Department of Orthopaedics, University of Maryland School of Medicine , Baltimore, MD, USA.
Department of Microbiology and Immunology, University of Maryland School of Medicine , Baltimore, MD, USA.
Bone Res. 2016 Aug 9;4:16021. doi: 10.1038/boneres.2016.21. eCollection 2016.
We report here a method for the use of poly-l-lysine (PLL) to markedly improve the adenoviral transduction efficiency of primary murine osteoblasts and bone marrow stromal cells (BMSCs) in culture and in situ, which are typically difficult to transduce. We show by fluorescence microscopy and flow cytometry that the addition of PLL to the viral-containing medium significantly increases the number of green fluorescence protein (GFP)-positive osteoblasts and BMSCs transduced with an enhanced GFP-expressing adenovirus. We also demonstrate that PLL can greatly enhance the adenoviral transduction of osteoblasts and osteocytes in situ in ex vivo tibia and calvaria, as well as in long bone fragments. In addition, we validate that PLL can improve routine adenoviral transduction studies by permitting the use of low multiplicities of infection to obtain the desired biologic effect. Ultimately, the use of PLL to facilitate adenoviral gene transfer in osteogenic cells can provide a cost-effective means of performing efficient gene transfer studies in the context of bone research.
我们在此报告一种使用多聚赖氨酸(PLL)的方法,可显著提高原代鼠成骨细胞和骨髓基质细胞(BMSCs)在培养和原位中的腺病毒转导效率,这两种细胞通常较难转导。我们通过荧光显微镜和流式细胞术观察到,在含病毒的培养基中添加 PLL 可显著增加转导增强型 GFP 表达腺病毒的绿色荧光蛋白(GFP)阳性成骨细胞和 BMSCs 的数量。我们还证明,PLL 可极大地增强体外胫骨和颅骨以及长骨片段中骨细胞和成骨细胞内的腺病毒转导。此外,我们通过允许使用低感染复数来获得所需的生物学效应,验证了 PLL 可以改善常规腺病毒转导研究。最终,使用 PLL 促进成骨细胞中的腺病毒基因转移,可以为骨研究背景下进行高效基因转移研究提供一种具有成本效益的方法。