Determination of S-phase cells by in situ hybridization for histone H3 mRNA in hepatocellular carcinoma: correlation with histologic grade and other cell proliferative markers

Mod Pathol. 1996 Feb;9(2):99-104.


Cell proliferation has a crucial importance in biologic potentialities of tumor cells. Several methods to measure S-phase fraction in tumor samples have been developed, with considerable limitations in practical applications. Histones are nucleosomal proteins that are responsible for packaging chromosomal DNA into nucleosomes. The expression of histone genes is a fundamental step constituting the process of cell proliferation. Because histone H3 mRNA accumulates in the cytoplasm during S-phase, and then decreases as cells approach G2-phase, demonstration of histone H3 mRNA expression in a tumor cell population may represent responsible S-phase fraction. Thus, we have assessed S-phase fraction by nonisotopic in situ hybridization for histone H3 mRNA in paraffin sections of hepatocellular carcinomas (HCCs); then, we compared this with the histologic grades and other cell proliferative markers. In contrast to radioisotopic detection, signals were distinctly visualized, and quantitation of the stained-cells was easily carried out. Histone H3 labeling index (LI) significantly correlated with other cell proliferative markers, including Ki-67 (MIB-1) and PCNA immunostainings, and mitotic index. In addition, a statistically significant correlation was seen between histone H3 LI and histologic grades of differentiation, i.e., histone H3 LI being higher in less-differentiated HCCs. Furthermore, histone H3 LI/Ki-67 LI ratio was significantly higher in the moderately and poorly differentiated subtypes (including one case of the undifferentiated subtype) than in well-differentiated HCCs. By using this nonisotopic in situ hybridization technique, it becomes possible to assess rapidly, retrospectively, safety, and easily a malignant potentiality of HCCs in paraffin-embedded tissues.

MeSH terms

  • Adult
  • Aged
  • Biomarkers, Tumor / analysis*
  • Carcinoma, Hepatocellular / genetics*
  • Carcinoma, Hepatocellular / pathology*
  • Cell Differentiation
  • Cell Division
  • Female
  • Histones / genetics*
  • Humans
  • In Situ Hybridization
  • Liver Neoplasms / genetics*
  • Liver Neoplasms / pathology*
  • Male
  • Middle Aged
  • RNA, Messenger / analysis*
  • S Phase / genetics*


  • Biomarkers, Tumor
  • Histones
  • RNA, Messenger