Sonoporation mediated transduction of pDNA/siRNA into joint Synovium in vivo
Sonoporation mediated transduction of pDNA/siRNA into joint Synovium in vivo
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DOI：
10.1002/jor.20392
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发表时间：
2007-10-01
影响因子：
2.8
通讯作者：
Kubo, Toshikazu
中科院分区：
文献类型：
--
作者：
Saito, Masazumi;Mazda, Osam;Kubo, Toshikazu
本研究的目的是探讨声孔法在体内将质粒DNA(PDNA)和小干扰RNA(SiRNA)导入关节组织中的有效性。将pGEG.GL3与微泡混合后注射到大鼠膝关节内。超声检查经皮进行。注射后3d，用荧光素酶测定法和RT-PCR法检测滑膜组织中GL3的表达。将针对GL3的siRNA(SiGL3)或非特异性的siRNA与pGEG.GL3混合，通过声学转导的方法进行转化。将针对EGFP的siEGFP(SiEGFP)基因导入转基因大鼠膝关节，检测内源性基因的基因沉默效果。为了确定转导的siRNA的定位，将荧光标记的siRNA转导到关节中。超声手术组大鼠滑膜组织中GL3的表达明显增强。该基因的表达仅见于膝关节滑膜。共转导siGL3可显著抑制GL3的表达，但非特异性siRNA不能抑制GL3的表达。超声修饰siEGFP使转基因大鼠滑膜表层绿色荧光减弱。荧光标记的siRNA位于膝盖骨、股骨和胫骨周围的滑膜中。超声修复被认为是一种新型的基因转导方法，其优点是侵袭性极小。在这项研究中，我们证明了PDNA/siRNA可以通过超声手术特异性地转导到关节滑膜中。本方法可用于关节疾病的核酸治疗。(C)2007年整形外科研究会。由威利期刊出版公司出版。
The purpose of this study was to investigate the usefulness of sonoporation method on in vivo transduction of plasmid DNA (pDNA) and small interfering RNA (siRNA) into joint tissue. pGEG.GL3 plasmid was mixed with microbubble and injected into knee joints of rats. Ultrasound sonication was performed percutaneously. Three days after injection, GL3 expression of synovial tissue was determined by luciferase assay and RT-PCR. siRNA specific for GL3 (siGL3) or nonspecific siRNA were mixed with pGEG.GL3 plasmid and transduced by sonoporation. siRNA specific for EGFP (siEGFP) was transduced into the knee joints of EGFP transgenic rats, and gene silencing effects for endogenous gene were examined. To determine the localization of transduced siRNA, fluorescently labeled siRNA was transduced into joints. The expression of GL3 in the synovium was significantly enhanced by sonoporation. The gene expression was only seen in the synovium of the knee joint. The expression of GL3 was remarkably suppressed by co-transduction of siGL3, but not suppressed by nonspecific siRNA. siEGFP transduced by sonoporation attenuated green fluorescence on the surface layer of synovium of EGFP transgenic :rats. The fluorescently labeled siRNA was seen in the synovium around the patella, femur, and tibia. Sonoporation is examined as a recent, novel, gene transduction method, and the advantage of this technique is minimal invasiveness. In this study, we showed that pDNA/siRNA can be transduced specifically into the joint synovium using sonoporation. The present method may be useful in nucleic acid therapy for joint disorders. (c) 2007 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.