| Nowadays, lung cancer has become one of the malignant tumorsthat have the highest incidence and mortality. As a major disease which isthreatening the whole human society, the incidence of lung cancer hasreached as high as 12.6％of new cancer patients, accounted for 17.8％ofcancer mortality, and is still increasing gradually. According to theWHO's estimation, China will become one of the countries that have therelatively high incidence of lung cancer in the 21century. Lung cancercan be divided tow major groups according to the histopathology, smallcell lung carcinoma (SCLC) and non-small cell lung carcinoma (NSCLC).NSCLC comprises a broad spectrum of tumors that include squamouscarcinoma, adenocarcinoma and large-cell carcinoma according tohistological diagnosis. Lung adenocarcinoma is one of the most commonhistology types of lung cancer; the carcinogenesis of lungadenocarcinoma is a complex process involving multiple events and steps.Though some molecular pathogenesis studies on lung adenocarcinomahave been undertaken successfully on the gene (DNA) and transcription(mRNA) levels, the carcinogenic mechanism is still unclear. At present,there is still no the special lung adenocarcinoma molecular marker for an"early-stage" diagnosis and prognosis evaluation. Furthermore,functional complexity of normal and neoplastic tissues is not dictatedsolely by the encoding genomic information, but also determined byexecution molecules, proteins. The proteomic study of lungadenocarcinoma may directly and comprehensively explore thecarcinogenic mechanism of lung cancer. Thus, the proteomic approach isessential to understand pathogenesis and development of lungadenocarcinoma, it is expected to obtain some clues to further study of the carcinogenic mechanisms, diagnosis, treatment and new drugexploitation of lung adenocarcinoma.This study was designed to use comparative proteomics technologyto find the differential expressed proteins between human lungadenocarcinoma and paired normal tumor-adjacent lung tissues, thenvalidated with the immunohistochemistry technology based on clinicalsample. The research was optimized to establish the two-dimensionalelectrophoresis profiles with high resolution and reproducibility fromhuman lung adenocarcinoma tissue and paired normal tumor-adjacentlung tissue, characterized the differential expressed spots between 2-DEprofiles by 2-DE image analysis software and identified the differentialexpressed proteins with two kinds of different mass spectrometries. Onthe other hand, the immunohistochemistry technology was used tovalidate the differential expressed proteins with clinical samples.In the first place, comparative proteome analysis with twenty casesof human lung adenocarcinoma tissues and the paired normal lung tissuesadjacent to tumor were carried through. Namely, the total proteins ofhuman lung adenocarcinoma tissues and paired normal tumor-adjacentlung tissues were extracted with 2D Quant kit and seperated by means ofimmobilized pH gradient-based two-dimensional gel electrophoresis(2-DE) and Commassie Blue staining. The differential expressed proteinswere analyzed with 2-DE image analysis software and then identified bymatrix-assisted laser desorption/ionization time of flight massspectrometry (MALDI-TOF-MS) and quadrupole time of flight massspectrometry (Q-TOF-MS/MS). RESULTS: (1) The results showed thatwell-resolved, highly reproducible 2-DE patterns of human lungadenocarcinoma and paired normal tumor adjacent lung tissues were obtained with 24cm IPGstrips (pH3-10).The average spots of 3 gels were1052±48, 1037±57 spots were matched among tumor tissues andnormal lung tissues. The average position deviation of matched spots was0.814±0.103 mm in IEF direction and 1.052±0.185 mm in SDS-PAGEdirection. (2) PDQUEST 2D image analysis software was used toanalysis twenty cases of lung adnocarcinoma and paired normal lungtissues, average matched spots were 1006±, twenty eight differentialexpressed spots were screened. (3) Twenty non-redundant differentialexpressed proteins were identified by MALDI-TOF-MS andQ-TOF-MS/MS, thirteen proteins were up regulated and Severn proteinswere down regulated, some proteins were involved in the regulation ofcell signal transduction or metabolized enzyme of cell.To validate the results screened by proteome research,immunohistochemistry technology was used to validate several lungadnocarcinoma differential expressed proteins included 14-3-3 sigma,annexin 1 and superoxide dismutase. The immunohistochemistry resultsshowed that 14-3-3 sigma, annexin 1 and superoxide dismutase wereup-regulated in lung adenocarcinoma, P＜0.05.In this study, compratative proteomics technologies were systemicallyperformed in the lung adenocarcinoma and paired normal tumor-adjacentlung tissues, twenty non-redundant lung adenocarcima related proteinswere successfully identified with MALDI-TOF-MS and Q-TOF-MS/MSmass spectrometry. 14-3-3 sigma, annexin 1 and superoxide dismutasewere validated by immunohistochemistry technology with clinicalsamples. The results showed that 14-3-3 sigma, annexin 1 and superoxidedismutase were really differential expressed between lungadenocarcinoma and paired normal lung tissues. These results will provide scenticfic foundation for screening the molecular biomarker usedto diagnose and treat lung adenocarcinoma, as well as to elevate thepatient's prognosis and provide new clue for the research of human lungadenocarcinoma.
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