Quantitative Proteomic Study Of Laser Capture Microdissection Purified Normal Human Bronchial Epithelia And Human Lung Squamous Carcinoma

Background:Lung cancer, the morbidity and mortality in the first rank of human malignancies, is cause of cancer-related deaths worldwide. When the first treatment, almost invasion or metastasis. Proteomics provide a new opportunity to reveal the carcinogensis, invasion of lung cancer, analysis of the protein composition, expression and modification, that's possible to identificate the cancer-related proteins, discuss the carcinogenic molecular mechanism of HLSC, provide scientific foundation for the identification of cancer-related protein. Looking for early diagnosis of the cancer is very important for improving the survival rate and cure rate, is positive for diagnosis and treatment for lung cancer.Methods:Proteomic analysis was performed to identify the differential expression proteins between HLSC and NHBE. The differential expression proteins (CKB,HSBP1,SCCA1,S100A8,S100A9) were verified by western blot in HLSC and NHBE and tissue microarray in 100 clinical samples. NCI-H520 cells were stably transfected with plasmids that constitutively expressed sense CKB, and with empty vector. MTT assay, flow cytometry(FCM) analysis were performed to examine the effect of CKB expression on proliferation, cell cycle distribution of stably transfected cells.Results:(1) To establish the differential protein profile of HLSC and NHBE, A total of 96 differential expression proteins in the LCM-purified HLSC and NHBE was identified. Compared with NHBE, 49 proteins were up-regulated and 47 proteins were down-regulated in HLSC. Differential expression proteins were cytoplasmic proteins mainly, then nuclear proteins, secreted proteins and mitochondrial protein, Consist of structural proteins, metabolic enzymes, signal transduction proteins, cell adhesion and proliferation-related proteins. (2) HSBP1,S100A8,S100A9 were up-regulated and CKB,SCCA1 were down-regulated in HLSC tissue compared with NHBE tissues. They were confirmed by Western blot analysis and tissue microarray, and were consistent with the results of quantitative proteomics. (3) CKB expression level was lower in NCI-H520 cells than HBE cells confirmed by Western blot analysis. Then NCI-H520 cells stably transfected with sense CKB expression vector. (4) Up-regulation o of CKB expression in NCI-H520 cells could increase the number of G0/G1 phase cells, inhibit cell proliferation and induce apoptosis.Conclusion:A stable isotope labeled strategy using iTRAQ, followed by 2D-LC/Q-STAR mass spectrometry was performed to separate and identify 96 differential expression proteins.49 proteins were up-regulated and 47 proteins were down-regulated in HLSC. HSBP1/CKB/S100A8/S100A9/SCCA1 were confirmed by Western blot analysis and tissue microarray. CKB in NCI-H520 cells could increase the number of G0/G1 phase cells, inhibit cell proliferation and induce apoptosis.The results will provide scientific foundation for studying the carcinogenic mechanism of HLSC.