miR-15b-5p通过靶向LATS2基因调控前列腺癌细胞侵袭、迁移以及凋亡的分子机制
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摘要
目的探讨miR-15b-5p靶向大肿瘤抑制分子(LATS2)基因对前列腺癌PC-3细胞侵袭、迁移及凋亡的影响及机制。方法将anti-miR-15b-5p及anti-miR-15b-5p+si-LATS2(同时抑制miR-15b-5p和LATS2表达)转染前列腺癌PC-3细胞,48 h后,通过qRT-PCR检测miR-15b-5p mRNA表达,Transwell小室检测细胞侵袭和迁移能力;膜联蛋白V-异硫氰酸荧光素/碘化丙啶(Annexin V-FITC/PI)双染细胞凋亡试剂盒检测细胞凋亡率。双荧光素酶报告系统检测miR-15b-5p与LATS2的靶向关系。Western blotting检测LATS2、β-catenin、MMP-2和Bax蛋白表达。结果 PC-3细胞转染anti-miR-15b-5p后,miR-15b-5p mRNA表达明显降低,细胞侵袭和迁移能力明显降低,细胞凋亡率明显升高(P <0. 05)。miR-15b-5p与LATS2存在靶向关系,抑制LATS2表达后可明显减弱anti-miR-15b-5p对PC-3细胞侵袭、迁移、凋亡及β-catenin、MMP-2和Bax蛋白表达的影响(P <0. 05)。结论抑制miR-15b-5p可明显降低前列腺癌PC-3细胞的侵袭和迁移能力,并诱导细胞凋亡,其机制与靶向LATS2抑制Wnt/β-catenin信号通路有关。
Objective To investigate the effect of LATS2 gene targeting LATS2 gene on invasion,migration and apoptosis of prostate cancer PC-3 cells and its mechanism. Methods anti-miR-15b-5p and anti-miR-15b-5p + si-LATS2( simultaneously inhibiting the expression of miR-15b-5p and LATS2) were transfected into PC-3 cells. After 48 hours,the expression of miR-15b-5p was detected by qRT-PCR,the invasion and migration ability of cells were detected by Transwell chamber,and the apoptotic rate was detected by Annexin V-FITC/PI double-transfection kit. Dual luciferase reporter system was used to detect the target relationship between miR-15b-5p and LATS2.Western blotting was used to detect the expression of LATS2,β-catenin,MMP-2 and Bax protein. Results After anti-microRNA-15b-5p were transfected into PC-3 cells,the expression of miR-15b-5p decreased significantly,the invasion and migration ability of cells decreased significantly,and the apoptotic rate increased significantly( P < 0. 05). miR-15b-5p has a targeting relationship with LATS2. Inhibition of LATS2 expression could significantly attenuate the effects of anti-mir-15b-5p on invasion,migration,apoptosis and expression of β-catenin,MMP-2 and Bax in PC-3 cells( P < 0. 05). Conclusion Inhibiting miR-15b-5p can significantly reduce the invasion and migration ability of PC-3 cells and induce apoptosis. The mechanism is related to targeting LATS2 by inhibiting the Wnt/β-catenin signaling pathway.
Objective To investigate the effect of LATS2 gene targeting LATS2 gene on invasion,migration and apoptosis of prostate cancer PC-3 cells and its mechanism. Methods anti-miR-15b-5p and anti-miR-15b-5p + si-LATS2( simultaneously inhibiting the expression of miR-15b-5p and LATS2) were transfected into PC-3 cells. After 48 hours,the expression of miR-15b-5p was detected by qRT-PCR,the invasion and migration ability of cells were detected by Transwell chamber,and the apoptotic rate was detected by Annexin V-FITC/PI double-transfection kit. Dual luciferase reporter system was used to detect the target relationship between miR-15b-5p and LATS2.Western blotting was used to detect the expression of LATS2,β-catenin,MMP-2 and Bax protein. Results After anti-microRNA-15b-5p were transfected into PC-3 cells,the expression of miR-15b-5p decreased significantly,the invasion and migration ability of cells decreased significantly,and the apoptotic rate increased significantly( P < 0. 05). miR-15b-5p has a targeting relationship with LATS2. Inhibition of LATS2 expression could significantly attenuate the effects of anti-mir-15b-5p on invasion,migration,apoptosis and expression of β-catenin,MMP-2 and Bax in PC-3 cells( P < 0. 05). Conclusion Inhibiting miR-15b-5p can significantly reduce the invasion and migration ability of PC-3 cells and induce apoptosis. The mechanism is related to targeting LATS2 by inhibiting the Wnt/β-catenin signaling pathway.
引文
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[22]邱承俊,程帆. MMP-2基因对前列腺癌细胞恶性表型的影响及其机制研究[J].临床和实验医学杂志,2018,17(4):365-370.
[23]Mohammadian J,Sabzichi M,Molavi O,et al. Combined Treatment with Stattic and Docetaxel Alters the Bax/Bcl-2 Gene Expression Ratio in Human Prostate Cancer Cells[J]. Asian Pac J Cancer Prev,2016,17(11):5031-5035.
