Douar A M, Themis M, Sandig V, Friedmann T, Coutelle C
Department of Biochemistry and Molecular Genetics, Imperial College, Medical School at St Mary's London, UK.
Gene Ther. 1996 Sep;3(9):789-96.
In preparation for foetal gene therapy by intra-amniotic gene application, we have investigated the effect of amniotic fluid on several gene transfer systems. In vitro lipofection of embryonically derived 3T3 cells by several of the tested cationic lipids decreases in the presence of human amniotic fluid, while two formulations, Lipid 67 and Tfx-50, remain highly active. As some body fluids are known to inactivate most retroviral vectors, we investigated the influence of amniotic fluid on the efficiency of infection of 3T3 cells by murine leukaemia virus (MoMLV)-based vectors, including amphotropic and ecotropic retrovirus, and a vesicular stomatitis virus G (VSV-G) glycoprotein pseudotyped retroviral vector. All showed a decrease of infectivity from 21 to 56% in the presence of amniotic fluid. The ecotropic retrovirus is the most infectious under normal conditions as well as in amniotic fluid. Our results suggest that intra-amniotic injection may allow efficient gene transfer using either amniotic fluid-resistant cationic lipids or ecotropic retroviral vectors in a murine in vivo model for foetal gene therapy. The VSV-G-pseudotyped vector, although displaying a decrease of infectivity, remains of great interest for gene delivery, because of its broad host range and because of the high virus titers achievable. Finally, a baculovirus-based vector shows no decrease of its infectivity and does not seem to be affected by amniotic fluid but has only low infectivity on the tested foetal fibroblast cell line.
为通过羊膜腔内基因应用进行胎儿基因治疗做准备，我们研究了羊水对几种基因转移系统的影响。在几种测试的阳离子脂质对胚胎来源的3T3细胞进行体外脂质转染时，在人羊水存在的情况下转染效率降低，而两种制剂Lipid 67和Tfx - 50仍保持高活性。由于已知一些体液会使大多数逆转录病毒载体失活，我们研究了羊水对基于小鼠白血病病毒（MoMLV）的载体感染3T3细胞效率的影响，这些载体包括双嗜性和嗜异性逆转录病毒，以及水疱性口炎病毒G（VSV - G）糖蛋白假型逆转录病毒载体。在羊水存在的情况下，所有载体的感染性均降低了21%至56%。嗜异性逆转录病毒在正常条件下以及在羊水中都是最具感染性的。我们的结果表明，在小鼠胎儿基因治疗的体内模型中，羊膜腔内注射可能允许使用抗羊水阳离子脂质或嗜异性逆转录病毒载体进行有效的基因转移。VSV - G假型载体虽然感染性降低，但由于其广泛的宿主范围和可达到的高病毒滴度，对于基因递送仍然具有很大的吸引力。最后，一种基于杆状病毒的载体其感染性没有降低，似乎也不受羊水影响，但对测试的胎儿成纤维细胞系只有低感染性。
