Skip to main content
NCBI home page
Japanese Journal of Cancer Research : Gann logoLink to Japanese Journal of Cancer Research : Gann
. 1999 Feb;90(2):188–193. doi: 10.1111/j.1349-7006.1999.tb00732.x

Detection of Apoptotic Cells in Human Colorectal Cancer by Two Different in situ Methods: Antibody against Single‐stranded DNA and Terminal Deoxynucleotidyl Transferase‐mediated dUTP‐biotin Nick End‐labeling (TUNEL) Methods

Ichizo Watanabe 1,✉, Masao Toyoda 1,✉, Junji Okuda 1,✉, Toshiyuki Tenjo 1,✉, Keitaro Tanaka 1,✉, Tetsuhisa Yamamoto 1,✉, Hiroshi Kawasaki 1,✉, Toshihiro Sugiyama 2,✉, Yoshihiko Kawarada 2,✉, Nobuhiko Tanigawa 1,✉
PMCID: PMC5926049  PMID: 10189889

Abstract

Wecomparatively investigated the extent of apoptotic cell loss in human colorectal cancers evaluated by two methods, namely the conventional terminal deoxynucleotidyl transferase (TdT)‐ mediated deoxyuridine triphosphate (dUTP)‐biotin nick end‐labeling (TUNEL) method and immunohistochemistry for single‐stranded (ss) DNA. The apoptotic index (AI) obtained with the TUNEL method was higher than that shown by the immunohistochemistry for ssDNA. However, a significant correlation in AIs evaluated by these methods was found. The AIs obtained by both methods were significantly higher in the advanced cancers than in the early cancers. Cellular proliferation activity was assessed in terms of positivity rate (PR) for expression of proliferating cell nuclear antigen (PCNA). The PR of advanced cancers was significantly higher than that of early cancers. The present results indicate that immunohistochemistry for ssDNA is useful (as is the TUNEL method) for evaluation of apoptotic tumor cells in colorectal carcinomas. In addition, it was confirmed that there is a remarkable increase of not only proliferation activity, but also tumor cell apoptosis in the process of progression of colon cancer from early to advanced stages of the disease.

Keywords: Apoptosis, TUNEL, Immunohistochemistry, Single‐stranded DNA, Colorectal cancer

Full Text

The Full Text of this article is available as a PDF (2.4 MB).

