BACKGROUND microRNA-627-5p(miR-627-5p)dysregulation has been observed in several cancer types,such as hepatocellular carcinoma,oral squamous cell carcinoma,glioblastoma multiforme,and gastric cancer.The biological fun...BACKGROUND microRNA-627-5p(miR-627-5p)dysregulation has been observed in several cancer types,such as hepatocellular carcinoma,oral squamous cell carcinoma,glioblastoma multiforme,and gastric cancer.The biological function of miR-627-5p in colorectal cancer(CRC)growth and metastasis is yet unclear.AIM To investigate the effects of miR-627-5p on the malignant biological properties of colorectal malignant tumour cells by targeting Wnt2.METHODS The levels of miR-627-5p in colorectal tumour tissues were assessed in Gene Expression Omnibus datasets.In order to identify Wnt2 transcript expression in CRC tissues,quantitative real-time polymerase chain reaction(qRT-PCR)analysis was used.Luciferase reporter tests were used to explore whether miR-627-5p might potentially target Wnt2.Wnt2 transcript and protein levels were detected in CRC cells with high miR-627-5p expression.To learn more about how miR-627-5p affects CRC development,migration,apoptosis,and invasion,functional experiments were conducted.Cotransfection with the overexpression vector of Wnt2 and miR-627-5p mimics was utilized to verify whether overexpression of Wnt2 could cancel the impact of miR-627-5p in CRC.Western blot and qRT-PCR were conducted to investigate the effects of miR-627-5p on the Wnt/β-catenin signalling pathway.RESULTS miR-627-5p was notably decreased in colorectal tumour tissues,while the gene level of Wnt2 was notably upregulated.A dual luciferase reporter assay revealed that miR-627-5p specifically targets the 3’-untranslated regions of Wnt2 and miR-627-5p upregulation markedly reduced the protein and gene expression of Wnt2 in CRC cells.In vitro gain-of-function assays displayed that miR-627-5p overexpression decreased CRC cells’capabilities to invade,move,and remain viable while increasing apoptosis.Wnt2 overexpression could reverse the suppressive functions of miR-627-5p.Moreover,upregulation of miR-627-5p suppressed the transcript and protein levels of the downstream target factors in the canonical Wnt/β-catenin signalling,such as c-myc,CD44,β-catenin,and cyclinD1.CONCLUSION miR-627-5p acts as a critical inhibitory factor in CRC,possibly by directly targeting Wnt2 and negatively modulating the Wnt/β-catenin signalling,revealing that miR-627-5p could be a possible treatment target for CRC.展开更多
BACKGROUND Exosomal miRNAs play crucial roles in many central nervous system diseases.Cerebral small vessel disease(CVSD)is a small vessel disease that is affected by various factors.This study aimed to investigate th...BACKGROUND Exosomal miRNAs play crucial roles in many central nervous system diseases.Cerebral small vessel disease(CVSD)is a small vessel disease that is affected by various factors.This study aimed to investigate the role of exosomal miR-320e in the Wnt/β-catenin pathway stimulated by oxidative stress and assess its clinical correlation with psychiatric symptoms in patients with CVSD.AIM To explore whether exosomal miR-320e could suppress the Wnt/β-catenin pathway and play a protective role in CVSD progression,as well as examine its potential correlation with cognitive impairment and depression in patients with CVSD.METHODS Differentially expressed exosomal miRNAs were filtered by sequencing plasma exosomes from patients with CVSD and healthy controls.Bioinformatics and dual luciferase analyses were used to confirm the binding of miR-320e to Wnt2,and the mRNA and protein levels of downstream components in the Wnt/β-catenin pathway were evaluated when overexpressed or with knockdown of miR-320e under H2O2-induced oxidative stress.In addition,Wnt2-targeting siRNA was used to confirm the role of miR-320e in the Wnt2-mediated inhibition of the Wnt/β-catenin pathway.A retrospective analysis was conducted among patients with CVSD to confirm the correlation between miR-320e expression and the severity of cognitive impairment and depression,which were quantified using the Montreal Cognitive Assessment(MoCA)/Executive Function Assessment(EFA),and the Hamilton Depression Scale(HAMD)/Beck Depression