Effect of Antisense Oligodeoxynucleotide Targeting hTERT on Telome-rase Activity and Cell Apoptosis in K562 Cell Line

The study was purposed to investigate the inhibitory effect of antisense phosphorothioate oligodeoxy nucleotide(ASPSODN)targeting hTERT mRNA on gene of interest in K562 cells and influence of ASPSODN on telomerase activity and apoptosis of K562 cells.Human leukemia cell line K562 was transfected by liposome with ASPSODN and SPSODN(sense phsophorothioate oligodeoxynucleotide)at different concentrations(0.2,0.6,1.0 μmol/L).At the same time,blank control,liposome control and SPSODN groups were designed for comparison.The transfected cells were collected and detected at 24 and 48 hours;the expression of target gene hTERT mRNA and telomerase activity were detected by RT-PCR and TRAP-ELISA respectively,and cell apoptosis was assayed by flow cytometry.The results showed that after K562 cells were transfected for 24 hours,the expression of hTERT mRNA had no difference between liposome control(0.80±0.24),0.2 μmol/L ASPSODN(0.69±0.12),0.2 μmol/L SPSODN(0.72±0.25)and blank control(0.85±0.28),but the expression of hTERT mRNA in 0.6 μmol/L ASPSODN group(0.42±0.16)remarkably decreased as compared with liposome control group,0.6 μmol/L SPSODN(0.69±0.26)had no obvious effect on the expression of hTERT mRNA,the expression of hTERT mRNA in 1.0 μmol/L ASPSODN and SPSODN groups both decreased;mortality of K562 cells transfected by liposome with 1.0 μmol/L ASPSODN and SPSODN remarkably increased.After 24 hours,telomerase relative activity of K562 cells showed no significant difference between blank control(88.9%)and liposome control(77.7%).The telomerase relative activities of K562 cells treated with 0.2,0.6,1.0 μmol/L ASPSODN were 60.6%,52%,58.2% respectively.There was significant difference as compared with blank control;0.6 μmol/L ASPSODN showed significant difference(p=0.037),as compared with liposome control group.The telomerase relative activities in K562 cells treated with 0.2,0.6,1.0 μmol/L SPSODN were 76.1%,72.2%,65.7% respectively,but the telomerase relative activities of K562 cells in 0.2,0.6 μmol/L SPSODN groups was not inhibited obviously.When K562 cells were treated for 48 hours,telomerase relative activity of K562 cells in each ASPSODN groups restored.It showed that telomerase relative activities of K562 cells treated with 0.2,0.6,1.0 μmol/L ASPSODN were 84.1%,82.3%,79.6% respectively,while teromerase relative activities of K562 cells treated with 0.2,0.6,1.0 μmol/L SPSODN for 48 hours were 74.8%,74.5%,67.9% respectively.Telomerase activity of K562 cells could not be inhibited by 0.2 and 0.6 μmol/L SPSODN.After culturing for 48 hours,the cell apoptosis rates of K562 in 0.6 μmol/L ASPSODN,0.6 μmol/L SPSODN,liposome control and blank control groups were(4.82±0.39)%,(1.83±0.34)%,1.84±1.04)%,(1.07±0.74)% respectively.There was difference between ASPSODN and SPSODN gproups(p0.05),but the significant difference was found in ASPSODN group as compared with liposome control and blank control(p0.01).It is concluded that the ASPSODN targeting hTERT can specifically inhibit the expression of hTERT mRNA in K562 cells and significantly suppress the telomerase activity of K562 cells at 0.6 μmol/L,which inhibitory time is short.The ASPSODN at high concentration(1.0 μmol/L)shows definite cytotoxity.0.6 μmol/L of ASPSODN significantly induces cell apoptosis,while no such effect was seen in SPSODN group.