2次元電気泳動法により検出された非小細胞肺癌関連蛋白質の免疫組織化学染色による検証  [in Japanese] Validation Analysis Using Immunohistochemistry of Non-small Cell Lung Cancer-associated Proteins Detected by Two-dimensional Polyacrylamide Gel Electrophoresis  [in Japanese]

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Abstract

<b>目的</b>.原発性肺癌の生物学的性格を反映するバイオマーカーの探索を目的にEttan-DIGE法による肺癌組織型関連蛋白の検出・同定を試みた.<b>方法</b>.組織型ごとに4~8症例の組織分化が中等度~高分化症例の2次元電気泳動(2-DE)用サンプルを混合し,組織型標準サンプルとして蛍光標識後に2-DEを施行,各組織型で高発現するスポットを検出した.各スポットは質量分析法で蛋白質分子を同定,検証は肺癌腫瘍組織の免疫組織化学染色によって行った.<b>結果</b>.19種類の扁平上皮癌関連蛋白質(eSq),16種類の腺癌関連蛋白質(eAd),17種類の神経内分泌癌関連蛋白質(eNE)を検出,このうち6種類のeSq,8種類のeAdを同定した.同定蛋白質の14種類のうち8種類がサイトケラチン(CK)(eSq:CK5,CK6A,CK6C,CK6D,CK17,eAd:CK8,CK18,CK19)であった.その後の免疫組織染色による検証で各組織型間でのCKの発現に特徴があることを示すことができた.<b>考察</b>.腫瘍関連蛋白質の発現量に基づいて肺癌を評価することで,腫瘍の生物学的性格を反映させることが可能となり,治療法選択に応用できると考える.<br>

<i><b>Objective</b></i>. We set out to detect primary lung cancer (PLC)-associated proteins using 2-dimensional polyacrylamide gel electrophoresis (2-DE) analysis and to identify the protein molecules for the exploration of biomarkers reflecting the biological characteristics of lung cancer. <i><b>Study Design</b></i>. We prepared samples for 2-DE from representative histological types of primary lung cancer; squamous cell carcinoma, adenocarcinoma and small cell carcinoma. We selected 8 cases of well or moderately differentiated squamous cell carcinoma confirmed postoperativly to possess typical histological features of primary lung squamous cell carcinoma. The mixture that contained equal amounts of these 8 samples was taken to be a standard sample of lung squamous cell carcinoma. Similary, we selected 8 cases of well or moderately differentiated adenocarcinoma to prepare a standard sample of primary lung adenocarcinoma. We also mixed specimens of 4 cases of small cell carcinoma, and prepared a standard sample of small cell carcinoma. We then labeled each standard sample with a fluorescent dye (either Cy2, Cy3 or Cy5) according to the manufacturer's protocol. Furthermore, using mass spectrometry we identified protein molecules that were detected in 2-DE analysis, followed by validation analysis by immunohistochemistry. <i><b>Results</b></i>. We detected 19 kinds of squamous cell carcinoma-associated spots (eSq), 16 kinds of adenocarcinoma-associated spots (eAd) and 17 kinds of neuroendocrine carcinoma-associated spots (eNE). From these spots, 6 kinds of eSq-protein molecules and 8 kinds of eAd-protein molecules were identified. Eight protein molecules out of these 14 identified molecules were cytokeratin (CK) molecules. CK5, CK6A, CK6C, CK6D and CK17 were identified as eSq-protein molecules, and furthermore CK8, CK18 and CK19 were identified as eAd-protein molecules. The results of a validation analysis using immunohistochemistry indicated a high possibility that CK5, CK5/6 and CK17 are biomarkers for squamous cell carcinoma, and that CK18 is a biomarker for adenocarcinoma. We succeeded in showing a significant relationship between the histopathological differentiation of PLC and the expression of CKs. <i><b>Conclusion</b></i>. There is a strong possibility that the biological characteristics of lung carcinoma may be clarified by classification based upon proteomic analysis, and the results of proteomic analysis may be applied to the selection of therapeutic strategies of PLC.<br>

Journal

  • Haigan

    Haigan 47(7), 861-869, 2007-12-20

    The Japan Lung Cancer Society

References:  22

Codes

  • NII Article ID (NAID)
    110006571323
  • NII NACSIS-CAT ID (NCID)
    AN00203978
  • Text Lang
    JPN
  • Article Type
    ART
  • ISSN
    03869628
  • Data Source
    CJP  NII-ELS  J-STAGE 
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