Proteomic identification of tumor biomarkers associated with primary gallbladder cancer.
OBJECTIVE
To identify potential biomarkers of primary gallbladder cancer (PGC).


METHODS
Fresh PGC, cholecystitis and normal gallbladder tissue specimens collected from 10 patients, respectively, were subjected to comparative proteomic analysis. The proteomic patterns of PGC were compared with those of cholecystitis and normal gallbladder tissues using two-dimensional gel electrophoresis (2-DE). The differentially expressed proteins were then identified using a MALDI-TOF mass spectrometer (MS) and database searches. To further validate these proteins, 20 samples of PGC tissues and normal tumor-adjacent tissues were collected for Western blot, quantitative real-time PCR, and immunohistochemical staining assay.


RESULTS
Seven differentially expressed protein spots were detected by 2-ED analysis by comparing the average maps of PGC, cholecystitis and normal gallbladder tissues. Six of the seven differentially expressed proteins were identified using MALDI-TOF MS, with three overexpressed and three underexpressed in PGC tissue. Protein levels of annexin A4 (ANXA4) were significantly elevated, and heat shock protein 90-beta (Hsp90β) and dynein cytoplasmic 1 heavy chain 1 (Dync1h1) were decreased in PGC tissues relative to the normal tumor-adjacent tissues as shown by Western blot analysis. However, levels of actin, aortic smooth muscle and gamma-actin were unchanged. In addition, the mRNA levels of all 5 proteins showed similar changes to those of the protein levels (P < 0.01). Further validation by immunohistochemical analysis showed the upregulated expression of ANXA4 and decreased expression of Hsp90β and Dync1h1 in the cytoplasm of PGC tissues relative to the normal tumor-adjacent tissues.


CONCLUSIONS
Three proteins are identified as potential biomarkers of PGC using proteomic analysis. The functions of these proteins in the carcinogenesis of PGC remain to be studied.
Hai-Lin Huang, Department of General Surgery, Hospital of China Armed Police Forces, Nanchang 330030, Jiangxi Province, China.
Hai-Lin
HL
Huang
Hai-Lin Huang, Department of General Surgery, Hospital of China Armed Police Forces, Nanchang 330030, Jiangxi Province, China.
Hou-Shan
HS
Yao
Hai-Lin Huang, Department of General Surgery, Hospital of China Armed Police Forces, Nanchang 330030, Jiangxi Province, China.
Yi
Y
Wang
Hai-Lin Huang, Department of General Surgery, Hospital of China Armed Police Forces, Nanchang 330030, Jiangxi Province, China.
Wei-Jun
WJ
Wang
Hai-Lin Huang, Department of General Surgery, Hospital of China Armed Police Forces, Nanchang 330030, Jiangxi Province, China.
Zhi-Qian
ZQ
Hu
Hai-Lin Huang, Department of General Surgery, Hospital of China Armed Police Forces, Nanchang 330030, Jiangxi Province, China.
Kai-Zhou
KZ
Jin
D000368:Aged; D014408:Biomarkers, Tumor / Q000032:analysis* / Q000235:genetics; D015153:Blotting, Western; D016022:Case-Control Studies; D030562:Databases, Protein; D015180:Electrophoresis, Gel, Two-Dimensional; D005260:Female; D005706:Gallbladder Neoplasms / Q000737:chemistry* / Q000235:genetics / Q000473:pathology; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D008875:Middle Aged; D009363:Neoplasm Proteins / Q000032:analysis* / Q000235:genetics; D011237:Predictive Value of Tests; D040901:Proteomics / Q000379:methods; D012333:RNA, Messenger / Q000032:analysis; D060888:Real-Time Polymerase Chain Reaction; D015203:Reproducibility of Results; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D014408:Biomarkers, Tumor; D009363:Neoplasm Proteins; D012333:RNA, Messenger
Annexin A4; Biomarker; Dynein cytoplasmic 1 heavy chain 1; Heat shock protein 90-beta; Primary gallbladder cancer; Proteomic analysis