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@#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.
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Osteoarthritis (OA) is a chronic health condition. MicroRNAs (miRs) are critical in chondrocyte apoptosis in OA. We aimed to investigate the mechanism of miR-130b in OA progression. Bone marrow mesenchymal stem cells (BMSCs) and chondrocytes were first extracted. Chondrogenic differentiation of BMSCs was carried out and verified. Chondrocytes were stimulated with interleukin (IL)-1β to imitate OA condition in vitro. The effect of miR-130b on the viability, inflammation, apoptosis, and extracellular matrix of OA chondrocytes was studied. The target gene of miR-130b was predicted and verified. Rescue experiments were performed to further study the underlying downstream mechanism of miR-130b in OA. miR-130b first increased and drastically reduced during chondrogenic differentiation of BMSCs and in OA chondrocytes, respectively, while IL-1β stimulation resulted in increased miR-130b expression in chondrocytes. miR-130b inhibitor promoted chondrogenic differentiation of BMSCs and chondrocyte growth and inhibited the levels of inflammatory factors. miR-130b targeted SOX9. Overexpression of SOX9 facilitated BMSC chondrogenic differentiation and chondrocyte growth, while siRNA-SOX9 contributed to the opposite trends. Silencing of SOX9 significantly attenuated the pro-chondrogenic effects of miR-130b inhibitor on BMSCs. Overall, miR-130b inhibitor induced chondrogenic differentiation of BMSCs and chondrocyte growth by targeting SOX9.
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MicroRNAs/genetics , Mesenchymal Stem Cells , Down-Regulation , Cell Differentiation , Cells, CulturedABSTRACT
Osteoarthritis (OA) is a chronic health condition. MicroRNAs (miRs) are critical in chondrocyte apoptosis in OA. We aimed to investigate the mechanism of miR-130b in OA progression. Bone marrow mesenchymal stem cells (BMSCs) and chondrocytes were first extracted. Chondrogenic differentiation of BMSCs was carried out and verified. Chondrocytes were stimulated with interleukin (IL)-1β to imitate OA condition in vitro. The effect of miR-130b on the viability, inflammation, apoptosis, and extracellular matrix of OA chondrocytes was studied. The target gene of miR-130b was predicted and verified. Rescue experiments were performed to further study the underlying downstream mechanism of miR-130b in OA. miR-130b first increased and drastically reduced during chondrogenic differentiation of BMSCs and in OA chondrocytes, respectively, while IL-1β stimulation resulted in increased miR-130b expression in chondrocytes. miR-130b inhibitor promoted chondrogenic differentiation of BMSCs and chondrocyte growth and inhibited the levels of inflammatory factors. miR-130b targeted SOX9. Overexpression of SOX9 facilitated BMSC chondrogenic differentiation and chondrocyte growth, while siRNA-SOX9 contributed to the opposite trends. Silencing of SOX9 significantly attenuated the pro-chondrogenic effects of miR-130b inhibitor on BMSCs. Overall, miR-130b inhibitor induced chondrogenic differentiation of BMSCs and chondrocyte growth by targeting SOX9.
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@#AIM: To study the expression and cancer-promoting mechanism of miR-130b in human retinoblastoma(RB).<p>METHODS: Detected the expression levels of miR-130b in human RB carcinoma tissues, adjacent tissues and human RB cell lines(HXO-Rb44 and Y79)by qRT-PCR; detected the expression levels of PTEN in HXO-Rb44 and Y79 cells by qRT-PCR, Western Blot and immunofluorescence; verified the target relationship between miR-130b and PTEN by dual-luciferase reporter gene test; the co-transfection test was used to investigate the relationship between PTEN and miR-130b on the expression of PI3K/Akt signaling pathway in RB cell line.<p>RESULTS: The expression level of miR-130b in cancer tissue of RB was significantly higher than that of paracancerous tissue(<i>P</i><0.05). Compared with axben-181 cells, the expression level of miR-130b in HXO-Rb44 and Y79 cells was significantly increased(<i>P</i><0.05). Compared with RB cancer tissue, the expression level of PTEN in its paracancerous tissue was significantly increased(<i>P</i><0.05). The expression level of miR-130b was negatively correlated with the expression level of PTEN(<i>P</i><0.001). The mRNA and protein expression levels of PTEN in HXO-Rb44 cells overexpressing miR-130b were significantly reduced, while the mRNA and protein expression levels of PTEN in Y79 cells after miR-130b interference were significantly increased(<i>P</i><0.05). Compared with miR-130b mimics+PTEN-NC group, the luciferase activity of miR-130b mimics+wt-PTEN group was significantly reduced(<i>P</i><0.05).In the HXO-Rb44 cells co-transfected with miR-130b mimics+PTEN-NC, the expression levels of p-Akt 308 and p-Akt 473 protein were significantly increased(<i>P</i><0.05), while the expression levels of PTEN protein were significantly decreased(<i>P</i><0.05). In the HXO-Rb44 cells co-transfected with miR-130b mimics+PTEN, no significant changes were observed in the above three proteins.<p>CONCLUSION: miR-130b is highly expressed in RB tissues and cell lines. PTEN is the target gene of miR-130b, and miR-130b may negatively regulate PTEN to affect the expression of PI3K/Akt signaling pathway and ultimately play a role in promoting cancer.
