The Study Of The Inhibitory Action Of TRAIL Combined With Paclitaxe On Prostate Cancer Cell

First part The inhibitory action of TRAIL combined with paclitaxe on prostate cancer cell linesObjective: Tumor necrosis factor-related apoptosis inducing ligand (TRAIL) induces apoptosis of multiple tumor cells including prostate cancer cells, but many human cancer cell are refractory to TRAIL-induced cell death. We intended to investigate the inhibitory action of TRAIL combined with paclitaxe on prostate cancer cell lines LNCAP, DU145 and PC3.Method: The survival rate of LNCAP, DU145 and PC3 cells treated respectively with TRAIL, paclitaxe and TRAIL combined with paclitaxe was tested by using MTT assay. Meanwhile, the apoptosis rate was evaluated by flow cytometric analysis.Result:(1) The inhibition effect of 2ng/ml TRAIL on LNCAP cell was obvious and in a time-dependent manner. The survival rate was (75.15±3.73)%, (66.93±2.90)% and (26.51±2.00)% when the cell was treated with 2ng/ml TRAIL for 24h, 48h and 72h respectively. To PC3 cells, the inhibition effect was lighter, the survival rate was (87.92±3.65)%,(70.54±3.43)%,(84.51±3.28)%, when PC3 cell treated with 2ng/ml TRAIL for 24h,48h,72h respectively To Du145 cell, the inhibition effect was very little, the survival rate was (92.69±3.82)%,(99.75±1.86)% by treated with 2ng/ml TRAIL for 48h,72h respectively. The inhibition effect was marked when LNCAP, DU145 and PC3 cells were treated with 2ng/ml TRAIL combined with 25nmol/ml paclitaxe, and it was strongest at 72h. The survival rate was (19.56±1.72)%, (41.24±1.84)%, (29.06±2.84)% respectively. The difference was significant as compared with cells treated with TRAIL or paclitaxe alone (P<0.01).(2) TRAIL could induce apoptosis of LNCAP, DU145 and PC3 cells. The apoptosis rate was (52.81±3.70)%, (21.68±1.48)%, (35.21±2.00)% when they were treated with 5ng/ml TRAIL for 24h. The apoptosis rate was (22.34±1.98)%, (14.34±2.23)%, (30.08±3.36)% when they were treated with 2ng/ml TRAIL for 24h. By comparison, the difference of the tow groups was significant (P<0.01), and the effect of inducing apoptosis showed a certain dose-effect relationship.(3) The effect of inducing apoptosis was enhanced obviously when LNCAP, DU145 and PC3 cells were treated with 2ng/ml TRAIL combined with 25nmol/ml paclitaxe. At 24h, 48h and 72h, the apoptosis rate were all higher obviously than that when treated with TRAIL or paclitaxe alone, the difference was significant (P<0.01).Conclusion: In comparison with TRAIL or paclitaxe, TRAIL combined with paclitaxe enhanced the inhibitory effect of cell growth and the effect of inducing apoptosis on LNCAP, DU145 and PC3 cells in a time- and dose-dependent manner.Second part The inhibitory action of TRAIL combined with paclitaxe on human primary prostate cancer cellsObjective: To investigate the inhibitory action of TRAIL combined with paclitaxe on human primary prostate cancer cells.Method: The human primary prostate cancer cells were separated from fresh tissue of prostate cancer 21 patients clinically. The survival rate of human primary prostate cancer cells treated respectively with TRAIL, paclitaxe and TRAIL combined with paclitaxe for 6h, 12h and 24h was measured by MTT method.Result:(1) The survival rate was respectively reduced to (90.30±6.04)%, (82.41±6.88)% and (73.31±7.50)%, when the human primary prostate cancer cells were treated with 5ng/ml TRAIL for 6h, 12h and 24h. Time-dependent relationship was obvious (P<0.01).(2) The survival rate was respectively reduced to (85.97±6.26)%, (75.28±7.16)% and (57.38±6.91)%, when the human primary prostate cancer cells were treated with 25nmol/ml paclitaxe for 6h, 12h and 24h. Time-dependence existed as well (P<0.01).(3) The survival rate was respectively reduced to (79.87±6.22)%, (68.81±7.13)% and (49.39±6.49)%, when the human primary prostate cancer cells were treated with 5ng/ml TRAIL combined with 25nmol/ml paclitaxe for 6h, 12h and 24h. Compare with the two groups above, the difference was significant (P<0.01). Moreover, the combined effect was also time-dependent.Conclusion: The inhibition effect on the human primary prostate cancer cells was markedly enhanced by the combination treatment of TRAIL and paclitaxe.Third Part The study of the mechanism of Paclitaxe-TRAIL combination treatments to prostate cancer cellObjective: To study the improvement of the inhibition of prostate cancer cell with the Paclitaxe-TRAIL combination treatments and its mechanism. Methods: JURKART, LNCAP, DU145, PC3 cancer cells were treated respectively by 2ng/ml TRAIL, 25nmol/ml and Paclitaxe- TRAIL combination. Their survival rates had been measured after 48 hours by MTT. The TRAIL-R2 mRNA and Caspase-3 mRNA expression in these four cancer cells had been detected by RT-PCR. The TRAIL-R2, Bcl-2 and Caspase-3 protein expression in the Du 145 cells with the 25nmol/ml Paclitaxe treatment had been detected by Western blot in 2hrs, 4hrs, 6hrs, 8hrs, 12hrs and 24hrs.Results:1. There had an inhibition effect in the JURKART, LNCAP, DU 145, PC3 cancer cells in 48hrs with the 2ng/ml TRAIL and 25nmol/ml Paclitaxe treatments, and there had a significant inhibition effect found in the Paclitaxe-TRAIL combination treatments (P<0.01). No significant inhibition effect was observed in DU145 cell with 2ng/ml TRAIL treatment alone, the survival rate was (94.86±4.18%). However, when Du145 cells was treated firstly by 25nmol/ml Paclitaxe for 24hrs, then by 2ng/ml TRAIL for 24hrs, there had a significant inhibition effect, its survival rate decreased to (41.63±3.39%)2. there had a high level expression of TRAIL-R2 mRNA in the JURKART, LNCAP, DU 145, PC3 cancer cells, and Paclitaxe had no influence on it. The Caspase-3 mRNA was expressed obviously in the JURKART, LNCAP and PC3 cancer cells, however, Paclitaxe had no effect on it. The Caspase-3 mRNA was hardly expressed in Du145 cells, but it increased sharply with the Paclitaxe treatments for 24hrs.3. 25nmol/ml Paclitaxe treated DU 145 cells in 2hrs, 4hrs, 6hrs, 8hrs, 12hrs and 24hrs respectively, there had no significant varieties found in TRAIL-R2 protein level, bcl-2 protein decreased gradually, and Caspase-3 protein increased. There had obvious differences among the times of the Paclitaxe treatments.(P<0.01)Conclusion: With the treatments of Paclitaxe in Du 145 cells for 24hrs, the inhibition effect of 2ng/ml TRAIL was improved. Z could depress the anti-apoptosis bcl-2 gene expression, increase Caspase-3 level, consequently improve the prostate cancer cell's sensitivity to TRAIL. Paclitaxe -TRAIL combination treatments could increase their abilities of inhibition effects against prostate cancer cells.