目的:利用羟基磷灰石纳米颗粒(hydroxyapatite nanoparticle,HAT)携带构建的神经营养因子-3(neurotrophic factor-3,NT-3)-绿色荧光蛋白基因(enhancement type Green fluorescent protein C2,pEGFPC2),通过耳蜗灌注方法转染兴奋毒性损...目的:利用羟基磷灰石纳米颗粒(hydroxyapatite nanoparticle,HAT)携带构建的神经营养因子-3(neurotrophic factor-3,NT-3)-绿色荧光蛋白基因(enhancement type Green fluorescent protein C2,pEGFPC2),通过耳蜗灌注方法转染兴奋毒性损伤后的豚鼠耳蜗螺旋神经节细胞(spinal ganglion cells,SGCs),观察pEGFPC2-NT3的表达及其对耳蜗螺旋神经节细胞的保护作用。方法:构建携带绿色荧光蛋白报告基因的重组质粒pEGFPC2-NT3。通过耳蜗灌注海人酸(kainic acid,KA)建立豚鼠耳蜗兴奋性损伤模型,在给KA1周后利用HAT携带重组质粒进行耳蜗灌注以转染耳蜗螺旋神经节细胞,免疫组化法观察转染后1周NT-3的表达及4周后电镜下螺旋神经节细胞形态学变化,同时观察对听觉脑干诱发电位(auditory brain-stem response,ABR)的影响。结果:成功构建豚鼠耳蜗兴奋损伤模型。灌注重组质粒后1周免疫组化法观察到螺旋神经节细胞胞浆内NT-3蛋白表达。4周后电镜下螺旋神经节细胞形态学损害减轻,ABR检测听功能较兴奋毒性损害后有恢复。结论:在豚鼠耳蜗灌注KA造成耳蜗兴奋性损伤后第7天,经耳蜗鼓阶转染羟基磷灰石纳米颗粒介导的NT-3基因仍可减轻KA对耳蜗螺旋神经节细胞的兴奋性毒性损伤。展开更多
Transfection efficiency of hydroxyapatite nanoparticles(HAnps)is relative to the particle size,morphology,surface charge,surface modifier and so on.This study prepared HAnps with doped Tb/Mg by hydrothermal synthesis ...Transfection efficiency of hydroxyapatite nanoparticles(HAnps)is relative to the particle size,morphology,surface charge,surface modifier and so on.This study prepared HAnps with doped Tb/Mg by hydrothermal synthesis method(HTSM)and investigated the effects of different Tb/Mg contents on the morphology,particle size,surface charge,composition and cellular endocytosis of HAnps.The results showed that Mg-HAnps possessed better dispersion ability than Tb-HAnps.With increasing doping content of Tb/Mg-HAnps,the granularity of Tb-HAnps increased,while that of Mg-HAnps declined.Both particle size and zeta potential of Mg-HAnps were lower than those of Tb-HAnps.7.5%Mg-doping HAnps presented relatively uniform slender rod morphology with average size of30nm,while10%Mg-doping HAnps were prone to agglomeration.Moreover,Mg-HAnps-GFP(green fluorescent protein)endocytosed by MG63cells was dotted in the perinuclear region,while Tb-HAnps were more likely to aggregate.In conclusion,as gene vectors,Mg-HAnps showed enhanced properties compared to Tb-HAnps.展开更多
文摘目的:利用羟基磷灰石纳米颗粒(hydroxyapatite nanoparticle,HAT)携带构建的神经营养因子-3(neurotrophic factor-3,NT-3)-绿色荧光蛋白基因(enhancement type Green fluorescent protein C2,pEGFPC2),通过耳蜗灌注方法转染兴奋毒性损伤后的豚鼠耳蜗螺旋神经节细胞(spinal ganglion cells,SGCs),观察pEGFPC2-NT3的表达及其对耳蜗螺旋神经节细胞的保护作用。方法:构建携带绿色荧光蛋白报告基因的重组质粒pEGFPC2-NT3。通过耳蜗灌注海人酸(kainic acid,KA)建立豚鼠耳蜗兴奋性损伤模型,在给KA1周后利用HAT携带重组质粒进行耳蜗灌注以转染耳蜗螺旋神经节细胞,免疫组化法观察转染后1周NT-3的表达及4周后电镜下螺旋神经节细胞形态学变化,同时观察对听觉脑干诱发电位(auditory brain-stem response,ABR)的影响。结果:成功构建豚鼠耳蜗兴奋损伤模型。灌注重组质粒后1周免疫组化法观察到螺旋神经节细胞胞浆内NT-3蛋白表达。4周后电镜下螺旋神经节细胞形态学损害减轻,ABR检测听功能较兴奋毒性损害后有恢复。结论:在豚鼠耳蜗灌注KA造成耳蜗兴奋性损伤后第7天,经耳蜗鼓阶转染羟基磷灰石纳米颗粒介导的NT-3基因仍可减轻KA对耳蜗螺旋神经节细胞的兴奋性毒性损伤。
基金Project(2015WK3012) supported by the Hunan Provincial Science and Technology Department Project,ChinaProject(81571021) supported by the National Natural Science Foundation of China+2 种基金Project(225) supported by the High Level Health Personnel in Hunan Province,ChinaProject(621020094) supported by the State Key Laboratory of Powder Metallurgy of Central South University,ChinaProject(20160301) supported by New Talent Project of the Third Xiangya Hospital of Central South University,China
文摘Transfection efficiency of hydroxyapatite nanoparticles(HAnps)is relative to the particle size,morphology,surface charge,surface modifier and so on.This study prepared HAnps with doped Tb/Mg by hydrothermal synthesis method(HTSM)and investigated the effects of different Tb/Mg contents on the morphology,particle size,surface charge,composition and cellular endocytosis of HAnps.The results showed that Mg-HAnps possessed better dispersion ability than Tb-HAnps.With increasing doping content of Tb/Mg-HAnps,the granularity of Tb-HAnps increased,while that of Mg-HAnps declined.Both particle size and zeta potential of Mg-HAnps were lower than those of Tb-HAnps.7.5%Mg-doping HAnps presented relatively uniform slender rod morphology with average size of30nm,while10%Mg-doping HAnps were prone to agglomeration.Moreover,Mg-HAnps-GFP(green fluorescent protein)endocytosed by MG63cells was dotted in the perinuclear region,while Tb-HAnps were more likely to aggregate.In conclusion,as gene vectors,Mg-HAnps showed enhanced properties compared to Tb-HAnps.
