Raji cells were also susceptible to DRB induced cytotoxicit

Experimental lung metastasis models MCF 7 and MCF 7 DOX cells were injected into the tail vein of Balb KU-55933 構造 c nude mice, Three months after injection, the animals were killed by CO2 inhalation and their lungs were excised. Lung tumor formation was observed and tumor nodules were counted under a dis secting microscope. All animal experiment procedures were approved by the Institutional Animal Care and Use Committee in Korea National Cancer Center. Gelatin and fibrinogen plasminogen zymography The proteolytic activity of MMP 2, MMP 9, and uPA in CM was analyzed by substrate gel electrophoresis using SDS PAGE gels containing 0. 2% gelatin or 0. 12% fibrinogen and plasminogen, CM from each treatment group was concen trated using an Amicon Ultra 4 centrifugal device and loaded onto gels.<br><br> After electrophoresis, the gels were washed with 2. 5% Triton X 100 and incubated overnight in zymogram incubation buffer at 37 C. Clear bands indicative of gela tinolytic activity were visualized by staining the gels with Coomassie blue. Gene expression analyses from whole genome Total RNA was isolated and purified from MCF 7 and MCF 7 DOX cells using purchase Linifanib the TRIzol reagent and RNease Mini kit, Of those, 500 ng RNA was biotinylated and amplified using the Illumina TotalPrep RNA Amplification Kit according to the manufacturers instructions. The cRNA yield was measured using RiboGreen RNA quantitation kit, and 750 ng of the cRNA sample was hybridized on a human HT 12 expression bead chip for profiling 48,804 tran scripts per sample. Bead chips were stained with strep tavidin and scanned using an Illumina BeadArray Reader.<br><br> BeadStudio V3 was used to quantile normalize the data. To find doxorubicin resistant phenotype associated genes, we applied expression data to search and include genes with significant difference in expression levels between MCF 7 and MCF 7 DOX. Gene sets with 2 fold or more difference in LY3009104 1187594-10-0 mRNA level and p value cut off are presented in Table 1. Statistical analysis The effect of doxorubicin or NS398 on breast cancer cell proliferation was analyzed using one way ANOVA followed by Turkeys multiple test, The data of in vitro cancer cell invasion and tumor incidence in the mice were analyzed using Students t test, Results Resistance of MCF 7 DOX cells to doxorubicin Doxorubicin is one of the most commonly used drugs in the treatment of cancer, but its inhibitory effect on cell proliferation varies in several cancer cell lines.<br><br> Therefore, we investigated whether doxorubicin has the same anti proliferative effect in MDA MB 231, MCF 7, MCF 7 DOX, and T 47D cell lines. The cells were treated with the indicated concentrations of doxorubicin for 72 h and cell proliferation was measured using the MTT assay. Doxorubicin inhibited cell proliferation in a concentra tion dependent manner in MCF 7 and T47D cells, and to a lesser extent in MDA MB 231 cells. By contrast, doxorubicin mediated inhibition was significantly reduced in MCF 7 DOX cells, We next mea sured the growth of MCF 7 and MCF 7 DOX cells at lower doxorubicin concentrations and MCF 7 DOX cells were consistently resistant to doxorubicin, We then tested whether the doxorubicin mediated growth inhibition was mediated by apoptosis.