REFERENCES

  • 1.Payne , C. M. and Cromey , D. W.An image analysis workstation designed for multiple users: application of quantitative digital imaging techniques to electron microscopy . Ultrastruct. Pathol. , 16 , 147 – 154 ( 1992. ). [DOI] [PubMed] [Google Scholar]
  • 2.Kerr , J. F. R. , Wyllie , A. H. and Currie , A. R.Apoptosis: a basic biological phenomenon with wide‐ranging implications in tissue kinetics . Br. J. Cancer , 26 , 239 – 257 ( 1972. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3.Compton , M. M.A biochemical hallmark of apoptosis: internucleosomal degradation of the genome . Cancer Metastasis Rev. , 11 , 105 – 119 ( 1992. ). [DOI] [PubMed] [Google Scholar]
  • 4.Gavrieli , Y. , Sherman , Y. and Ben‐Sasson , S. A.Identification of programmed cell death in situ via specific labeling of nuclear DNA fragmentation . J. Cell Biol. , 119 , 493 – 501 ( 1992. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5.Tomei , L. D.Apoptosis: a program death or survival ? In“Apoptosis, The Molecular Basis of Cell Death ” ed. Tomei L. D. and Cope F. , pp. 279 – 316 ( 1991. ). Cold Spring Harbor Laboratory Press; , New York . [Google Scholar]
  • 6.Naruse , I. , Keino , H. and Kawarada , Y.Antibody against single‐stranded DNA detects both programmed cell death and drug‐induced apoptosis . Histochemistry , 101 , 73 – 78 ( 1994. ). [DOI] [PubMed] [Google Scholar]
  • 7.Japanese Society for Cancer of the Colon and Rectum . “Japanese Classification of Colorectal Carcinoma ” pp. 3 – 74 ( 1997. ). Kanehara Co. , Tokyo . [Google Scholar]
  • 8.Thompson , C. B.Apoptosis in the pathogenesis and treatment of disease . Science , 267 , 1456 – 1462 ( 1995. ). [DOI] [PubMed] [Google Scholar]
  • 9.Cohen , J. J. and Duke , R. C.Glucocorticoid activation of a calcium‐dependent endonuclease in thymocyte nuclei leads to cell death . J. Immunol. , 132 , 38 – 42 – 1984. ). [PubMed] [Google Scholar]
  • 10.Fortini , P. , Bigmami , M. and Dogliotti , E.Evidence for AP site formation related to DNA‐oxygen alkylation in CHO cells treated with ethylating agents . Mutat. Res. , 236 , 129 – 137 ( 1990. ). [DOI] [PubMed] [Google Scholar]
  • 11.Laval , J. , Boiteux , S. and O'Connor , T. R.Physiological properties and repair of apurinic/apyrimidinic sites and imidazole ring‐opened guanines in DNA . Mutat. Res. , 233 , 73 – 79 ( 1990. ). [DOI] [PubMed] [Google Scholar]
  • 12.Umemura , S. , Yasuda , M. , Osamura , Y. , Kawarada , Y. , Sugiyama , T. and Tsutsumi , Y.Enhancement of TdTmediated dUTP‐biotin nick end labeling (TUNEL) method using mung bean nuclease, a single‐strand DNA digestion enzyme . J. Histochem. Cytochem. , 44 , 125 – 132 ( 1996. ). [DOI] [PubMed] [Google Scholar]
  • 13.Ansari , B. , Coates , P. J. , Greenstein , B. D. and Hall , P. A.In situ end‐labeling detects DNA strand breaks in apoptosis and other physiological and pathological states . J. Pathol. , 170 , 1 – 8 ( 1993. ). [DOI] [PubMed] [Google Scholar]
  • 14.Grasl‐Kraupp , B. , Ruttkay‐Nedecky , B. , Koudelka , H. , Bukowska , K. , Bursch , W. and Schulte‐Hermann , R.In situ detection of fragmented DNA (TUNEL assay) fails to discriminate among apoptosis, necrosis, and autolytic cell death: a cautionary note . Hepatology , 21 , 1465 – 1468 ( 1995. ). [DOI] [PubMed] [Google Scholar]
  • 15.Desjardins , L. M. and Macmanus , J. P.An adherent cell model to study different stages of apoptosis . Exp. Cell Res. , 216 , 380 – 387 ( 1995. ). [DOI] [PubMed] [Google Scholar]
  • 16.Frankfurt , O. S. , Seckinger , D. and Sugarbaker , E. V.Pleiotropic drug resistance and survival advantage in leukemic cells with diminished apoptotic response . Int. J. Cancer , 59 , 217 – 224 ( 1994. ). [DOI] [PubMed] [Google Scholar]
  • 17.Frankfurt , O. S. , Robb , J. A. , Sugarbaker , E. V. and Villa , L.Monoclonal antibody to single‐stranded DNA is a specific and sensitive cellular marker of apoptosis . Exp. Cell Res. , 226 , 387 – 393 ( 1996. ). [DOI] [PubMed] [Google Scholar]
  • 18.Leoncini , L. , Vecchio , M. T. D. , Megha , T. , Barbini , P. , Galieni , P. , Pileri , S. , Sabattini , E. , Gherlinzoni , F. , Tosi , P. , Kraft , R. and Cottier , H.Correlations between apoptotic and proliferative indices in malignant non‐Hodgkin/rquote slymphomas . Am. J. Pathol. , 142 , 755 – 763 ( 1993. ). [PMC free article] [PubMed] [Google Scholar]
  • 19.Kasagi , N. , Gomyo , Y. , Shirai , H. , Tsujitani , S. and Ito , H.Apoptotic cell death in human gastric carcinoma: analysis by terminal deoxynucleotidyl transferase‐mediated dUTPbiotin nick end labeling . Jpn. J. Cancer Res. , 85 , 939 – 945 ( 1994. ). [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 20.Tatebe , S. , Ishida , M. , Kasagi , N. , Tsujitani , S. , Kaibara , N. and Ito , H.Apoptosis occurs more frequently in metastatic foci than in primary lesions of human colorectal carcinomas: analysis by terminal‐deoxynucleotidyl‐transferasemediated dUTP‐biotin nick end labeling . Int. J. Cancer , 65 , 173 – 177 ( 1996. ). [DOI] [PubMed] [Google Scholar]
  • 21.Hawkins , N. , Lees , J. , Hargrave , R. , O'Connor , T. , Meagher , A. and Ward , R.Pathological and genetic correlates of apoptosis in the progression of colorectal neoplasia . Tumour Biol. , 18 , 146 – 156 ( 1997. ). [DOI] [PubMed] [Google Scholar]
  • 22.Compton , M. M.Laboratory methods: development of an apoptosis endonuclease assay . DNA Cell Biol. , 10 , 133 – 141 ( 1991. ). [DOI] [PubMed] [Google Scholar]

Articles from Japanese Journal of Cancer Research : Gann are provided here courtesy of Wiley

RESOURCES