Mir-340 Inhibits The Oncogenic Potential Of Prostate Cancer Cells By Targeting HMGN5

Prostate cancer is a common cancer in worldwide, which seriously threat the health of men. microRNA is abnormal expression in several tumor which participated in the tumorigenesis, in which the mi R-340 have been reported to inhibit tumor growth in different cancer types. However, it is still unknown whether miR-340 plays an important role in prostate cancer. Therefore, this study aims to explore the expression pattern of mi R-340 and its functions in prostate cancer. Then continue to validated whether HMGN5 is the target gene of miR-340 and regulating the expression through its promotion, then affect the proliferation and invasion of prostate cancer. Above in all, this study will maybe provide a new therapeutic target in the treatment of prostate cancer.This study totaly include the three parts: Part one: The expression pattern of miR-340 in prostate cancer cells and tissues; The second part: The biological behavior of miR-340 in prostate cancer; Part three: The preliminary mechannism study of miR-340 which how to regulate the HMDN5 expressionPart I The expression pattern of miR-340 in prostatecancer cells and tissues Methods:1) Clinical specimens collection, including 20 cases of prostate cancer tissues and 20 cases of paired peritumoral tissue specimens.2) The prostate cancer cell lines culture, including PC-3, 22RV-1, DU-145 and LNCaP prostate cancer cells with the RPMI 1640 medium. Normal human prostate epithelial cells rwpe1 was cultured in serum-free medium as the control.3) miR-340 mRNA expression levels examination using RT-PCR, including prostate cancer tissues, adjacent normal tissue, prostate cancer cell lines and normal human prostate epithelial cells. Results:1) Prostate cancer tissues with lower expression of miR-340 compared with adjacent tissues.2) Compared with normal prostate epithelial cells, prostate cancer cell lines with lower expression of miR-340 significantly. In conclusion:The expression of miR-340 was significantly down-regulated in prostate cancer cell lines and prostate cancer cell lines.Part II The biological behavior of miR-340 in prostatecancer Methods:1) Prostate cancer cells were transfected with miR-340 mimics and anti-mir-340, then to investigate the effect of mi R-340 on proliferation and migration activity in prostate cancer cells.2) Prostate cancer cells were transfected with miR-340 mimics and anti-mir-340, then to investigate the effect on proliferation by MTT assay and colony formation assay in prostate cancer cells.3) Prostate cancer cells were transfected with miR-340 mimics and anti-mir-340, then to investigate the effect on apoptosis of prostate cancer cells using the TUNEL method.4) Prostate cancer cells were transfected with miR-340 mimics and anti-mir-340, then to investigate the expression level of apoptosis related protein caspase-3.5) Prostate cancer cells were transfected with miR-340 mimics and anti-mir-340, then to investigate the invasion activity of prostate cancer cell. Results:1) Overexpression of miR-340 can inhibit the proliferation of prostate cancer cell, down regulation miR-340 may promote the proliferation.2) Overexpression of mi R-340 can inhibit the colony formation of prostate cancer cell, downregulation miR-340 may increase the cells colony formation.3) Overexpression of miR-340 can promote prostate cancer cell apoptosis, repression miR-340 can inhibit the apoptosis of prostate cancer cells.4) Overexpression of mi R-340 can inhibit the invasion activity of prostate cancer cell, repression miR-340 can promote the invasion of prostate cancer cells.In conclusion:Overexpressing mi R-340 can significantly inhibit the proliferation of prostatecancer cells and induce apoptosis. In the while, repression mi R-340 can inhibit theinvasion ability.Part III The mechanism study of miR-340 on target gene HGMN5 and the tumor suppressor role in prostate cancer cellsMethods:1) Construction the vector for luciferase assay, which used to investigate thebinding activity of miR-340 to HGMN5 3'UTR. miR-340 mimics or anti-miR-340were co-transfected into the DU-145 cells to examinate whether HMGN5 is the directtarget gene mi R-340.2) Detecting the expression of HMGN5 by real-time PCR in prostate cancercells, which were treated with miR-340 mimics or anti-miR-340.3) Detecting the level of HMGN5, Bcl-2, cyclin B1 and MMP9 proteinexpression, by Western blot analysis in prostate cancer cells, which were treated withmiR-340 mimics or anti-mi R-340.4) Examination of the biological behavior of prostate cancer cells which treatedwith co-transfection HMGN5 and miR-340 mimics. So that to investigate theoverexpression of HMGN5 affect on miR-340.5) The correlation analysis of the HMGN5 mRNA and miR-340 with Spearmanmethod in prostate cancer tissues.Results:1) HMGN5 maybe the direct target gene of miR-340.2) Overexpression mi R-340 can significantly reduce the expression of HGMN5in prostate cancer cells. In contrast, inhibition of miR-340 can significantly increasedthe expression of HGMN5.3) Overexpression miR-340 can significantly reduce the expression of Bcl-2, cyclin B1 and MMP9. Instead, inhibition of miR-340 can significantly increased the expression of these proteins.4) Overexpression miR-340 can suppressed the proliferation and invasion activity of prostate cancer cells, and these effect can be reversed by overexpression of HMGN5.5) There was be a significant negative correlation between miR-340 and HMGN5 mRNA level in prostate cancer tissues. In conclusion:We make sure that the HMGN5 gene was the direct target gene of mi R-340 in prostate cancer cells in this study, and overexpression miR-340 could repress the proliferation of prostate cancer cells by negatively regulates HMGN5. In this study, we firstly found that the key relation of mi R-340 and HMGN5 in prostate cancer. Therefore, this study not only provides the molecular mechanism of prostate cancer tumorigenesis, but also suggests a potential therapeutic target for clinical prostate cancer treatment.