Aim： To investigate the effect of inhibition of telomerase with human telomerase reverse transcriptase （hTERT） antisense on tumor necrosis factor-α （TNF-α-induced apoptosis in prostate cancer cells （PC3）. Meth...Aim： To investigate the effect of inhibition of telomerase with human telomerase reverse transcriptase （hTERT） antisense on tumor necrosis factor-α （TNF-α-induced apoptosis in prostate cancer cells （PC3）. Methods： Antisense phosphorothioate oligodeoxynucleotide （AS PS-ODN） was synthesized and purified. Telomerase activity was measured using the telomeric repeat amplification protocol （TRAP） and polymerase chain reaction enzyme-linked immunoassay （PCR-ELISA）. hTERT mRNA was measured by reverse transcription PCR （RT-PCR） assay and gel-image system, hTERT protein was detected by immunochemistry and flow cytometry. Cell viability was detected by 3-（4, 5-dimethylthiazol-2-yl）-2,5-Diphenyltetrazolium （MTT） assay. Cell apoptosis was observed by morphological method and determined by flow cytometry. Results： The telomerase activity decreased with time after hTERT AS PS-ODN treatment. The levels of hTERT mRNA decreased with time after hTERT AS PS-ODN treatment, which appeared before the decline of the telomerase activity. The percentage of positive cells of hTERT protein declined with time after hTERT AS PS-ODN treatment, which appeared after the decline of hTERT mRNA. There was no difference in telomerase activity, hTERT mRNA and protein levels between hTERT sense phosphorothioate oligodeoxynucleotide （S PS-ODN） and the control group. The cell viability decreased with time after hTERT AS PS-ODN combined with TNF-α treatment. The percentage of apoptosis increased with time after hTERT AS PS-ODN combined with TNF-α treatment. There was no difference in cell viability and the percentage of apoptosis between hTERT S PS-ODN and the control group. Conclusion： hTERT AS PS-ODN can significantly inhibit telomerase activity by downregulating the hTERT mRNA and protein expression, and inhibition of telomerase with hTERT antisense can enhance TNF-α- induced apoptosis of PC3 cells.展开更多
文摘Aim： To investigate the effect of inhibition of telomerase with human telomerase reverse transcriptase （hTERT） antisense on tumor necrosis factor-α （TNF-α-induced apoptosis in prostate cancer cells （PC3）. Methods： Antisense phosphorothioate oligodeoxynucleotide （AS PS-ODN） was synthesized and purified. Telomerase activity was measured using the telomeric repeat amplification protocol （TRAP） and polymerase chain reaction enzyme-linked immunoassay （PCR-ELISA）. hTERT mRNA was measured by reverse transcription PCR （RT-PCR） assay and gel-image system, hTERT protein was detected by immunochemistry and flow cytometry. Cell viability was detected by 3-（4, 5-dimethylthiazol-2-yl）-2,5-Diphenyltetrazolium （MTT） assay. Cell apoptosis was observed by morphological method and determined by flow cytometry. Results： The telomerase activity decreased with time after hTERT AS PS-ODN treatment. The levels of hTERT mRNA decreased with time after hTERT AS PS-ODN treatment, which appeared before the decline of the telomerase activity. The percentage of positive cells of hTERT protein declined with time after hTERT AS PS-ODN treatment, which appeared after the decline of hTERT mRNA. There was no difference in telomerase activity, hTERT mRNA and protein levels between hTERT sense phosphorothioate oligodeoxynucleotide （S PS-ODN） and the control group. The cell viability decreased with time after hTERT AS PS-ODN combined with TNF-α treatment. The percentage of apoptosis increased with time after hTERT AS PS-ODN combined with TNF-α treatment. There was no difference in cell viability and the percentage of apoptosis between hTERT S PS-ODN and the control group. Conclusion： hTERT AS PS-ODN can significantly inhibit telomerase activity by downregulating the hTERT mRNA and protein expression, and inhibition of telomerase with hTERT antisense can enhance TNF-α- induced apoptosis of PC3 cells.
