CAJ | 학술논문

本研究用细菌内同源重组的方法构建了针对VEGF受体FLT-1,KDR的siRNA表达重组腺病毒AdH1-siRNA/FLT-1和AdH1-siRNA/KDR,感染HUVEC细胞,观察两种病毒对HUVEC细胞体外增殖和形成微血管的干扰作用.通过RT-PCR实验表明, AdH1-siRNA/FLT-1和AdH1-siRNA/KDR均可特异性地下调血管内皮细胞(HUVEC)中的 FLT-1mRNA 和 KDR mRNA 水平.与对照组相比,FLT-1的 mRNA 水平下降为40%,KDR 的mRNA水平下降为52%. AdH1-siRNA/FLT-1和AdH1-siRNA/KDR对HUVEC细胞增殖均有干扰作用,加入病毒9 d后,对照组的细胞平均计数为31.5×104/μL,AdH1-siRNA/FLT-1干扰病毒对照组为28×104/μL,AdH1-siRNA/KDR干扰组为21.5×104/μL. AdH1-siRNA/FLT-1和AdH1-siRNA/KDR对HUVEC细胞在Matrigel上形成微血管均有干扰作用,对照组每HPF形成微血管数量为10.75条,AdH1-siRNA/FLT-1干扰病毒对照组为10.25条,实验组AdH1-siRNA/KDR每HPF形成微血管数量为7条.证明AdH1-siRNA/FLT-1,AdH1-siRNA/KDR均可干扰血管形成.
본연구용세균내동원중조적방법구건료침대VEGF수체FLT-1,KDR적siRNA표체중조선병독AdH1-siRNA/FLT-1화AdH1-siRNA/KDR,감염HUVEC세포,관찰량충병독대HUVEC세포체외증식화형성미혈관적간우작용.통과RT-PCR실험표명, AdH1-siRNA/FLT-1화AdH1-siRNA/KDR균가특이성지하조혈관내피세포(HUVEC)중적 FLT-1mRNA 화 KDR mRNA 수평.여대조조상비,FLT-1적 mRNA 수평하강위40%,KDR 적mRNA수평하강위52%. AdH1-siRNA/FLT-1화AdH1-siRNA/KDR대HUVEC세포증식균유간우작용,가입병독9 d후,대조조적세포평균계수위31.5×104/μL,AdH1-siRNA/FLT-1간우병독대조조위28×104/μL,AdH1-siRNA/KDR간우조위21.5×104/μL. AdH1-siRNA/FLT-1화AdH1-siRNA/KDR대HUVEC세포재Matrigel상형성미혈관균유간우작용,대조조매HPF형성미혈관수량위10.75조,AdH1-siRNA/FLT-1간우병독대조조위10.25조,실험조AdH1-siRNA/KDR매HPF형성미혈관수량위7조.증명AdH1-siRNA/FLT-1,AdH1-siRNA/KDR균가간우혈관형성.
This study used the homologous recombination technique constructed a recombinant adenovirus AdH1-siRNA/FLT-1 and AdH1-siRNA/KDR targeting FLT-1 and KDR infects human endothelial cells ( HUVEC ) , observation the effect of the adenovirus interfere the proliferation of HUVEC in vitro and the tube formation. Reverse transcription-PCR showed that this adenovirus could specifically decrease level of the FLT-1 and KDR mRNA of HUVEC. Compared with control group,the FLT-1 and KDR mRNA expression of decreased to 40% and 52%. AdH1-siRNA/FLT-1 and KDR can efficiently interfere the proliferation of HUVEC,9 d after infection of adenovirus,the average cell number of control group and vector group are 31. 5×104/μL and 28×104/μL,that of groups respectively infected with AdH1-siRNA/VEGF are 21. 5×104/μL,and can efficiently interfere the tube formation of HUVEC on matrigel. Averagely,the tube number of every HPF is 10. 75 in the control group,and 10. 25 in the vector group. In the group treated with AdH1-siRNA/VEGF,there are 7 tubes in the HPE. It is sowed that AdH1-siRNA/FLT-1 and AdH1-siRNA/KDR can also interfere tube formation.