Inventory(BDI),respectively.RESULTS High-throughput sequencing revealed that exosomal miR-320e was downregulated in patients with CVSD.Bioinformatics analysis and dual-luciferase reporter gene experiments showed that exosomal miR-320e inhibited the Wnt/β-catenin pathway in response to oxidative stress by targeting the 3'noncoding region of Wnt2.Uptake of exosomes carrying miR-320e into endothelial cells could also target Wnt2 and inhibit the Wnt2/β-catenin pathway.Elevated miR-320e expression may protect patients with CVSD from relatively severe cognitive impairment and depression,as it was found to have a positive correlation with the MoCA/EFA and HAMD/BDI scores.CONCLUSION Our results suggest that exosomal miR-320e suppresses the Wnt/β-catenin pathway and may play a protective role in CVSD progression.展开更多
目的探讨分析microRNA-21(miR-21)靶向调控Wnt2基因对肝癌细胞HepG2增殖与迁移的影响。方法采用实时荧光定量PCR法检测肝癌细胞HepG2及正常肝细胞株LO2 miR-21的表达情况。对HepG2细胞转染miR-21抑制剂,分析转染后抑制剂组与对照组miR-2...目的探讨分析microRNA-21(miR-21)靶向调控Wnt2基因对肝癌细胞HepG2增殖与迁移的影响。方法采用实时荧光定量PCR法检测肝癌细胞HepG2及正常肝细胞株LO2 miR-21的表达情况。对HepG2细胞转染miR-21抑制剂,分析转染后抑制剂组与对照组miR-21的表达情况、细胞增殖、迁移及凋亡情况,检测两组细胞Wnt2蛋白表达水平,并采用双荧光素酶报告基因实验验证miR-21与Wnt2基因的关系。计量资料两组间比较采用t检验。结果HepG2细胞miR-21相对表达水平明显高于LO2细胞(1.978±0.035 vs 1.586±0.022,t=16.424,P<0.05)。转染miR-21抑制剂后,抑制剂组miR-21相对表达水平较对照组显著降低(0.857±0.017 vs 1.684±0.039,t=33.669,P<0.05)。转染miR-21抑制剂24、48、72 h后,抑制剂组HepG2细胞增殖能力均较对照组显著降低(P值均<0.05);抑制剂组穿过Tranwell小室的细胞数显著低于对照组(83.72±15.06 vs 147.85±20.64,t=4.347,P<0.05);抑制剂组细胞凋亡率明显高于对照组(25.67%±3.95%vs 10.27%±2.14%,t=5.937,P<0.05)。抑制剂组HepG2细胞的Wnt2相对表达水平明显低于对照组(0.862±0.127 vs 1.306±0.218,t=3.048,P<0.05)。TargetScan软件显示,miR-21抑制剂可显著抑制野生型Wnt2-3′UTR质粒转染细胞的荧光素酶活性(0.972±0.102 vs 0.612±0.092,t=4.219,P<0.05),而对突变型Wnt2-3′UTR质粒转染细胞的荧光素酶活性并无明显影响(0.982±0.093 vs 0.911±0.128,t=0.972,P>0.05)。结论抑制miR-21表达可有效抑制HepG2细胞增殖和迁移,促进HepG2细胞凋亡,并抑制Wnt信号通路的过度激活,可能成为肝癌治疗的潜在靶基因之一。展开更多
基金Supported by the National Key Development Plan for Precision Medicine Research,No.2017YFC0910002.
文摘BACKGROUND microRNA-627-5p(miR-627-5p)dysregulation has been observed in several cancer types,such as hepatocellular carcinoma,oral squamous cell carcinoma,glioblastoma multiforme,and gastric cancer.The biological function of miR-627-5p in colorectal cancer(CRC)growth and metastasis is yet unclear.AIM To investigate the effects of miR-627-5p on the malignant biological properties of colorectal malignant tumour cells by targeting Wnt2.METHODS The levels of miR-627-5p in colorectal tumour tissues were assessed in Gene Expression Omnibus datasets.In order to identify Wnt2 transcript expression in CRC tissues,quantitative real-time polymerase chain reaction(qRT-PCR)analysis was used.Luciferase reporter tests were used to explore whether miR-627-5p might potentially target Wnt2.Wnt2 transcript and protein levels were detected in CRC cells with high miR-627-5p expression.To learn more about how miR-627-5p affects CRC development,migration,apoptosis,and invasion,functional experiments were conducted.Cotransfection with the overexpression vector of Wnt2 and miR-627-5p mimics was utilized to verify whether overexpression of Wnt2 could cancel the impact of miR-627-5p in CRC.Western blot and qRT-PCR were conducted to investigate the effects of miR-627-5p on the Wnt/β-catenin signalling pathway.RESULTS miR-627-5p was notably decreased in colorectal tumour tissues,while the gene level of Wnt2 was notably upregulated.A dual luciferase reporter assay revealed that miR-627-5p specifically targets the 3’-untranslated regions of Wnt2 and miR-627-5p upregulation markedly reduced the protein and gene expression of Wnt2 in CRC cells.In vitro gain-of-function assays displayed that miR-627-5p overexpression decreased CRC cells’capabilities to invade,move,and remain viable while increasing apoptosis.Wnt2 overexpression could reverse the suppressive functions of miR-627-5p.Moreover,upregulation of miR-627-5p suppressed the transcript and protein levels of the downstream target factors in the canonical Wnt/β-catenin signalling,such as c-myc,CD44,β-catenin,and cyclinD1.CONCLUSION miR-627-5p acts as a critical inhibitory factor in CRC,possibly by directly targeting Wnt2 and negatively modulating the Wnt/β-catenin signalling,revealing that miR-627-5p could be a possible treatment target for CRC.
