UNASSIGNED: A triple-plasmid transfection protocol was used to produce rAAV subtype-9 vectors containing firefly luciferase genomes in HEK293 cells. Lewis rat liver grafts were flushed and stored in cold HTK solution. Three experimental groups received rAAV at different doses, administered via the portal vein as a bolus during SCS. A control group did not receive rAAV (N = 2). Recipients then underwent syngeneic liver transplantation. Bioluminescence imaging to quantify in vivo luciferase expression was performed on post-operative days 7, 14, 28, and 56.
UNASSIGNED: Control animals demonstrated no bioluminescent activity, while animals receiving rAAV-treated livers had increasing bioluminescence, peaking at four weeks but sustained to the eight-week endpoint. This result was confirmed by experimental endpoint tissue luciferase activity assay.
UNASSIGNED: rAAV mediates gene transduction in liver grafts when administered during SCS and has potential for gene therapy applications in solid organ transplantation.
使用三重质粒转染方案在HEK293细胞中产生含有萤火虫荧光素酶基因组的rAAV亚型-9载体。冲洗Lewis大鼠肝脏移植物并储存在冷的HTK溶液中。三个实验组接受不同剂量的rAAV,在SCS期间通过门静脉作为推注给药。对照组不接受rAAV(N=2)。受者随后接受了同基因肝移植。在术后第7、14、28和56天进行生物发光成像以定量体内荧光素酶表达。
■对照动物没有生物发光活性,虽然接受rAAV处理的肝脏的动物的生物发光增加,在四周达到峰值,但持续到八周终点。该结果通过实验终点组织荧光素酶活性测定得到证实。
■rAAV在SCS期间施用时介导肝移植物中的基因转导,并且具有在实体器官移植中基因治疗应用的潜力。