MicroRNA-140-5p通过蛋白激酶C亚型ε促进肾癌细胞增殖（英文）
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摘要
目的探讨microRNA-140-5p(miR-140-5p)在肾癌发生中的作用。方法 Real-time PCR方法检测肾癌细胞(Caki-1、ACHN、OS-RC-2和786-O)中miR-140-5p的表达。MTS,流式细胞术和Western blotting检测细胞增殖,细胞周期和相关蛋白的改变。结果降低Caki-1细胞中miR-140-5p的水平后,细胞增殖被抑制,G0/G1期细胞数量显着增加,cyclin D、cyclin E和CDK2、CDK4也随之减少;此外,增加786-O细胞中miR-140-5p的水平后,促进体外细胞增殖和体内肿瘤发生。基因芯片结果发现,蛋白激酶C亚型ε(PKCε)基因在改变Caki-1和786-O的细胞中显著性升高。si PKCε在降低肾癌细胞(RCC)中PKCε蛋白表达的同时,还能显著性反转抑制物/模拟物诱导的细胞增殖和细胞周期的改变。结论 miR-140-5p通过PKCε促进肾癌细胞增殖。
Objective To explore the role of micro RNA-140-5 p( miR-140-5 p) in renal cell carcinoma( RCC)tumorigenesis. Methods The expression of miR-140-5 p was determined by Real-time PCR in RCC( Caki-1,ACHN,OSRC-2 and 786-O),and compared with kidney cortex proximal tubule cell( HK-2). After Caki-1 and 786-O cells were transfected with miR-140-5 p inhibitor or mimic,the cell proliferation,cell cycle and related proteins were evaluated by MTS assay,flow cytometry and Western blotting. Results Caki-1 and 786-O showed the highest and lowest expression of miR-140-5 p respectively. After the level of miR-140-5 p in Caki-1 cells was decreased,cell proliferation was inhibited and cell number in G0/G1 phase was increased significantly,along with the reduction of cyclin D, E and CDK2,4.Furthermore,the increasing level of miR-140-5 p in 786-O cells promoted cell proliferation in vitro and tumorigenesis in vivo. Moreover,microarray demonstrated that protein kinase C isoform ε( PKCε),a markedly enhanced gene in RCC cells with the changed level of miR-140-5 p. The combination treatment with down-regulation of PKCε,inhibitor/mimic induced changes of cell proliferation and cell cycle arrests were marked reversed in Caki-1 and 786-O cells. Conclusion These result indicate that miR-140-5 p may promote cell proliferation via PKCε in RCC.
Objective To explore the role of micro RNA-140-5 p( miR-140-5 p) in renal cell carcinoma( RCC)tumorigenesis. Methods The expression of miR-140-5 p was determined by Real-time PCR in RCC( Caki-1,ACHN,OSRC-2 and 786-O),and compared with kidney cortex proximal tubule cell( HK-2). After Caki-1 and 786-O cells were transfected with miR-140-5 p inhibitor or mimic,the cell proliferation,cell cycle and related proteins were evaluated by MTS assay,flow cytometry and Western blotting. Results Caki-1 and 786-O showed the highest and lowest expression of miR-140-5 p respectively. After the level of miR-140-5 p in Caki-1 cells was decreased,cell proliferation was inhibited and cell number in G0/G1 phase was increased significantly,along with the reduction of cyclin D, E and CDK2,4.Furthermore,the increasing level of miR-140-5 p in 786-O cells promoted cell proliferation in vitro and tumorigenesis in vivo. Moreover,microarray demonstrated that protein kinase C isoform ε( PKCε),a markedly enhanced gene in RCC cells with the changed level of miR-140-5 p. The combination treatment with down-regulation of PKCε,inhibitor/mimic induced changes of cell proliferation and cell cycle arrests were marked reversed in Caki-1 and 786-O cells. Conclusion These result indicate that miR-140-5 p may promote cell proliferation via PKCε in RCC.
引文
[1]Siegel RL,Miller KD,Jemal A.Cancer statistics,2016[J].Ca ACancer J Clini,2016,66(1):7-30.
[2]Gong BS,Feng Q.Netrin-1:the new tumor markers in renal clear cell carcinoma[J].Asian Pac J Trop Med,2015,8(6):488-492.
[3]Bartel DP.MicroRNAs:genomics,biogenesis,mechanism,and function[J].Cell,2004,116(2):281-297.
[4]Cekaite L,Eide P W,Lind GE,et al.MicroRNAs as growth regulators,their function and biomarker status in colorectal cancer[J].Oncotarget,2016,7(6):6476-6505.
[5]Ryou-UT,Hiroaki M,Takahiro O.The role of microRNAs in the regulation of cancer stem cells[J].Front Genet,2013,4(4):295.
[6]Linehan WM.The genetic basis of kidney cancer:implications for management and use of targeted therapeutic approaches[J].Eur Urol,2012,61(5):896-898.
