Yu Qing, Que Loretta G, Rockey Don C
Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710, USA.
Am J Physiol Gastrointest Liver Physiol. 2002 Mar;282(3):G565-72. doi: 10.1152/ajpgi.00512.2000.
Adenovirus-mediated gene transfer has become an important tool with which to introduce genetic material into cells. Available data emphasize efficient adenoviral transduction of parenchymal liver cells (i.e., hepatocytes) in both in vitro and in vivo model systems, typically in normal cells. The aim of this study was to evaluate gene transfer to nonparenchymal (and parenchymal) cells of the normal and injured rat liver. Hepatocytes, stellate cells, and endothelial cells were isolated by standard methods. Liver injury was induced by bile duct ligation or carbon tetrachloride administration. Cells were transduced in vitro with an adenovirus encoding beta-galactosidase (Ad.beta-gal) over a range of viral titers, and transduced cells were identified by detection of X-gal. In vivo transduction efficiency was studied in normal and injured livers using cell isolation techniques. Nonparenchymal cells were transduced with greater frequency than hepatocytes at all adenoviral titers tested, both in vitro and in vivo. After liver injury, adenoviral transduction was reduced for all liver cell types compared with that for cells from normal livers (at all virus titers). Notably, transduction efficiency remained greater in nonparenchymal cells than in hepatocytes after liver injury. This work implies that, to achieve comparable gene expression in the injured liver, higher adenoviral titers may be required, an important consideration as gene therapy in disease states is considered.
腺病毒介导的基因转移已成为将遗传物质导入细胞的一种重要工具。现有数据强调在体外和体内模型系统中，通常在正常细胞中，腺病毒能有效地转导肝实质细胞（即肝细胞）。本研究的目的是评估基因向正常和受损大鼠肝脏的非实质（和实质）细胞的转移情况。通过标准方法分离肝细胞、星状细胞和内皮细胞。通过胆管结扎或给予四氯化碳诱导肝损伤。用编码β-半乳糖苷酶的腺病毒（Ad.β-gal）在一系列病毒滴度下体外转导细胞，并通过检测X-gal鉴定转导的细胞。使用细胞分离技术研究正常和受损肝脏中的体内转导效率。在体外和体内测试的所有腺病毒滴度下，非实质细胞的转导频率均高于肝细胞。肝损伤后，与正常肝脏的细胞相比（在所有病毒滴度下），所有肝细胞类型的腺病毒转导均降低。值得注意的是，肝损伤后非实质细胞的转导效率仍高于肝细胞。这项工作表明，为了在受损肝脏中实现相当的基因表达，可能需要更高的腺病毒滴度，这是在考虑疾病状态下的基因治疗时的一个重要考量因素。
