Pharyngeal cartilage morphogenesis is crucial for the formation of craniofacial structures.Cranial neural crest cells are specified at the neural plate border,migrate to pharyngeal arches,and differentiate into pharyn...Pharyngeal cartilage morphogenesis is crucial for the formation of craniofacial structures.Cranial neural crest cells are specified at the neural plate border,migrate to pharyngeal arches,and differentiate into pharyngeal chondrocytes,which subsequently flatten,elongate,and stack like coins during maturation.Although the developmental processes prior to chondrocyte maturation have been extensively studied,their subsequent changes in morphology and organization remain largely elusive.Here,we show that wnt2bb is expressed in the pharyngeal ectoderm adjacent to the chondrogenic precursor cells in zebrafish.Inactivation of Wnt2bb leads to a reduction in nuclearβ-catenin,which impairs chondrogenic precursor proliferation and disrupts chondrocyte morphogenesis and organization,eventually causing a severe shrinkage of pharyngeal cartilages.Moreover,the decrease ofβ-catenin in wnt2bb^(-/-)mutants is accompanied by the reduction of Yap expression.Reactivation of Yap can restore the proliferation of chondrocyte progenitors as well as the proper size,shape,and stacking of pharyngeal chondrocytes.Our findings suggest that Wnt/β-catenin signaling promotes Yap expression to regulate pharyngeal cartilage formation in zebrafish.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support of the National Natural Science Foundation of China(32025014 and 32330029 to Q.W.)the National Key Research and Development Program of China(2020YFA0804000 to Q.W.)+2 种基金Guangdong Excellent Youth Team Project(2024B1515040019 to Q.W.)Guangzhou Science and Technology Plan Project(202201010323 to X.H.)the Fundamental Research Funds for the Central Universities(to Q.W.).
文摘Pharyngeal cartilage morphogenesis is crucial for the formation of craniofacial structures.Cranial neural crest cells are specified at the neural plate border,migrate to pharyngeal arches,and differentiate into pharyngeal chondrocytes,which subsequently flatten,elongate,and stack like coins during maturation.Although the developmental processes prior to chondrocyte maturation have been extensively studied,their subsequent changes in morphology and organization remain largely elusive.Here,we show that wnt2bb is expressed in the pharyngeal ectoderm adjacent to the chondrogenic precursor cells in zebrafish.Inactivation of Wnt2bb leads to a reduction in nuclearβ-catenin,which impairs chondrogenic precursor proliferation and disrupts chondrocyte morphogenesis and organization,eventually causing a severe shrinkage of pharyngeal cartilages.Moreover,the decrease ofβ-catenin in wnt2bb^(-/-)mutants is accompanied by the reduction of Yap expression.Reactivation of Yap can restore the proliferation of chondrocyte progenitors as well as the proper size,shape,and stacking of pharyngeal chondrocytes.Our findings suggest that Wnt/β-catenin signaling promotes Yap expression to regulate pharyngeal cartilage formation in zebrafish.
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。
