ABSTRACT
<p><b>OBJECTIVE</b>To investigate the effect of osteopontin silencing on the invasion and apoptosis of U251 cells.</p><p><b>METHODS</b>The invasion, apoptosis and levels of uPA, MMP-2 and MMP-9 were determined by invasion assay, flow cytometry, Western blot and real-time fluorescence quantitative PCR respectively.</p><p><b>RESULTS</b>Osteopontin small interference RNA (siRNA) inhibited osteopontin expression and cell invasion, promoted apoptosis in U251 cells. In addition, the expression of Bcl-2, uPA, MMP-2 and MMP-9 was decreased, while Bax level was elevated.</p><p><b>CONCLUSION</b>Osteopontin siRNA can inhibit U251 cells invasion via the down-regulation of uPA, MMP-2 and MMP-9 levels, and promote apoptosis through induction of Bax expression and inhibition of Bcl 2 level. It suggests that osteopontin plays an important role in human glioma progression.</p>
Subject(s)
Humans , Apoptosis , Cell Line, Tumor , Gene Expression Regulation, Neoplastic , Gene Silencing , Gene Transfer Techniques , Genetic Vectors , Genetics , Metabolism , Glioma , Genetics , Metabolism , Pathology , Lentivirus , Genetics , Metabolism , Neoplasm Invasiveness , Osteopontin , Genetics , Metabolism , RNA, Small Interfering , GeneticsABSTRACT
<p><b>OBJECTIVE</b>To study inhibitory efficacy of combined gene therapy for malignant gliomas transfected with antisense human telomerase reverse transcriptase (hTERT)/PTEN in vitro and in vivo.</p><p><b>METHODS</b>To construct two adenovirus recons which contained antisense hTERT and wild-type PTEN respectively with high performance homologous recombination system in bacteria. The two adenovirus recons were transfected into U251 human malignant glioma cells combinedly or respectively in vitro and in vivo. U251 cell proliferation in vitro was determined by MTT assay and flow cytometry, tumor growth in vivo was measured by the volume of glioma in nude mice. Telomerase activity was detected by telomeric repeat amplification protocol (TRAP) assay. Expression of hTERT and PTEN protein was detected by Western blotting methods.</p><p><b>RESULTS</b>After transfection in vitro, the growth of U251 cells was inhibited significantly. The inhibitory effect was time-dependent. The strongest inhibition was observed in combined transfection group, at the 6th day, the survival rate was 37.6%, telomerase activity (only 28.8TPG) was inhibited significantly, hTERT protein expression was inhibited significantly too, which was 0.2106, but PTEN protein expression was increased significantly, which was 0.9630. In vivo, the growth of tumors was also effectively inhibited.</p><p><b>CONCLUSION</b>Growth of malignant glioma cells is effectively inhibited after transfection with combined antisense hTERT and PTEN in vitro and in vivo.</p>