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Objective To further understand potential mechanism of miRNAs in bladder cancer .Mtehods Human microarray was used to analyze the expression of miRNAs in four pair human BC cancer tissues and adjacent normal tissues.Then RT-Qpcr was used to verified that the expression of two most up -regulated miRNAs and target genes for results of miRNA/Mrna microarray.Subsequently, it was deduced that miR-130b-3p could target at PTEN through a bioinformatics approach and dual-luciferase reporter assay.CCK8, EDU, flow cytometry, wound healing, Transwell and cytoskeleton assay were applied to prove that miR-130b could affect proliferation, apoptosis, migration and invasion of BC cells.The effects of PTEN regula-ted by miR-130b-3p on the key target protein expression of PI 3K/AKT and integrin β1/FAK signaling pathway were determined by Western blot.Resulst miR-130b-3p was significantly up-regulated and nega-tively correlated with PTEN expression in clinical bladder cancer specimens as compared with normal urothelial tissue.miR-130b-3p mimics could down-regulate PTEN expression, which leads to the activation of PI3K/AKT and integrinβ1/FAK signaling pathway related to cell proliferation , migration and invasion in bladder cancer EJ cells.When the cells were transfected with miR-130b-3p inhibitors, they could be sur- veyed with rearranging cytoskeleton.Conclusions miR-130b/PTEN may be used as a marker for bladder cancer.
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Objective To study the effects of miR-130b on the neuronal migration in the developing embryonic mice cortex.Methods Pregnancy E15.5 mice were selected and plasmid of miR-130b was injected into the lateral ventricle of the embryonic brain.Two days after eletroporation,murine embryos were collected and cut into frozon coronal slices,then surveyed the neuronal migration under the fluorescence microscope.Results The neuronal migration rates were higher in miR-130b overexpression embryonic cortex,of which 75.1％ eletroporated neurons migrated into MZ,compared with the 22.1％ in control group.On the contrary,only 7.9％ eletroporated neurons stayed in the VZ/ SVZ in the experimental group,compared with the 69.3％ in control group.Conclusions miR-130b accelerates the neuronal migration in the developing embryonic mice cortex,its regulational role is worth studying.
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[Summary] The aim of this study was to detect the levels of serum miR-130b expression in patients with type 2 diabetes mellitus and to analyze their correlation with diabetic renal damage. 243 patients with type 2 diabetes mellitus were divided into three groups according to urinary albumin/creatinine ratio ( UACR ): normoalbuminuria group (UACR300 mg/g, n=54). The levels of serum miR-130b were validated by realtime polymerase chain reaction. Serum transforming growth factor-β1 (TGF-β1) and tumor necrosis factor-α(TNF-α) were determined by enzyme-linked immunosorbent assay ( ELISA) in all patients and 59 healthy volunteers. Compared with control group, the level of serum miR-130b in the type 2 diabetes mellitus group were significantly decreased, gradually with the increases of UACR. The level of serum miR-130b was inversely correlated with blood urea nitrogen ( r=-0. 295, P<0.05), serum creatinine(r=-0. 316, P<0. 05), UACR(r=-0. 463, P<0. 05), but positively related to the estimated glomerular filtration rate(r=0. 367, P<0. 01). The level of serum miR-130b was also negatively correlated to homeostasis model assessment for insulin resistance, triglyceride, low density lipoprotein-cholesterol, TNF-α, and TGF-β1 (r=-0. 257,-0. 345,-0. 242,-0. 562,-0. 622, all P<0. 01). The present study indicates that serum miR-130b might be a potential new biomarker for early diagnosis of diabetic nephropathy. Serum miR-130b might be involved in the pathogenesis of diabetic nephropathy.