Faculty of Electrical Engineering, Laboratory of Biocybernetics, University of Ljubljana, Tržaška 25, 1000, Ljubljana, Slovenia.
Faculty of Medicine, Institute of Biophysics, University of Ljubljana, Vrazov trg 2, 1000, Ljubljana, Slovenia.
Biomed Eng Online. 2021 Aug 21;20(1):85. doi: 10.1186/s12938-021-00922-3.
Gene electrotransfer is an established method that enables transfer of DNA into cells with electric pulses. Several studies analyzed and optimized different parameters of gene electrotransfer, however, one of main obstacles toward efficient electrotransfection in vivo is relatively poor DNA mobility in tissues. Our aim was to analyze the effect of impaired mobility on gene electrotransfer efficiency experimentally and theoretically. We applied electric pulses with different durations on plated cells, cells grown on collagen layer and cells embedded in collagen gel (3D model) and analyzed gene electrotransfer efficiency. In order to analyze the effect of impaired mobility on gene electrotransfer efficiency, we applied electric pulses with different durations on plated cells, cells grown on collagen layer and cells embedded in collagen gel (3D model) and analyzed gene electrotransfer efficiency.
We obtained the highest transfection in plated cells, while transfection efficiency of embedded cells in 3D model was lowest, similarly as in in vivo. To further analyze DNA diffusion in 3D model, we applied DNA on top or injected it into 3D model and showed, that for the former gene electrotransfer efficiency was similarly as in in vivo. The experimental results are explained with theoretical analysis of DNA diffusion and electromobility.
We show, empirically and theoretically that DNA has impaired electromobility and especially diffusion in collagen environment, where the latter crucially limits electrotransfection. Our model enables optimization of gene electrotransfer in in vitro conditions.
基因电转移是一种将 DNA 导入细胞的成熟方法，通过电脉冲实现。已有多项研究分析和优化了基因电转移的不同参数，但在体内实现高效电转染的主要障碍之一是 DNA 在组织中的迁移率相对较差。我们旨在从实验和理论两方面分析 DNA 迁移率受损对基因电转移效率的影响。我们在铺板细胞、在胶原层上生长的细胞和嵌入胶原凝胶（3D 模型）的细胞上施加不同持续时间的电脉冲，并分析基因电转移效率。为了分析 DNA 迁移率受损对基因电转移效率的影响，我们在铺板细胞、在胶原层上生长的细胞和嵌入胶原凝胶（3D 模型）的细胞上施加不同持续时间的电脉冲，并分析基因电转移效率。
我们在铺板细胞中获得了最高的转染效率，而 3D 模型中嵌入细胞的转染效率最低，与体内情况相似。为了进一步分析 3D 模型中的 DNA 扩散，我们将 DNA 置于顶部或注入 3D 模型中，并表明对于前者，基因电转移效率与体内相似。实验结果通过 DNA 扩散和电泳迁移率的理论分析得到解释。
我们从实验和理论上表明，DNA 在胶原环境中的电泳迁移率，特别是扩散受到损害，后者是限制电转染的关键因素。我们的模型可以优化体外条件下的基因电转移。
