Pages J C, Andreoletti M, Bennoun M, Vons C, Elcheroth J, Lehn P, Houssin D, Chapman J, Briand P, Benarous R
ICGM, H pital Cochin, Paris, France.
Hum Gene Ther. 1995 Jan;6(1):21-30. doi: 10.1089/hum.1995.6.1-21.
The ex vivo approach to hepatic gene therapy involves several steps, which include the isolation and culture of hepatocytes, followed by their transduction with a retrovirus. Subsequently, autologous hepatocytes are transplanted. The number of hepatocytes that can be transduced by retroviruses bearing the therapeutic gene is one of the limiting steps that can impair the success of this strategy. We presently describe an experimental approach that leads to improved transduction efficiency in mouse and human hepatocytes in vitro. By using a recombinant retrovirus bearing the Escherichia coli beta-galactosidase gene, we show that addition of growth factors to the cells, namely human hepatocyte growth factor (HGF), allows marked increase in the transduction efficiency in mouse (up to 80%) and human (40%) hepatocytes. Familial hypercholesterolemia (FH) is due to mutation in the low-density lipoprotein (LDL) receptor gene and results in a deficiency in LDL receptors. Transduction of the human LDL receptor cDNA under the transcriptional control of the L-type pyruvate kinase promoter-activator into mouse hepatocytes led to an elevated tissue-specific expression of the human protein. These results suggest that the ex vivo approach remains a promising alternative for hepatic gene therapy.
肝脏基因治疗的离体方法包括几个步骤，其中包括肝细胞的分离和培养，随后用逆转录病毒进行转导。接着，进行自体肝细胞移植。携带治疗性基因的逆转录病毒能够转导的肝细胞数量是可能影响该策略成功的限制步骤之一。我们目前描述了一种实验方法，该方法可提高体外小鼠和人肝细胞的转导效率。通过使用携带大肠杆菌β-半乳糖苷酶基因的重组逆转录病毒，我们发现向细胞中添加生长因子，即人肝细胞生长因子（HGF），可使小鼠（高达80%）和人（40%）肝细胞的转导效率显著提高。家族性高胆固醇血症（FH）是由于低密度脂蛋白（LDL）受体基因突变导致LDL受体缺乏所致。在L型丙酮酸激酶启动子-激活剂的转录控制下，将人LDL受体cDNA转导入小鼠肝细胞，导致人蛋白的组织特异性表达升高。这些结果表明，离体方法仍然是肝脏基因治疗的一种有前景的替代方法。
