Studies On The Expressions Of MicroRNA-129 In Prostate Cancer And Its Functional Mechanisms

Prostate cancer has become one of urinary tract malignances with the fastest rising incidence rates. Currently there are lack of ideal indicators for early diagnosis and condition monitoring of prostate cancer. The current regimens are also not helpful for improving the curative effects further. miRNAs are a class of non-coding RNA molecules, about 21-25 nucleotides in length, regulate the expression of target genes after transcription, involve in the formation and development of tumor. The oncology researchs based on miRNAs provides a new target for the diagnosis and treatment of prostate cancer and also a train of thought. The study showed the expression of miR-129 associated with the malignant degree of tumor indicated that miR-129 could be used in the diagnosis and monitoring of tumor. In order to know the relationship between miR-129 and prostate cancer and its function mechanisms, we tested the miR-129 expression in prostate cancer organizations, analyzed the relationships between miR-129 expression and clinical pathological characteristics of prostate cancer, as well as the impacts of miR-129 overexpression on the PC3 cell’s proliferation, apoptosis and invasion. Finally we deteced the expression of miR-129’s target gene and its downstream gene, and discussed the action mechanisms of miR-129. The experiment was divided into three parts:Objective To detect the miR-129 expression in prostate cancer organizations, investigate its relationship with clinical pathological characteristics of prostate cancer, and explore its clinical significance.Methods The expression of miR-129 was detected using real-time fluorescent quantitative PCR in 37 cases of prostate cancer and adjacent tissues, then to investigate expression differences among different clinical and pathologic group, and analyse its diagnostic value with ROC curve.Results The expression of endogenous miR-129 in prostate cancer tissues was lower than that in control groups (P<0.05). In the different clinical and pathologic group, all the expression differences of miR-129 had no statistical significance (P>0.05). The AUC (area under the ROC curve) of miR-129 was less than that of PSA.Conclusion 1. The expressions of miR-129 was low in prostate cancer, may be one of the reasons for prostate cancer occurrence.2. The expressions of miR-129 were not associated with the clinical features of prostate cancer, miR-129 was not a good clinical monitoring indicators.3. The diagnostic-efficacy of miR-129 was poor, not a good diagnosis index of prostate cancer.Objective To detect the biology behavior changes of PC-3 cells after overexpressing miR-129, observe the biological effects of miR-129 overexpression in PC3 cells.Methods Has-miR-129 mimic was transfeced into PC-3 cells, and then transfection efficiency was detected by real-time PCR. The cell proliferation activities were detected by CCK8 methods. The changes of cell cycle were detected by flow cytometry. The cell apoptosis was detected by Annexin V/7-AAD double staining methods. The abilities of cell migration and invasion were detected by transwell experiments.Results After transfection, the relative expression of miR-129 in PC-3 cells was 435 times of which in control group (P<0.01). Within 24h the cell proliferation abilities of transfection group were no differences with control group (P>0.05). After 48h, which was obviously lower than that of control group(P<0.05). The G1 phase cell rates in transfection group increased significantly compared with the control group (P<0.05), the S phase cell ratio decreased significantly compared with controls (P<0.05). The cell apoptosis rates in transfection group increased significantly compared with control group (P＜0.05). Migration experiments showed that the average transmembrane cells in transfection group reduced significantly compared with controls (P<0.01). Invasion experiments showed the invasive cells in transfection group were significantly less than the control group (P<0.01).Conclusion 1. Transfection with mimic could effectively form the state of miR-129 high-expression.2. The overexpression of miR-129 could negatively regulate PC-3 cell proliferation, cycle, apoptosis, invasion and other biological behavior.3. miR-129 might play the role of tumor suppressor genes in prostate cancer, could be used as a target for the prostate cancer treatment.Objective To detect the target gene of miR-129 in PC-3 cells and to explore its possible molecular mechanisms.Methods The target gene of miR-129 was predicted by using software (Targetscan, MIRanda and PicTar) and was verified through the dual-luciferase reporter assay system. After miR-129 over-expression, the mRNA or protein expressions of target gene were detected by real-time PCR and western blot methods respectively. The protein expressions of downstream genes were detected with western blot methods.Results The softwares predicted that MAPK1 gene might be a potential target gene of miR-129. Dual-luciferase reporter assay system showed that overexpression of miR-129 could obviously inhibit the luciferase activities of wild-type pMIR_MAPK1_WT (vs control, P<0.01), but basically no effections on the luciferase activities of mutation plasmids (vs controls, P>0.05). After miR-129 over-expression, MAPK1 mRNA and protein expressions were both less than control groups (P＜0.01). The variations of the total STAT3 protein expression were not obvious (P>0.05), but the phosphorylated STAT3 protein (p-STAT3) expressions decreased significantly compared with the control groups (P<0.01). The protein expressions of the downstream BCL-XL gene were significantly lower than control groups (P＜0.01).Conclusion 1. MAPK1 was a target gene of miR-129.2. MiR-129 might reduce the expressions of MAPK1 through mRNA degradation or inhibition of translation.3. miR-129 might affect the STAT3 transcription activity via MAPK1, there might exist MAPK1-STAT3-BCL-XL signal transduction pathway in prostate cancer.