ABSTRACT
AIM: To explore the effect of hTERT antisense phosphorothioate oligodeoxynucleotide (ASODN) on apoptosis induced by chemotherapeutic drugs in Jurkat cell lines. METHODS: Cell viability was determined using the trypan blue dye exclusion assay. Apoptosis was detected by morphological observation, DNA gel electrophoresis and flow cytometry analysis. RESULTS: The survival rates of Jurkat cells cultured with daunorubicin, vincristin, and etoposide, respectively were similar with that cultured with those chemotherapic drugs plus hTERT ASODN. The survival rates of Jurkat cells cultured with cis-diamminedichicloroplatinum(DDP) added 24 hours later were higher than that cultured with hTERT ASODN and DDP added 24 hours later. The survival rates of Jurkat cells cultured with DDP were similar with that cultured with hTERT SODN and DDP. In morphological observation of apoptotic cells using Giemsa staining, cells displayed classic apoptotic changes treated with DDP or DDP combined with hTERT ASODN or SODN at 48 hours. Agarose gel electrophoresis of genomic DNA from Jurkat cells treated with ASODN and DDP combination for 48 hours showed typical DNA “ladder”. Neither the DNA from Jurkat cells treated with SODN plus DDP nor the DNA from the cells treated with DDP alone showed ladder pattern. Apoptosis rates of Jurkat cells treated with DDP for 48 hours after 24 hours of exposure to ASODN significantly increased. There was significant difference in the percentage of apoptotic Jurkat cells between hTERT ASODN plus DDP and SODN plus DDP or DDP alone, respectively. CONCLUSION: The hTERT ASODN complementary to the translation initiation region of hTERT mRNA enhanced DDP-induced apoptosis in Jurkat cells.
ABSTRACT
AIM: To explore the effect of human telomerase reverse transcriptase (hTERT) gene antisense phosphorothioate oligodeoxynucleotide (AS PS-ODN) on cisplatin-induced apoptosis in cultured primary acute lymphoblastic leukemia (ALL) cells. METHODS: The expression levels of hTERT protein were detected by immunofluorescence using fluoresce isothiocyanate (FITC) lable. Cell surviving fraction was determined using the trypan blue dye exclusion assay. Apoptosis was detected by morphological observation and flow cytomertric cell cycle analysis. RESULTS: The expression of hTERT protein was inhibited after treated by hTERT AS PS-ODN. Treatment with cisplatin after 24 h of exposure to AS PS-ODN had significantly reduced the number of viable ALL cells. However,there was no difference on ALL cells survival between sense oligodeoxynucleotide (S PS-ODN) /CDDP combination and CDDP-treated cells alone. In morphological observation of apoptotic cells using Hoechst 33258 and PI double staining techniques,cells displayed classic apoptotic changes treated with CDDP or CDDP combined with hTERT AS PS-ODN or S PS-ODN at 48 h. Apoptotic rates of cells added CDDP and AS PS-ODN were higher than that of cells added CDDP only ( P 0.05). CONCLUSION: hTERT AS PS-ODN inhibited the expression of hTERT protein and increased the CDDP-induced apoptosis in primer acute lymphoblastic leukemic cells.
ABSTRACT
AIM: To explore the effect of hTERT gene antisense phosphorothioate oligodeoxynucleotide (ASODN) on telomerase activity in Raji cells. METHODS: Polymerase chain reaction enzyme-linked immuonassay (PCR-ELISA) was used to determine telomerase activity. The expression levels of hTERT mRNA and protein were assayed by reverse transcriptase polymerase chain reaction (RT-PCR) and immunofluorescence using fluoresce isothiocyanate (FITC) label,respectively. RESULTS: RT-PCR and immunofluorescence assay showed that the expression levels of hTERT mRNA and protein from Raji cells decreased with time after hTERT ASODN treatment. There was no difference in hTERT mRNA and protein levels between control and SODN-treated cells. Telomerase activity decreased when Raji cells were treated with ASODN for 48 h. Telomerase activity of Raji cells was significantly inhibited when treated with ASODN for 72 h. There was no difference in telomerase activity levels between control and hTERT SODN-treated cells. CONCLUSION: hTERT ASODN could inhibit telomerase activity in Raji cells.