[2]Li Y,Chen Y,Li J,et al. Co-delivery of microRNA-21 antisense oligonucleotides and Gemcitabine using nanomedicine for pancreatic cancer therapy.[J]. Cancer Sci,2017,108(7):1493-1503.
[3]Ruan L,Chen J,Ruan L,et al. MicroRNA-186 suppresses lung cancer progression by targeting SIRT6[J]. Cancer Biomark,2017,21(2):415-423.
[4]Zhao C,Zhao Q,Zhang C,et al. miR-15b-5p resensitizes colon cancer cells to 5-fluorouracil by promoting apoptosis via the NF-κB/XIAP axis[J]. Sci Rep,2017,7(1):4194-4206.
[5]Chen R,Sheng L,Zhang HJ,et al. miR-15b-5p facilitates the tumorigenicity by targeting RECK and predicts tumour recurrence in prostate cancer[J]. J Cell Mol Med,2018,22(3):1855-1863.
[6]Ladiz MAR,Najafi M,Korditamandani DM. Contribution of LATS1 and LATS2 promoter methylation in OSCC development[J]. J Cell Commun Signal,2017,11(1):49-55.
[7]Zheng YB,Xiao K,Xiao GC,et al. MicroRNA-103 promotes tumor growth and metastasis in colorectal cancer by directly targeting LATS2[J]. Oncol Lett,2016,12(3):2194-2200.
[8]Guo Y,Cui J,Ji Z,et al. miR-302/367/LATS2/YAP pathway is essential for prostate tumor-propagating cells and promotes the development of castration resistance[J]. Oncogene,2017,36(45):6336-6347.
[9]Chua SK,Wang BW,Lien L M,et al. Mechanical Stretch Inhibits MicroRNA499 via p53 to Regulate Calcineurin-A Expression in Rat Cardiomyocytes[J]. Plos One,2016,11(2):e0148683.
[10]Inoguchi S,Seki N,Chiyomaru T,et al. Tumour-suppressive microRNA-24-1 inhibits cancer cell proliferation through targeting FOXM1in bladder cancer[J]. Febs Letters,2016,588(17):3170-3179.
[11]付晓亮,杨增悦,舒涛,等. miR-103通过靶向PDCD10逆转顺铂诱导的前列腺癌细胞耐药[J].现代肿瘤医学,2017,25(4):511-515.
[12]Gui B,Hsieh CL,Kantoff PW,et al. Androgen receptor-mediated downregulation of microRNA-221 and-222 in castration-resistant prostate cancer[J]. Plos One,2017,12(9):e0184166.
[13]Zhao C,Li Y,Chen G,et al. Overexpression of miR-15b-5p promotes gastric cancer metastasis by regulating PAQR3[J]. Oncol Rep,2017,38(1):352-358.
[14]Li H,Zhang J,Lee MJ,et al. OIP5,a target of miR-15b-5p,regulates hepatocellular carcinoma growth and metastasis through the AKT/m TORC1 andβ-catenin signaling pathways[J]. Oncotarget,2017,8(11):18129-18144.
[15]Wan L,Sun M,Liu G J,et al. Long non-coding RNA PVT1 promotes non-small cell lung cancer cell proliferation through epigenetically regulating LATS2 expression[J]. Mol Cancer Ther,2016,15(5):1082-1094.
[16]刘武军,李焕祥,马彦寿,等.微小RNA-373靶向调控LATS2的表达及其对肝癌细胞HepG2增殖与凋亡的影响[J].临床肿瘤学杂志,2016,21(12):1074-1078.
[17]Wu KF,Liang WC,Feng L,et al. H19 mediates methotrexate resistance in colorectal cancer through activating Wnt/β-catenin pathway[J]. Exper Cell Res,2017,350(2):312-317.
[18]Ohsugi T,Yamaguchi K,Zhu C,et al. Decreased expression of interferon-induced protein 2(IFIT2)by Wnt/β-catenin signaling confers anti-apoptotic properties to colorectal cancer cells[J]. Oncotarget,2017,8(59):100176-100186.
[19]Yu EJ,Hooker E,Johnson DT,et al. LZTS2 and PTEN collaboratively regulate-catenin in prostatic tumorigenesis[J]. Plos One,2017,12(3):e0174357.
[20]Chen Q,Zhao X,Zhang H,et al. MiR-130b suppresses prostate cancer metastasis through down-regulation of MMP2[J]. Mol Carcinog,2015,54(11):1292-1300.
[21]Emeline T,Fran?ois B,Jean-Yves P,et al. MMP2 and MMP9 serum levels are associated with favorable outcome in patients with inflammatory breast cancer treated with bevacizumab-based neoadjuvant chemotherapy in the BEVERLY-2 study[J]. Oncotarget,2016,7(14):18531-18540.
[22]邱承俊,程帆. MMP-2基因对前列腺癌细胞恶性表型的影响及其机制研究[J].临床和实验医学杂志,2018,17(4):365-370.
[23]Mohammadian J,Sabzichi M,Molavi O,et al. Combined Treatment with Stattic and Docetaxel Alters the Bax/Bcl-2 Gene Expression Ratio in Human Prostate Cancer Cells[J]. Asian Pac J Cancer Prev,2016,17(11):5031-5035.