Cancer Res Treat.  2006 Apr;38(2):99-107.

Protein Expression Profile using Two-Dimensional Gel Analysis in Squamous Cervical Cancer Patients

• Bae SM
• Min HJ
• Ding GH
• Kwak SY
• Cho YL
• Nam KH
• Park CH
• Kim YW
• Kim CK
• Han BD
• Lee YJ
• Kim DK
• Ahn WS

Abstract

PURPOSE: Screening in cervical cancer is now progressing to discover candidate genes and proteins that may serve as biological markers and that play a role in tumor progression. We examined the protein expression patterns of the squamous cell carcinoma (SCC) tissues from Korean women with using two-dimensional polyacrylamide gel electrophoresis (2-DE) and matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometer.
MATERIALS AND METHODS
Normal cervix and SCC tissues were solubilized and 2-DE was performed using pH 3~10 linear IPG strips of 17 cm length. The protein expression was evaluated using PDQuest 2-D software(TM). The differentially expressed protein spots were identified with a MALDI-TOF mass spectrometer, and the peptide mass spectra identifications were performed using the Mascot program and by searching the Swiss-prot or NCBInr databases.
RESULTS
A total of 35 proteins were detected in SCC. 17 proteins were up-regulated and 18 proteins weredown-regulated. Among the proteins that were identified, 12 proteins (pigment epithelium derived factor, annexin A2 and A5, keratin 19 and 20, heat shock protein 27, smooth muscle protein 22 alpha, alpha-enolase, squamous cell carcinoma antigen 1 and 2, glutathione S-transferase and apolipoprotein a1) were protein previously known to be involved in tumor, and 21 proteins were newly identified in this study.
CONCLUSION
2-DE offers the total protein expression profiles of SCC tissues; further characterization of these differentially expressed proteins will give a chance to identify the badly needed tumor-specific diagnostic markers for SCC.

Keyword

Cervical cancer; Squamous cell carcinoma; Two-dimensional polyacrylamide gel electrophoresis; Matrix assisted laser desorption/ ionization-time of flight mass spectrometer

MeSH Terms

Annexin A2
Apolipoproteins
Biomarkers
Carcinoma, Squamous Cell
Cervix Uteri
Databases, Protein
Electrophoresis, Polyacrylamide Gel
Epithelium
Female
Glutathione Transferase
HSP27 Heat-Shock Proteins
Humans
Hydrogen-Ion Concentration
Keratin-19
Mass Screening
Muscle, Smooth
Phosphopyruvate Hydratase
Uterine Cervical Neoplasms*
Annexin A2
Apolipoproteins
Glutathione Transferase
HSP27 Heat-Shock Proteins
Keratin-19
Phosphopyruvate Hydratase

Figure

• 

  Fig. 1 Comparison of the proteome by performing two-dimensional gel electrophoresis on normal tissues and cervical SCC tissues. Representative examples of the 2-DE gels derived from normal cervix tissue and from the cervical SCC tissue. The normal cervix (A) and cervical SCC (B) total proteins were separated by 2-DE with using IPG strips (pH 3~10) in the first dimension and 12% SDS-PAGE in the second dimension. The identified protein spots are indicated by numbers. The down-regulated (A) or up-regulated (B) proteins in the cervical SCC are indicated.

• 

  Fig. 2 Comparison of the protein expressions of the normal samples and the SCC samples. The up-regulated and down-regulated proteins (14-3-3ε, annexin A1 and SCCA-2) were selected and the magnified gel images are presented (A). The differences of expression were statistically meaningful according to the PDQuest 2-D software quantification (p-value <0.05) (B).

• 

  Fig. 3 MALDI-TOF peptide mass fingerprint of the tryptic digest of the tropomyosin beta chain. The MALDI-TOF mass spectrum of tropomyosin beta chain (A) and the matched peptide sequences are underlined (B).

• 

  Fig. 4 Western blot analysis of several proteins. The proteins obtained from the normal cervix control samples (N) and the cervical squamous cell carcinoma samples (C) were subjected to western blot analysis. Equal amounts of proteins (40 µg) were analyzed with SDS-PAGE and then they were Western blotted with anti-SCCA-1, anti-SCCA-2, anti-Hsp27, anti-tropomyosin and anti-annexin 1 antibodies.

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