Merle Uta, Encke Jens, Tuma Sabine, Volkmann Martin, Naldini Luigi, Stremmel Wolfgang
Department of Gastroenterology, University Hospital Heidelberg, Germany.
Scand J Gastroenterol. 2006 Aug;41(8):974-82. doi: 10.1080/00365520600554790.
Wilson disease is a copper storage disorder caused by mutations in the ATP7B gene leading to liver cirrhosis. It has previously been shown that lentiviral vectors can govern an efficient delivery and stable expression of a transgene. The aim of this pilot study was to prove the principle of a lentiviral gene transfer in the Long-Evans cinnamon (LEC) rat, an animal model of Wilson disease.
LEC rats were treated either by systemic application of lentiviral vectors or by intrasplenic transplantation of LEC-rat hepatocytes lentivirally transduced with ATP7B. The ATP7B gene expression was analyzed by RT-PCR and immunofluorescence analysis. The therapeutic effect was assessed by analysis of liver histology, serum ceruloplasmin oxidase activity, and liver copper content.
Hepatic expression of the transgene was detected at different time-points post-treatment and lasted for up to 24 weeks (end of experiment). Liver copper levels were lowered in all treatment groups compared to untreated LEC rats. Twenty-four weeks after treatment, the area of the examined liver-tissue sections occupied by fibrosis was 48.3-57.9% in untreated LEC rats and 10.7-19.8% in rats treated with cell therapy. In systemically treated rats, only small fibrous septa could be observed.
These data prove for the first time that lentiviral ATP7B gene transfer is feasible in Wilson disease. In our pilot study the systemic approach was more promising in ameliorating disease progression than the transplantation of lentivirally transduced hepatocytes.
威尔逊病是一种由ATP7B基因突变导致的铜储存障碍性疾病，可引发肝硬化。此前已有研究表明，慢病毒载体能够实现转基因的高效递送和稳定表达。本初步研究的目的是在威尔逊病动物模型长-伊文斯肉桂色大鼠（LEC大鼠）中验证慢病毒基因转移的原理。
对LEC大鼠进行全身应用慢病毒载体治疗，或进行脾内移植经慢病毒转病毒转导ATP7B基因的LEC大鼠肝细胞治疗。通过逆转录聚合酶链反应（RT-PCR）和免疫荧光分析对ATP7B基因表达进行分析。通过肝组织学分析、血清铜蓝蛋白氧化酶活性分析和肝脏铜含量分析评估治疗效果。
在治疗后的不同时间点均检测到转基因的肝脏表达，且持续长达24周（实验结束时）。与未治疗的LEC大鼠相比，所有治疗组的肝脏铜水平均降低。治疗24周后，未治疗的LEC大鼠肝脏组织切片中纤维化所占面积为48.3% - 57.9%，而接受细胞治疗的大鼠中该面积为10.7% - 19.8%。在全身治疗的大鼠中，仅观察到小的纤维间隔。
这些数据首次证明慢病毒ATP7B基因转移在威尔逊病中是可行的。在我们的初步研究中，全身治疗方法在改善疾病进展方面比移植经慢病毒转导的肝细胞更具前景。
