assessment of proposed mechanisms for sensing hydrogen peroxide in mammalian systems. Arch Biochem Biophys 2004;422:119-24 https://doi.org/10.1016/j.abb.2003.12.029
  10. Finkel T, Holbrook NJ. Oxidants, oxidative stress and the biology of ageing. Nature 2000;408:239-47 https://doi.org/10.1038/35041687
  11. Rhee SG, Chang TS, Bae YS, Lee SR, Kang SW. Cellular regulation by hydrogen peroxide. J Am Soc Nephrol 2003;14:S211-5 https://doi.org/10.1097/01.ASN.0000077404.45564.7E
  12. Forman HJ, Torres M. Redox signaling in macrophages. Mol Aspects Med 2001;22:189-216 https://doi.org/10.1016/S0098-2997(01)00010-3
  13. Chae HZ, Kim IH, Kim K, Rhee SG. Cloning, sequencing, and mutation of thiol-specific antioxidant gene of Saccharomyces cerevisiae. J Biol Chem 1993;268:16815-21
  14. Chae HZ, Rhee SG. A thiol-specific antioxidant and sequence homology to various proteins of unknown function. Biofactors 1994;4:177-80
  15. Chae HZ, Robison K, Poole LB, Church G, Storz G, Rhee SG. Cloning and sequencing of thiol-specific antioxidant from mammalian brain: alkyl hydroperoxide reductase and thiol-specific antioxidant define a large family of antioxidant enzymes. Proc Natl Acad Sci U S A 1994;91:7017-21
  16. Hofmann B, Hecht HJ, Flohe L. Peroxiredoxins. Biol Chem 2002;383:347-64 https://doi.org/10.1515/BC.2002.040
  17. Yanagawa T, Ishikawa T, Ishii T, Tabuchi K, Iwasa S, Bannai S, et al. Peroxiredoxin I expression in human thyroid tumors. Cancer Lett 1999;145:127-32 https://doi.org/10.1016/S0304-3835(99)00243-8
  18. Noh DY, Ahn SJ, Lee RA, Kim SW, Park IA, Chae HZ. Overexpression of peroxiredoxin in human breast cancer. Anticancer Res 2001;21:2085-90
  19. Lee SC, Na YP, Lee JB. Expression of peroxiredoxin II in vascular tumors of the skin: a novel vascular marker of endothelial cells. J Am Acad Dermatol 2003;49:487-91 https://doi.org/10.1067/S0190-9622(03)01485-3
  20. Dierick JF, Wenders F, Chainiaux F, Remacle J, Fisher AB, Toussaint O. Retrovirally mediated overexpression of peroxiredoxin VI increases the survival of WI-38 human diploid fibroblasts exposed to cytotoxic doses of tert-butylhydroperoxide and UVB. Biogerontology 2003;4:125-31 https://doi.org/10.1023/A:1024154024602
  21. Woo HA, Kang SW, Kim HK, Yang KS, Chae HZ, Rhee SG. Reversible oxidation of the active site cysteine of peroxiredoxins to cysteine sulfinic acid: immunoblot detection with antibodies specific for the hyperoxidized cysteine-containing sequence. J Biol Chem 2003;278:47361-4 https://doi.org/10.1074/jbc.C300428200
  22. Biteau B, Labarre J, Toledano MB. ATP - dependent reduction of cysteine-sulphinic acid by S. cerevisiae sulphiredoxin. Nature 2003;425:980-4 https://doi.org/10.1038/nature02075
  23. Budanov AV, Sablina AA, Feinstein E, Koonin EV, Chumakov PM. Regeneration of peroxiredoxins by p53-regulated sestrins, homologs of bacterial AhpD. Science 2004;304:596-600 https://doi.org/10.1126/science.1095569
  24. Jang HH, Lee KO, Chi YH, Jung BG, Park SK, Park JH, et al. Two enzymes in one: two yeast peroxiredoxins display oxidative stress-dependent switching from a peroxidase to a molecular chaperone function. Cell 2004;117:625-35 https://doi.org/10.1016/j.cell.2004.05.002
  25. Oh YJ, Kim YS, Choi YI, Sheen SS, Park JH, Hwang SC, et al. Oxidative inactivation of peroxiredoxin isoforms by $H_{2}O_{2}$ in pulmonary epithelial, macrophage, and other cell lines with their subsequent regeneration. Tuberc Respir Dis 2005;58:31-42 https://doi.org/10.4046/trd.2005.58.1.31
  26. Chang TS, Cho CS, Park S, Yu S, Kang SW, Rhee SG. Peroxiredoxin III, a mitochondrion-specific peroxidase, regulates apoptotic signaling by mitochondria. J Biol Chem 2004;279:41975-84 https://doi.org/10.1074/jbc.M407707200
  27. Kim HJ, Chae HZ, Kim YJ, Kim YH, Hwangs TS, Park EM, et al. Preferential elevation of Prx I and Trx expression in lung cancer cells following hypoxia and in human lung cancer tissues. Cell Biol Toxicol 2003;19:285-98 https://doi.org/10.1023/B:CBTO.0000004952.07979.3d
  28. Park SH, Chung YM, Lee YS, Kim HJ, Kim JS, Chae HZ, et al. Antisense of human peroxiredoxin II enhances radiation-induced cell death. Clin Cancer Res 2000;6:4915-20
  29. Koo KH, Lee S, Jeong SY, Kim ET, Kim HJ, Chae HZ et al. Regulation of thioredoxin peroxidase activity by C-terminal truncation. Arch Biochem Biophys 2002;397:312-8 https://doi.org/10.1006/abbi.2001.2700