[Improvement of protein preparation methods for bronchial epithelial tissues and establishment of 2-DE profiles from carcinogenic process of human bronchial epithelial tissues].
OBJECTIVE
To optimize the protein preparation methods for bronchial epithelial tissues, to establish two-dimensional gel electrophoresis profiles from different stages tissues of human bronchial epithelial carcinogenic process, and to provide a base for identifying carcinogenesis-associated proteins of lung squamous carcinoma.


METHODS
After obtaining samples of the normal-metaplasia-dysplasia-carcinoma tissues of human bronchial epithelia, improved deoxycholate-trichloroaetic acid (DOC-TCA) precipitation was used to extract and purify the total proteins of bronchial epithelial samples. Immobilized pH gradient two-dimensional polyacrylamide gel electrophoresis (2-DE) was used to seperate the total proteins of the samples. After silver staining, ImageMaster 2-DE image analysis software was applied to analyze 2-DE images.


RESULTS
The total proteins extracted with improved DOC-TCA precipitation was used to perform 2-DE. 2-DE patterns with high resolution and reproducibility from different stages were obtained. The average spots for normal epithelium, metaplasia, dysplasia and invasive carcinoma were 1190 +/- 63, 1 227 +/- 69, 1272 +/- 71 and 1326 +/- 82, respectively. One of the squmous metaplasia tissues was chosen and repeated 2-DE for 3 times. Averagely 1216 +/- 75 spots were detected among the 3 gels of metsplasia tissues and 1082 +/- 67 spots were matched. The average matching rate was 89.3% and protein spots in the 3 gels had a good reproducibility. The average position deviation of matched spots in different gels was 0.835 +/- 0.247 mm in IEF direction, and 0.921 +/- 0.104 mm in SDS-PAGE direction.


CONCLUSIONS
Improved DOC-TCA precipitation is a proper method for protein sample preparation of bronchial epithelial tissues. The well-resolved, reproducible 2-DE profiles from different stage tissues of human bronchial epithelial carcinogenic process have been primarily established.
Wu
Xiao-Ying
XY
Key Laboratory of Cancer Proteomics of Ministry of Health, Xiangya Hospital, Central South University, Changsha 410008, China.
Xiao
Zhi-Qiang
ZQ
Chen
Zhu-Chu
ZC
Li
Cui
C
Li
Jian-Ling
JL
Feng
Xue-Ping
XP
Yi
Hong
H
Li
Mao-Yu
MY
D001980:Bronchi / Q000378:metabolism / Q000473:pathology*; D002294:Carcinoma, Squamous Cell / Q000378:metabolism / Q000473:pathology; D015180:Electrophoresis, Gel, Two-Dimensional; D004848:Epithelium / Q000473:pathology*; D006801:Humans; D008175:Lung Neoplasms / Q000378:metabolism* / Q000473:pathology; D008679:Metaplasia / Q000378:metabolism / Q000473:pathology; D009363:Neoplasm Proteins / Q000378:metabolism; D011230:Precancerous Conditions / Q000473:pathology*; D025941:Protein Interaction Mapping; D011506:Proteins / Q000378:metabolism*; D020543:Proteome / Q000378:metabolism
D009363:Neoplasm Proteins; D011506:Proteins; D020543:Proteome