The Expression And Significance Of PARP-1in Prostate Cancer And Effect Of PARP-1on The Proliferation In Androgen Independent Prostate Cancer PC3Cell Line

Objective To investigate the expression and significance of Poly(ADP-ribose)polymerase-1(PARP-1) and its active polymer poly(ADP-ribose)(PAR) in ProstateCancer (PCa) and Benign prostatic hyperplasia (BPH) tissues from Chinese patients.And to observe the effects and underlying mechanism of siRNA targeting PARP-l onthe proliferation of androgen independent prostate cancer PC3cell line for providing anew way of the treatment of prostate cancer.Methods The expression of PARP-1and PAR assessed by immunohistochemistry SPmethod were compared between78PCa patients and49BPH patients in thehospital-based study. And to analyze the correlation of PARP-1and PAR with initialserum TPSA (TPSA＞20and TPSA≤20) in Pca, as well as in well-differentiated PCa(Gleason score≤6) and poorly differentiated PCa (Gleason score≥8) tissues. Threespecific siRNA sequences targeting PARP-l were designed and synthesized．And twosequences which had better interfering effect on the expression of PARP-l wereevaluated and selected through lipofectamine transfection， RT-PCR and WesternBlot．The effeet of PARP-l silencing on the proliferation of PC3cells was observed withMTS assay and the levels of the phosphorylation of Akt and GSK3β were detected byWestern Blot.Results1. Immunohistochemical analysis revealed that both PARP-1and PAR proteinswere mainly expressed in the nucleus of PCa or BPH tissue cells. The over-expressionof PARP-1and PAR was significantly higher in PCa than in BPH (P<0.05).2. Although spearman correlations analysis showed the over-expression of PARP-1and PAR in prostate cancer tissue was not correlated with age, initial serum TPSA andGleason scores (P>0.05), over-expression of PARP-1and PAR had an upward trendwith the initial TPSA levels (TPSA＞20vs TPSA≤20) and Gleason Score grade (GS≥8vs GS≤6).3. Compared to the blank control group，the transfected group with the negativecontrol sequence had no significant impact on the expression of PARP-l，however the transfected group with siRNA-1706，-2003or-2907could significantly suppress themRNA and protein expression of PARP-l．And the transfected group with siRNA-1706had the strongest interferential effect, while the transfected group with siRNA-2907hadthe weakest interferential effect. The differences were statistically significant (P<0.05).4. The siRNA-1706and-2003of PARP-1could significantly inhibit the proliferationof PC3cells. The differences were statistically significant (P<0.01).5. PARP-1targeted siRNA-1706and siRNA-2003could significantly down-regulatethe intracelln1ar levels of phosphorylated Akt (Ser473) and GSK3β(Ser9) in PC3cells.But the expression of total Akt1and GSK3β were not significantly affected.Conclusions1. The expression of PARP-1and its active product PAR is markedlyelevated in PCa than that in BPH tissues, which may implicate that PARP-1and PARare involved in the occurrence of PCa.2. There was no significant correlation between PARP-1or PAR and serum TPSA orGleason score in patients with prostate cancer, but with an increasing trend, indicatingthat it may serve as a marker for prostate cancer progression, but still need to beimproved in future research.3. PARP-l targeted siRNA was significantly suppress the expression of endogenousPARP-l and inhibit the proliferation of PC3cells, which was related to the inhibition ofAkt activity and the activation of GSK3β．It could contribute to the study of molecularmechanisms and treatment exploration of prostate cancer. But PARP-l targeted siRNAon the ranscriptional regulation mechanism of cell proliferation-related genes remainsto be further studied.4. Inhibition of PARP-1function is expected to become a new target for the treatment ofprostate cancer.