[7]Petillo D,Kort EJ,Anema J,et al.MicroRNA profiling of human kidney cancer subtypes[J].Int J Oncol,2009,35(1):109-114.
[8]Heinzelmann J,Henning B,Sanjmyatav J,et al.Specific miRNAsignatures are associated with metastasis and poor prognosis in clear cell renal cell carcinoma[J].World J Urol,2011,29(3):367-373.
[9]Youssef YM,White NM,Grigull J,et al.Accurate molecular classification of kidney cancer subtypes using microRNA signature[J].European Urology,2011,59(5):721-730.
[10]Li H,Guan SB,Lu Y,et al.MiR-140-5p inhibits synovial fibroblasts proliferation and inflammatory cytokines secretion through targeting TLR4[J].Biomed Pharmacother,2017,96:208-214.
[11]Guo H,Yang S,Li S,et al.LncRNA SNHG20 promotes cell proliferation and invasion via miR-140-5p-ADAM10 axis in cervical cancer[J].Biomed Pharmacother,2018,102:749-757.
[12]Miao X,Wang Z,Chen B,et al.miR-140-5p suppresses retinoblastoma cell proliferation,migration,and invasion by targeting CEMIP and CADM3[J].Cell Mol Biol,2018,64(6):42-47.
[13]Hu Y,Li Y,Wu C,et al.MicroRNA-140-5p inhibits cell proliferation and invasion by regulating VEGFA/MMP2 signaling in glioma[J].Tumour Biol,2017,39(4):1-12.
[14]Zhuang H,Meng X,Li Y,et al.Cyclic AMP responsive elementbinding protein promotes renal cell carcinoma proliferation probablyviathe expression of spindle and kinetochore-associated protein 2[J].Oncotarget,2016,7(13):16325-16337.
[15]Wang X,Ren Y,Zhuang H,et al.Decrease of phosphorylated proto-oncogene CREB at Ser 133 site inhibits growth and metastatic activity of renal cell cancer[J].Expert Opinion on Therapeutic Targets,2015,19(7):985-995.
[16]Chen X,Wang X,Ruan A,et al.miR-141 is a key regulator of renal cell carcinoma proliferation and metastasis by controlling EphA2 expression[J].Clini Cancer Res,2014,20(10):2617-2630.
[17]Croce CM,Calin GA.miRNAs,cancer,and stem cell division[J].Cell,2005,122(1):6-7.
[18]Hanahan D,Weinberg RA.Hallmarks of cancer:the next generation[J].Cell,2011,144(5):646-674.
[19]Kahl CR,Means AR.Regulation of cell cycle progression by calcium/calmodulin-dependent pathways[J].Endocr Revi,2003,24(6):719-736.
[20]Wang P,Huang S,Wang F,et al.Cyclic AMP-response element regulated cell cycle arrests in cancer cells[J].PLoS One,2013,8(6):e65661.
[21]Desdouets C,Matesic G,Molina CA,et al.Cell cycle regulation of cyclin A gene expression by the cyclic AMP-responsive transcription factors CREB and CREM[J].Mol Cell Biol,1995,71(2):233-234.
[22]Jaken S.Protein kinase C isozymes and substrates[J].Curr Opini Cell Biol,1996,8(2):168-173.
[23]Gorin MA,Pan Q.Protein kinase C epsilon:an oncogene and emerging tumor biomarker[J].Mol Cancer,2009,8:9.
[24]Jiang Z,Kong C,Zhang Z,et al.Reduction of protein kinase Cα(PKC-α)promote apoptosis via down-regulation of Dicer in bladder cancer[J].J Cell Mol Medi,2015,19(5):1085-1093.
[25]Wu D,Foreman TL,Gregory CW,et al.Protein kinase cepsilon has the potential to advance the recurrence of human prostate cance[J]r.Cancer Res,2002,62(8):2423-2429.
[26]Xu XW,Yang YL,Sun ZhL,et al.Raptor promoting invasion and metastasis of breast cancer through the signaling pathway Wnt3a/β-catenin[J].Acta Anatomica Sinica,2017,48(6):682-687.
[27]Pan Q,Li WB,Teknos TN,et al.Targeted disruption of protein kinase Cεreduces Cell invasion and motility through inactivation of RhoA and RhoC GTPases in head and neck squamous cell carcinoma[J].Cancer Res,2006,66(19):9379-9384.
[28]Huang B,Cao K,Li X,et al.The expression and role of protein kinase C(PKC)epsilon in clear cell renal cell carcinoma[J].JExp Clini Cancer Res,2011,30:88.
[2]Gong BS,Feng Q.Netrin-1:the new tumor markers in renal clear cell carcinoma[J].Asian Pac J Trop Med,2015,8(6):488-492.
[3]Bartel DP.MicroRNAs:genomics,biogenesis,mechanism,and function[J].Cell,2004,116(2):281-297.
[4]Cekaite L,Eide P W,Lind GE,et al.MicroRNAs as growth regulators,their function and biomarker status in colorectal cancer[J].Oncotarget,2016,7(6):6476-6505.
