Hepatocellular carcinoma (HCC) is one of the most frequent visceral neoplasia worldwide and is a multifactorial and multistage pathogenesis that finally leads to the deregulation of cell homeostasis. Laser capture microdissection (LCM) may allow a more ready identification of differences in protein expression in selected cell types or areas of tissue, and microscopic regions as small as 3-5 microm in diameter can be sampled. Here we applied the LCM to the proteomic study of hepatitis B-related HCC and surrounding non-tumor tissues. Proteome alterations were observed using 2-DE and ESI-MS/MS, and alterations in the proteome were examined. Twenty protein spots were selected, of which 11 proteins were significantly altered in the HCC compared with the surrounding non-tumor tissues. Of the proteins that were selected, peroxiredoxin 2, apolipoprotein A-I precursor, 3-hydroxyacyl-CoA dehydrogenase type II, and 14.5-kDa translational inhibitor protein appear to be novel candidates as useful hepatitis B-related HCC markers. This study indicates that LCM is a useful technological method in the proteomic study of cancer tissue. The proteins revealed in this experiment can be used in the future for studies pertaining to hepatocarcinogenesis, or as diagnostic markers and therapeutic targets for HCC associated with hepatitis B virus infection.
Laser capture microdissection (LCM) is a powerful tool that enables the isolation of specific cell types from tissue sections, overcoming the problem of tissue heterogeneity and contamination. We combined the LCM with isotope-coded affinity tag (ICAT) technology and two-dimensional liquid chromatography to investigate the qualitative and quantitative proteomes of hepatocellular carcinoma (HCC). The effects of three different histochemical stains on tissue sections have been compared, and toluidine blue stain was proved as the most suitable stain for LCM followed by proteomic analysis. The solubilized proteins from microdissected HCC and non-HCC hepatocytes were qualitatively and quantitatively analyzed with two-dimensional liquid chromatography tandem mass spectrometry (2D-LC-MS/MS) alone or coupled with cleavable isotope-coded affinity tag (cICAT) labeling technology. A total of 644 proteins were qualitatively identified and 261 proteins were unambiguously quantified. These results showed that the clinical proteomic method using LCM coupled with ICAT and 2D-LC-MS/MS can carry out not only large-scale but also accurate qualitative and quantitative analysis.
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