OBJECTIVES: To investigate the role of apelin in regulating proliferation, migration and angiogenesis of bladder cancer cells and the possible regulatory mechanism. METHODS: GEO database was used to screen the differentially expressed genes in bladder cancer tissues and cells. Bladder cancer and paired adjacent tissues were collected from 60 patients for analysis of apelin expressions in relation to clinicopathological parameters. In cultured bladder cancer J82 cells and human umbilical vein endothelial cells (HUVECs), the effects of transfection with an apelin-overexpressing plasmid or specific siRNAs targeting apelin, fibroblast growth factor 2 (FGF2) and fibroblast growth factor receptor 1 (FGFR1) on proliferation and migration of J82 cells and tube formation in HUVECs were examined using plate cloning assay, Transwell assay, and angiogenesis assay; the changes in FGF2 expression and FGFR1 phosphorylation were detected using Western blotting. RESULTS: The expression level of apelin was significantly higher in bladder cancer tissues than adjacent tissues, and bladder cancer cell lines (T24 and J82) also expressed higher mRNA and protein levels of apelin than SV-HUC-1 cells. Apelin expression level in bladder cancer tissues was correlated with tumor invasion, distant metastasis and advanced TNM stages. Apelin knockdown significantly suppressed proliferation and migration of J82 cells and decreased the total angiogenic length of HUVECs. In contrast, apelin overexpression significantly promoted proliferation and migration and enhanced FGFR1 phosphorylation in J82 cells, and increased the total angiogenesis length in HUVECs, but this effects were effectively mitigated by transfection of the cells with FGF2 siRNA or FGFR1 siRNA. CONCLUSIONS High expression of apelin promotes J82 cell proliferation and migration and HUVEC angiogenesis by promoting activation of the FGF2/FGFR1 pathway.
Humans ; Urinary Bladder Neoplasms/blood supply* ; Receptor, Fibroblast Growth Factor, Type 1/metabolism* ; Cell Proliferation ; Cell Movement ; Fibroblast Growth Factor 2/metabolism* ; Neovascularization, Pathologic ; Human Umbilical Vein Endothelial Cells ; Cell Line, Tumor ; Signal Transduction ; Apelin ; Intercellular Signaling Peptides and Proteins/genetics* ; Female ; Male ; Angiogenesis

Objective To investigate the effects of miR-130b-5p targeting E26 transformation specific-1(ETS1)on proliferation,migration and invasion of prostatic cancer(PCa)cells and its mechanism. Methods The mRNA transcription level of miR-130b-5p gene in PCa tissues,adjacent tissues,(LNCap,PC-3,DU-145)and normal prostate cells(RPWE-1)PCa cells was measured by qRT-PCR,and the expression of ETS1 protein in PCa cells was detected by Western blot. Bioinformatics,fluorescein experiment,qRT-PCR and Western blot were used to predict and verify the targeting relationship between miR-130b-5p and ETS1. PC-3 cells were divided into control group(without any treatment),mimic group(transfected with miR-130b-5p mimic)and mimic + ETS1 group(transfected with miR-130b-5p mimic + pcDNA-ETS1). The cells were detected for the proliferation and viability by clone formation assay and CCK-8 respectively,measured for the migration and invasion by scratch test and Transwell chamber assay,and detected for the expression of invasion-related proteins and PI3K/AKT/mTOR pathway-related proteins by Western blot. Results The transcription level of miR-130b-5p mRNA in PCa tissues was significantly lower than that in adjacent tissues(t = 12. 450,P < 0. 001);Compared with RPWE-1 cells,the transcription level of miR-130b-5p mRNA in LNCap,PC-3 and DU-145 cells decreased significantly(t = 4. 463,7. 103 and 5. 741,P = 0. 001 2,< 0. 001 and < 0. 001,respectively),while the expression level of ETS1protein increased significantly(t = 4. 850,9. 325 and 7. 723,P = 0. 008,< 0. 001 and = 0. 002,respectively). miR-130-5p targeted and negatively regulated the expression of ETS1. Compared with the control group,the cloning rate,viability and scratch healing rate of cells in mimic group decreased significantly(t = 11. 370,10. 640 and 15. 660,respectively,each P < 0. 001),the number of invasive cells decreased significantly(t = 10. 160,P < 0. 001),the expression levels of matrix metalloproteinase-2(MMP-2),MMP-9 and vimentin decreased significantly(t = 15. 120,9. 992 and 12. 600,P < 0. 001,< 0. 001 and = 0. 002,respectively),while the expression level of E-cadherin increased significantly(t = 6. 928,P < 0. 001),and the phosphorylation levels of phosphatidylinositol-3 kinase(PI3K),protein kinase B(AKT)and mammalian target of rapamycin(mTOR)decreased significantly(t = 7. 746,8. 041 and 11. 510,P = 0. 002,0. 002,and < 0. 001,respectively);Compared with mimic group,the cell cloning rate,viability,scratch healing rate significantly increased in mimic + ETS1 group(t = 6. 988,6. 642 and 6. 660,respectively,each P < 0. 001),the number of invasive cells significantly increased(t = 4. 082,P = 0. 002),the expression levels of MMP-2,MMP-9 and vimentin proteins were significantly up-regulated(t = 10. 410,6. 754 and 8. 521,P = 0. 002,0. 003 and 0. 002,respectively),however,the expression level of E-cadherin was significantly down-regulated(t = 4. 648,P < 0. 01),and the phosphorylation levels of PI3K,AKT and mTOR were significantly up-regulated(t = 4. 850,4. 323 and 10. 840,P = 0. 008,0. 008 and < 0. 001,respectively)Conclusion miR-130b-5p targets ETS1 to inhibit the proliferation,migration and invasion of PC-3 cells,which may be through the regulation of PI3K/AKT/mTOR signaling pathway.