A Study On The Effects Of Nm23-H1 Gene On The Cytoskeleton And Biological Behavior In Human Lung Cancer Cell Line L9981

Tumor metastasis is not only the malignant marker and charateristcs of lung cancer, but also the key cause of failure to cure and high mortality in lung cancer. At present, it has been known that lung cancer metastasis was controlled by many genes, especially the tumor metastatic suppressive gene. nm23 gene was discovered in 1988 as a first and the most important tumor metastatic suppressive gene. It has been proved that a close correlation existed between the high metastatic rate and the low expression or alleic deletion of nm23-H1 gene by DNA, mRNA and protein levels in lung cancer. The lung cancer with nm23-H1 gene deletion is always combined with abnormal expression ofsome other metastastic-related genes. Dr. Zhou Qinghua has hypothesized that nm23-H1 gene may be an upstpeam regulatory gene of metastatic-related genes and it can regulate the downstream genes to inhibit metastasis. In order to explore the effect of nm23-Hl gene on cytoskeleton and biological behavior of human lung cancer and the molecular mechanisms regulating the cytoskeleton of lung cancer cell, a human high metastasis lung cancer cell line L9981 with nm23-Hl gene deletion has been screened and identified from 9 human lung cancer lines; a expression plasmid of pLXSN-nm23-Hl-EGFP with EGFP and wild nm23-Hl gene has been successfully constructed and transfected into the L9981 cell line, so as to observe the changes of cytoskeleton and biological behavior of the L9981 cell line, and the molecular mechanisms before and after transfecting with nm23-H1 gene. The results showed that:1. A human high metastasis lung cancer cell line L9981 with nm23-H1 gene deletion was screened and identified from 9 human lung cancer lines.2. A retrovirus eukaryocytic expression vectorpLXSN-nm23-H1-EGFP was successfully constructed through gene engineering techniques. DNA sequencing result showed that the terminative code of nm23-Hl gene in pLXSN-nm23-H1-EGFP has been mutated from ATG to TGA. The fusion protein of nm23-Hl-EGFP is stablely, continuously and high efficiently expressed in transfected L9981 cell line.3 .Trans fecti on of nm23-H1 gene can remarkably decrease the refraction , projection and adhesion ability of the L9981 lung cancer cell line.4.The vitro-invasion of L9981 lung cancer cell line was significantly decreased after transfecting with nm23-Hl gene.5.Transfection of nm23-H1 gene can significantly reduce the aggregation of microfilament, increase the polymerization of tubulin and also decrease the motility of the L9981 cell line.6.Transfection of nm23-Hl gene can remarkably downregulate the expression of integrin αv and a-actin in the L9981 cell line, but the same effect was not observed in the beta -tubulin expression.Conclusions: (1) The human high metastasis lung cancer cell line L9981 has deletion of nm23-H1 gene. (2) nm23-H1 gene canremarkably decrease the ability of vitro adhesion and invasion , reduce the aggregation of microfilament and increase the polymerization of tubulin. (3) The molecular mechanisms which nm23-H1 regulate the cytoskeleton of human lung cancer cell line L9981 might be related to the downregulation of integrinαv and α-actin expression.