[Effect of overexpression of Smad7 gene on cell proliferation].

OBJECTIVE: To study the effect of overexpression of Smad7 gene on cell proliferation in human bronchial epithelial cell lines. METHODS: Human bronchial epithelial cell lines, BEP2D and BERP35T2 cells, were cotransfected with the mammalian expression vectors PCISmad7.neo and pMyc-SEAP, the latter was ac-myc cis-acting enhancer element fused with alkaline phosphatase (SEAP) reporter gene. Expression of c-myc, p15 and p21 mRNA was detected by RT-PCR before and after stable transfection of Smad7 into BEP2D and BERP35T2 cells in order to study the regulation of TGF-beta-mediated growth inhibition. RESULTS: After BEP2D and BERP35T2 cells transfected with Smad7, the transcriptional activity of c-myc was significantly increased. Smad7 overexpressing cells showed upregulation of c-myc expression and downregulation of p15 and p21 expression, which contributed to the loss of TGF-beta responses in these cells. CONCLUSION: Overexpression of Smad7 may facilitate cell proliferation by antagonizing TGF-beta-mediated antiproliferative gene responses.

[Expression and clinical significance of methylthioadenosine phosphorylase gene in patients with non-small cell lung cancer].

OBJECTIVE: To investigate the change of expression of methylthioadenosine phosphorylase (MTAP) gene in patients with non-small cell lung cancer and its clinical significance. METHODS: Thirty fresh samples of cancer with adjacent tissues were collected from 15 patients with lung cancer, 12 males and 8 females, aged 53.6 (38 approximately 72), 8 with squamous cell carcinoma and 7 with adenocarcinoma. The expressions of MTAP mRNA and of its protein were analyzed by RT-PCR and Western blotting. RESULTS: In 11 out of the 15 tumor samples (73.3%) MTAP mRNA was not expressed or expressed only after the additional 5 circulations. However, the MTAP mRNA expression rate was 93.3% (14/15) in the adjacent tissues. The result in the 11 samples with none or low MTAP mRNA expression was confirmed by Western blot analysis. The low expression rates were not significantly different between lung adenocarcinoma and squamous cell carcinoma. But the low expression rate of MTAP in intermediate and poorly differentiated lung cancer was significantly higher than that in well-differentiated cancer. CONCLUSION: The low expression or loss of MTAP gene may be relevant closely to the differentiation degree in lung cancer.

[Proteomics-based identification of Maspin differential expression in bronchial epithelial immortalized cells and malignant transformation cells].

BACKGROUND & OBJECTIVE: Maspin, a serepin inhibitor, plays a key role in tumor growth and metastasis. The aim of this study was to identify the differential expression of Maspin in malignant transformation process of bronchial epithelial cells by proteomics. METHODS: Functional proteomics analysis of Maspin on bronchial epithelial immortalized cells and malignant transformation cells was carried out using immobilized pH gradient (IPG) two-dimensional electrophoresis, peptide mass fingerprinting (PMF), and post source decay (PSD) of bio-mass spectrometry. RESULTS: Nearly 1500 expressed proteins profile on bronchial epithelial immortalized cells and malignant transformation cells were obtained in the range of MW 14.4-94 kDa, PI 3-10. Image analysis showed that Maspin was down-regulated in malignant transformation cells compared with that in immortalized cells. Northern blot analysis showed that the mRNA abundance of Maspin in malignant transformation cells was much lower than that in immortalized cells. CONCLUSION: Alteration expression of Maspin at transcription and translation levels might be involved in carcinogenesis of lung.

[Regulation of Smad7 gene by TGF-beta 1 in process of malignant transformation].

BACKGROUND & OBJECTIVE: Escape from transforming growth factor-beta(TGF-beta)-induced inhibition of growth and proliferation may contribute to tumorigenesis. Smad7 is inhibitory Smads of TGF-beta s signal transduction pathway and prevents TGF-beta signaling. The disorder of Smad7 may lead to the perturbation of TGF-beta signal pathway. In this study, The authors analyzed the expression of Smad7 mRNA and the regulation of Smad7 gene by TGF-beta 1 in the process of malignant transformation of BEP2D cells to investigate the mechanism of cells malignant transformation. METHODS: Cells were cultured and stimulated with TGF-beta 1 followed by RNA extraction. Purified total RNA from TGF-beta 1 treated cells and untreated controls and performed an expression analysis with a human Smad7-specific probe applying Northern blot. As a loading control for the Northern experiment, the membrane was hybridized with a human glyceraldehyde-3-phosphate dehydrogenase(GAPDH) probe. Proteins were extracted from BEP2D and BERP35T-2 cells, then perform Western blot to examine the expression level of TGF-beta 1. RESULTS: Before stimulation with TGF-beta 1, the expression level of Smad7 in the BERP35T-2 cells were higher than that in the BEP2D cells. When stimulated with TGF-beta 1, Smad7 expression levels was upregulated evidently in BEP2D cells, but not significant in BERP35T-2 cells. The expression level of endogenetic TGF-beta 1, BERP35T-2 cells was a little higher than BEP2D cells. CONCLUSION: Over expression of Smad7 mRNA and down-regulation of the cells' responsiveness to TGF-beta 1 in human lung cancer cell line which induced by alpha-particles should be one of the mechanism of radiation induced lung cancer.