Long-term reduction of serum bilirubin levels in Gunn rats by retroviral gene transfer in vivo.
Long-term reduction of serum bilirubin levels in Gunn rats by retroviral gene transfer in vivo.
复制标题
通过体内逆转录病毒基因转移长期降低 Gunn 大鼠血清胆红素水平。
DOI：
10.1002/lt.500040111
复制
发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Chowdhury,JR
中科院分区：
文献类型：
--
作者：
Tada,K;Chowdhury,NR;Neufeld,D;Bosma,PJ;Heard,M;Prasad,VR;Chowdhury,JR
由胆红素-尿苷二磷酸-葡萄糖醛酸糖醛酸转移酶（胆红素- ugt）介导的与葡萄糖醛酸的结合对于胆红素的有效胆汁排泄是必不可少的。这种酶活性的遗传缺失导致人类可能致命的Crigler-Najjar综合征I型和Gunn大鼠终生高胆红素血症。为了开发一种治疗胆红素- ugt缺乏症的基因疗法，我们构建了一种高滴度的复制缺陷性两性重组逆转录病毒（MFG-S hB-UGT 1），该病毒能够转移编码胆红素- ugt - 1的基因，这是人肝脏中胆红素- ugt的主要亚型。为了刺激肝细胞增殖，Gunn大鼠进行66%肝切除术。24h后夹持肝静脉上下门静脉、肝动脉及下腔静脉，对门静脉及离体段腔静脉插管。将MFG-S hb - ugt1制剂以5 ml/min的速度经门静脉灌注肝脏10分钟，恢复肝脏循环。对照大鼠肝脏灌注表达大肠杆菌β-半乳糖苷酶的重组逆转录病毒。在MFG-S hb - ugt1灌注的大鼠中，而不是在对照组中，通过免疫转印迹法、肝匀浆胆红素- ugt测定和胆红素二脲和单脲的胆红素排泄显示了人胆红素- ugt1的表达。平均血清胆红素水平在3周内下降了20%至25%，并在整个研究期间（18个月）保持在该水平。这是首个在Crigler-Najjar综合征动物模型中通过逆转录病毒定向基因治疗长期改善遗传性黄疸的报道。
Conjugation with glucuronic acid, mediated by bilirubin-uridinediphosphoglucuronate glucuronosyltransferase (bilirubin-UGT), is essential for efficient biliary excretion of bilirubin. Inherited absence of this enzyme activity results in the potentially lethal Crigler-Najjar syndrome type I in humans and lifelong hyperbilirubinemia in Gunn rats. To develop a gene therapy for bilirubin-UGT deficiency, we constructed a high-titer replication-deficient amphotropic recombinant retrovirus (MFG-S hB-UGT 1) capable of transferring the gene encoding bilirubin-UGT 1 the principal bilirubin-UGT isoform in human liver. To stimulate hepatocyte proliferation, Gunn rats were subjected to 66% hepatectomy. After 24 hours, the portal vein, the hepatic artery, and the inferior vena cava above and below the hepatic vein were clamped, and the portal vein and the isolated segment of the vena cava were cannulated. The liver was perfused with the MFG-S hB-UGT 1 preparation through the portal vein at 5 ml/min for 10 minutes, then circulation was restored. Control rat livers were perfused with a recombinant retrovirus expressingEscherichia coliβ-galactosidase. In MFG-S hB-UGT 1-perfused rats, but not in controls, expression of human bilirubin-UGT 1 was shown by immunotransblotting, bilirubin-UGT assay of liver homogenates, and biliary excretion of bilirubin diglucuronide and monoglucuronide. Mean serum bilirubin levels decreased by 20% to 25% in 3 weeks and remained at that level throughout the study period (18 months). This is the first report of long-term amelioration of inherited jaundice by retrovirus-directed gene therapy in an animal model for Crigler-Najjar syndrome.