The Study Of Increased Sensitivity To Cisplatin By Nm23-H1

Since the initial description of the nm23 gene as an antimetastatic factor whose expression is correlated inversely with tumor metastatice potential in murine melanoma cell lines. Numerous studies have supported its suppressive effects on tumor metastatasis. Recently, the role of nm23-Hl in sensitivity to anticancer agents has attracted a great deal of attention. In addition to the antimetastastic property, nm23-Hl has been showen to be associated with sensitivity to cisplatin in breast carcinoma.Firstly, to transfect nm23-Hl into Tca8113 cell line using Lepfect liposomal transfection reagent and get stable Tca8113 cell line. Detect the protein expression of nm23-Hl in stable cell line using immunohistochemistry, western-blot and flow cytometer. we get pCMV-Bam- neo-nm23-Hl successfully and establish the stable expression of nm23-Hl. It is showed that 986bp of nm23-Hl is inserted into pCMV-neo-Bam vector by electrophoresis on a 1% agarose gel and stained with ethidium bromide. nm23-Hl expression was increased in the stable expression cell line by immunohistochemistry, western-blot and flow cytometer.Secondly, Using MTT, flow cytometer and VG PQ Excell, we detectthe changes of cell mortality rate, mitochondrial membrane potential, apoptosis and intracelleular platinum (Pt) by nm23-H13. we found that, in vitro, the cell mortality rate, apoptosis and intracellular platinum are increased significantly in stable expression cell line of nm23-Hl, comparing with Tca8113 cell line of non-transfection; this effect can be inhibited by oubain which is an inhibitor of Na(+)/K(+)-ATPase. mitochondrial membrane potential is decreased significantly in stable expression cell line of nm23-Hl.Thirdly, to establishing nude mice xenotransplanted model of Tca8113 cell, we detect the changes of tumor volume and weight by nm23-Hl and cisplatin. In vivo, the therapy of nm23-Hl plasmid and cisplatin can inhibit significantly the growth of tumor which include the volume and weight in nude mice xenotransplanted model, comparing with non-therapy group and therapy of cisplatin group.we establish the stable expression cell line of nm23-Hl. nm23-Hl can increase the chemosensitivity of cisplatin significantly; the mechanism is correlated with the activity of Na(+)/K(+)-ATPase. The therapy of nm23-Hl plasmid and cisplatin can inhibit the growth of tumor in nude mice. All thesis results are important for the combination of gene and chemotherapy to use in clinics.