文摘BACKGROUND Exosomal miRNAs play crucial roles in many central nervous system diseases.Cerebral small vessel disease(CVSD)is a small vessel disease that is affected by various factors.This study aimed to investigate the role of exosomal miR-320e in the Wnt/β-catenin pathway stimulated by oxidative stress and assess its clinical correlation with psychiatric symptoms in patients with CVSD.AIM To explore whether exosomal miR-320e could suppress the Wnt/β-catenin pathway and play a protective role in CVSD progression,as well as examine its potential correlation with cognitive impairment and depression in patients with CVSD.METHODS Differentially expressed exosomal miRNAs were filtered by sequencing plasma exosomes from patients with CVSD and healthy controls.Bioinformatics and dual luciferase analyses were used to confirm the binding of miR-320e to Wnt2,and the mRNA and protein levels of downstream components in the Wnt/β-catenin pathway were evaluated when overexpressed or with knockdown of miR-320e under H2O2-induced oxidative stress.In addition,Wnt2-targeting siRNA was used to confirm the role of miR-320e in the Wnt2-mediated inhibition of the Wnt/β-catenin pathway.A retrospective analysis was conducted among patients with CVSD to confirm the correlation between miR-320e expression and the severity of cognitive impairment and depression,which were quantified using the Montreal Cognitive Assessment(MoCA)/Executive Function Assessment(EFA),and the Hamilton Depression Scale(HAMD)/Beck Depression Inventory(BDI),respectively.RESULTS High-throughput sequencing revealed that exosomal miR-320e was downregulated in patients with CVSD.Bioinformatics analysis and dual-luciferase reporter gene experiments showed that exosomal miR-320e inhibited the Wnt/β-catenin pathway in response to oxidative stress by targeting the 3'noncoding region of Wnt2.Uptake of exosomes carrying miR-320e into endothelial cells could also target Wnt2 and inhibit the Wnt2/β-catenin pathway.Elevated miR-320e expression may protect patients with CVSD from relatively severe cognitive impairment and depression,as it was found to have a positive correlation with the MoCA/EFA and HAMD/BDI scores.CONCLUSION Our results suggest that exosomal miR-320e suppresses the Wnt/β-catenin pathway and may play a protective role in CVSD progression.
文摘目的探讨分析microRNA-21(miR-21)靶向调控Wnt2基因对肝癌细胞HepG2增殖与迁移的影响。方法采用实时荧光定量PCR法检测肝癌细胞HepG2及正常肝细胞株LO2 miR-21的表达情况。对HepG2细胞转染miR-21抑制剂,分析转染后抑制剂组与对照组miR-21的表达情况、细胞增殖、迁移及凋亡情况,检测两组细胞Wnt2蛋白表达水平,并采用双荧光素酶报告基因实验验证miR-21与Wnt2基因的关系。计量资料两组间比较采用t检验。结果HepG2细胞miR-21相对表达水平明显高于LO2细胞(1.978±0.035 vs 1.586±0.022,t=16.424,P<0.05)。转染miR-21抑制剂后,抑制剂组miR-21相对表达水平较对照组显著降低(0.857±0.017 vs 1.684±0.039,t=33.669,P<0.05)。转染miR-21抑制剂24、48、72 h后,抑制剂组HepG2细胞增殖能力均较对照组显著降低(P值均<0.05);抑制剂组穿过Tranwell小室的细胞数显著低于对照组(83.72±15.06 vs 147.85±20.64,t=4.347,P<0.05);抑制剂组细胞凋亡率明显高于对照组(25.67%±3.95%vs 10.27%±2.14%,t=5.937,P<0.05)。抑制剂组HepG2细胞的Wnt2相对表达水平明显低于对照组(0.862±0.127 vs 1.306±0.218,t=3.048,P<0.05)。TargetScan软件显示,miR-21抑制剂可显著抑制野生型Wnt2-3′UTR质粒转染细胞的荧光素酶活性(0.972±0.102 vs 0.612±0.092,t=4.219,P<0.05),而对突变型Wnt2-3′UTR质粒转染细胞的荧光素酶活性并无明显影响(0.982±0.093 vs 0.911±0.128,t=0.972,P>0.05)。结论抑制miR-21表达可有效抑制HepG2细胞增殖和迁移,促进HepG2细胞凋亡,并抑制Wnt信号通路的过度激活,可能成为肝癌治疗的潜在靶基因之一。
