de Poorter Jolanda J, Lipinski Kai S, Nelissen Rob G H H, Huizinga Tom W J, Hoeben Rob C
Leiden University Medical Center, Leiden, The Netherlands.
J Gene Med. 2007 Aug;9(8):639-48. doi: 10.1002/jgm.1057.
Predictable and adequate transgene expression is essential for clinical gene therapy. Several studies have focused on optimization of transgene expression. In this study the effect of sodium butyrate (NaB) and a ubiquitous chromatin opening element (UCOE) on short-term gene expression after adenovirus-mediated gene transfer in fibroblastic interface cells from periprosthetic tissue in loosened orthopedic implants is investigated.
Cultures of diploid human interface cells from four patients were infected with an adenovirus type-5 vector that carries the luciferase gene driven by the cytomegalovirus (CMV) promoter as a reporter. In addition, viruses with a UCOE were evaluated. Twenty-four hours after infection NaB was added in concentrations of 0 to 9 mM. Luciferase activity was tested after a further 24 h.
NaB in a concentration of 6 mM caused a 7- to 16-fold increase in reporter gene expression compared to control condition. There was no difference in reporter gene expression when cells were infected with Ad.1.5UCOE-CMV.Luc compared to Ad.CMV.Luc. A combination of NaB and a UCOE had no advantage over NaB alone.
Addition of NaB results in a marked increase in transgene expression in cultured cells. This would allow the enhancement of the expression of the transgene, without requiring a higher vector dose. Butyrate administration could not be substituted by inclusion of UCOEs in the vector. It remains to be established whether the effective concentrations of butyrate can be obtained in vivo.
可预测且充足的转基因表达对于临床基因治疗至关重要。多项研究聚焦于转基因表达的优化。本研究调查了丁酸钠（NaB）和一种普遍存在的染色质开放元件（UCOE）对腺病毒介导的基因转移后，来自松动骨科植入物周围假体组织的成纤维细胞界面细胞中短期基因表达的影响。
来自四名患者的二倍体人界面细胞培养物用携带由巨细胞病毒（CMV）启动子驱动的荧光素酶基因作为报告基因的5型腺病毒载体进行感染。此外，对带有UCOE的病毒进行了评估。感染后24小时，加入浓度为0至9 mM的NaB。再经过24小时后检测荧光素酶活性。
与对照条件相比，6 mM浓度的NaB使报告基因表达增加了7至16倍。与Ad.CMV.Luc相比，用Ad.1.5UCOE-CMV.Luc感染细胞时报告基因表达没有差异。NaB和UCOE的组合并不比单独使用NaB更具优势。
添加NaB可使培养细胞中的转基因表达显著增加。这将允许在不增加载体剂量的情况下增强转基因的表达。丁酸盐的施用不能通过在载体中包含UCOE来替代。丁酸盐的有效浓度在体内是否能够达到仍有待确定。
