Huang Wei, Queen Nicholas J, Cao Lei
Department of Cancer Biology and Genetics, The Ohio State University, Columbus, OH, USA.
The Comprehensive Cancer Center, The Ohio State University, Columbus, OH, USA.
Methods Mol Biol. 2019;1950:389-405. doi: 10.1007/978-1-4939-9139-6_23.
Recombinant adeno-associated virus (rAAV) vectors are attractive vehicles for gene therapy. Yet, it is challenging to genetically manipulate adipose tissue in adults due to the low transduction efficiency of naturally occurring AAV serotypes. We recently demonstrated that a novel engineered hybrid serotype Rec2 achieves high transduction of adipose tissue that is superior to naturally occurring serotypes via direct injection to adipose depots. Furthermore, the administration route influences the tropism and efficacy of Rec2 vector: oral administration transduces interscapular brown fat, while intraperitoneal injection preferentially targets visceral fat. Multiple in vivo studies by our lab and others have demonstrated that Rec2 vector provides a powerful tool to genetically manipulate adipose tissue for basic research and potential gene therapies of genetic and acquired diseases. Here we provide detailed protocols for AAV production and delivery to adipose tissue by direct injection, oral administration, and intraperitoneal injection.
重组腺相关病毒(rAAV)载体是基因治疗中颇具吸引力的载体。然而,由于天然存在的AAV血清型转导效率低,对成年动物的脂肪组织进行基因操作具有挑战性。我们最近证明,一种新型工程化杂交血清型Rec2通过直接注射到脂肪库中,能够实现对脂肪组织的高效转导,其效果优于天然血清型。此外,给药途径会影响Rec2载体的靶向性和疗效:口服给药可转导肩胛间棕色脂肪,而腹腔注射则优先靶向内脏脂肪。我们实验室和其他机构的多项体内研究表明,Rec2载体为在基础研究以及对遗传和后天性疾病进行潜在基因治疗时对脂肪组织进行基因操作提供了一个强大的工具。在此,我们提供了通过直接注射、口服给药和腹腔注射生产AAV并将其递送至脂肪组织的详细方案。