| Background: LKB1/STK11 (serine theronine protein kinase) is a tumor supressive gene newly identified in patients suffered with Peutz-Jeghers syndrome. Till now, a large amount of researches related with tumor suppress gene have been conducted. Some pathological study demonstrated that mutation of LKBl contributes to vascular abnormalities in embryo and up-regulation of VEGF in embryonic fibroblast cells. Also, in 58 percent of those patients with lung carcinoma metastased to the brain, mutation of LKBl gene had been reported. It can be infered from these results that LKB1 may play a role in the growth and metastasis of tumor by regulating the expression of VEGF in tumor cells. Growth and metastasis of tumor depend on the forming of newborn blood vessel, and tumor cells can secrete vascular endothelial growth factor (VEGF) and express its receptor, which accelerate the forming of newborn blood vessel. As a new tumor suppress gene LKB1, the role of which in the development of lung cancer have litte been studied. Therefore, it is of great importance to elucidate the mechanism of tumor growth and metastasis by studying the role of LKBl in regulating the expression of VEGF and other related genes in the lung cancer cells.Objective: To construct the cell models in which expression of LKBl was suppressed by RNA interference technique and to eluciate the role of the LKBl gene in the genesis and development of the lung cancer.Methods: SiRNA-LKBl Mediated by liposome system was transfected into 95D cells, which overexpressed LKBl gene, and stable transfectants were selected. The further study was made on the basis of these cell models. Growth curve and flow cytometry were utilized to analysis the cells proliferation. To elucidate the underlyingmolecular mechanisms, the region of VEGF promoter was analyzed by the chemiluminescence of the double luciferases and bioinformation, then the expression of SP1 and VEGF were detected by RT-PCR in Magnocellular lung cancer cells (95D). Furthermore, some transcription factors and genes related with cell signal pathways were investigated. The expression of the mRNA and the protein of these genes were determined using northern blotting and western blotting.Results: Our results showed that cell models were set up successfully, in which expression of LKB 1 was significantly suppressed by siRNA. The growth of the cultured cells with LKB 1 gene knocked down was proved more vibrant obviously. Meanwhile, the expression of VEGF in lung cancer cells was enhanced. In the analysis of the VEGF promoter region, we found that the activity of the region from -131 to +54 was increased obviously and there were some binding sites for transcription factor SP1. In the further experiment we verified that the changes in the expression between SP1 and VEGF had the similar tendency. By investigating some transcription factors and genes related to cell signal pathways, we found that the activity of MAP kinase pathway was decreased, while, the expression of some genes such as SPK VEGF and c-Myc was increased. Our results also showed that the protein expression of oncogene c-myc increased, but its transaction activity was not activated in Magnocellular lung cancer cells whose expression of LKB 1 was suppressed.Conclusions: (l)Cell models were set up successfully by RNA interference technique, which expression of LKB 1 was significantly suppressed. A series of studies were carried out, which set a technical foundation for gene study. (2) LKB1 plays an important role in the regulation negatively of the lung cancer cell proliferation and the inhibition of the tumor hyperplasy, which didn't activate the MAPK pathway but other ones perhaps did. (3) LKB1 gene regulated the expression of VEGF might be partly depended on the activity of SP1. (4)LKB1 gene might play a vital role in the process of the genesis and development of lung tumor through regulating SPK VEGF and c-Myc gene.
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