[5]Ryou-UT,Hiroaki M,Takahiro O.The role of microRNAs in the regulation of cancer stem cells[J].Front Genet,2013,4(4):295.
[6]Linehan WM.The genetic basis of kidney cancer:implications for management and use of targeted therapeutic approaches[J].Eur Urol,2012,61(5):896-898.
[7]Petillo D,Kort EJ,Anema J,et al.MicroRNA profiling of human kidney cancer subtypes[J].Int J Oncol,2009,35(1):109-114.
[8]Heinzelmann J,Henning B,Sanjmyatav J,et al.Specific miRNAsignatures are associated with metastasis and poor prognosis in clear cell renal cell carcinoma[J].World J Urol,2011,29(3):367-373.
[9]Youssef YM,White NM,Grigull J,et al.Accurate molecular classification of kidney cancer subtypes using microRNA signature[J].European Urology,2011,59(5):721-730.
[10]Li H,Guan SB,Lu Y,et al.MiR-140-5p inhibits synovial fibroblasts proliferation and inflammatory cytokines secretion through targeting TLR4[J].Biomed Pharmacother,2017,96:208-214.
[11]Guo H,Yang S,Li S,et al.LncRNA SNHG20 promotes cell proliferation and invasion via miR-140-5p-ADAM10 axis in cervical cancer[J].Biomed Pharmacother,2018,102:749-757.
[12]Miao X,Wang Z,Chen B,et al.miR-140-5p suppresses retinoblastoma cell proliferation,migration,and invasion by targeting CEMIP and CADM3[J].Cell Mol Biol,2018,64(6):42-47.
[13]Hu Y,Li Y,Wu C,et al.MicroRNA-140-5p inhibits cell proliferation and invasion by regulating VEGFA/MMP2 signaling in glioma[J].Tumour Biol,2017,39(4):1-12.
[14]Zhuang H,Meng X,Li Y,et al.Cyclic AMP responsive elementbinding protein promotes renal cell carcinoma proliferation probablyviathe expression of spindle and kinetochore-associated protein 2[J].Oncotarget,2016,7(13):16325-16337.
[15]Wang X,Ren Y,Zhuang H,et al.Decrease of phosphorylated proto-oncogene CREB at Ser 133 site inhibits growth and metastatic activity of renal cell cancer[J].Expert Opinion on Therapeutic Targets,2015,19(7):985-995.
[16]Chen X,Wang X,Ruan A,et al.miR-141 is a key regulator of renal cell carcinoma proliferation and metastasis by controlling EphA2 expression[J].Clini Cancer Res,2014,20(10):2617-2630.
[17]Croce CM,Calin GA.miRNAs,cancer,and stem cell division[J].Cell,2005,122(1):6-7.
[18]Hanahan D,Weinberg RA.Hallmarks of cancer:the next generation[J].Cell,2011,144(5):646-674.
[19]Kahl CR,Means AR.Regulation of cell cycle progression by calcium/calmodulin-dependent pathways[J].Endocr Revi,2003,24(6):719-736.
[20]Wang P,Huang S,Wang F,et al.Cyclic AMP-response element regulated cell cycle arrests in cancer cells[J].PLoS One,2013,8(6):e65661.
[21]Desdouets C,Matesic G,Molina CA,et al.Cell cycle regulation of cyclin A gene expression by the cyclic AMP-responsive transcription factors CREB and CREM[J].Mol Cell Biol,1995,71(2):233-234.
[22]Jaken S.Protein kinase C isozymes and substrates[J].Curr Opini Cell Biol,1996,8(2):168-173.
[23]Gorin MA,Pan Q.Protein kinase C epsilon:an oncogene and emerging tumor biomarker[J].Mol Cancer,2009,8:9.
[24]Jiang Z,Kong C,Zhang Z,et al.Reduction of protein kinase Cα(PKC-α)promote apoptosis via down-regulation of Dicer in bladder cancer[J].J Cell Mol Medi,2015,19(5):1085-1093.
[25]Wu D,Foreman TL,Gregory CW,et al.Protein kinase cepsilon has the potential to advance the recurrence of human prostate cance[J]r.Cancer Res,2002,62(8):2423-2429.
[26]Xu XW,Yang YL,Sun ZhL,et al.Raptor promoting invasion and metastasis of breast cancer through the signaling pathway Wnt3a/β-catenin[J].Acta Anatomica Sinica,2017,48(6):682-687.
[27]Pan Q,Li WB,Teknos TN,et al.Targeted disruption of protein kinase Cεreduces Cell invasion and motility through inactivation of RhoA and RhoC GTPases in head and neck squamous cell carcinoma[J].Cancer Res,2006,66(19):9379-9384.
[28]Huang B,Cao K,Li X,et al.The expression and role of protein kinase C(PKC)epsilon in clear cell renal cell carcinoma[J].JExp Clini Cancer Res,2011,30:88.