CQM - Centro de Química da Madeira, MMRG, Universidade da Madeira, Campus Universitário da Penteada, 9000-390 Funchal, Portugal.
Int J Pharm. 2012 Sep 15;434(1-2):199-208. doi: 10.1016/j.ijpharm.2012.05.066. Epub 2012 Jun 1.
The present study aimed at developing a new approach in gene delivery of calcium phosphate nanoparticles through simulated body fluid (CaP-SBF). The physicochemical and biological characteristics of the CaP-SBF nanoparticles were compared with those made in pure water (CaP-water) via a similar procedure. The CaP-SBF and CaP-water solutions were then adjusted to two different pH values of 7.4 and 8.0, mixed with plasmid DNA (pDNA), and added in varying amounts to human embryonic kidney (HEK 293T) cells. The transfection efficiency and cell viability were studied in vitro by reporter gene (luciferase and Enhanced Green Fluorescent Protein) expression and the resazurin reduction assay, respectively, 24 and 48 h after the incubation with the nanoparticles. Our results indicated considerably high in vitro transfection efficiency for CaP-SBF/DNA complexes at physiological pH (7.4) with high amounts of CaP. Additionally, the SBF solution exhibited the ability to reduce the rapid growth of CaP particles over time, leading to higher transfection efficiency of CaP-SBF/DNA complexes than those made in water (CaP-water/DNA).
本研究旨在通过模拟体液（CaP-SBF）开发一种新的磷酸钙纳米颗粒基因传递方法。通过类似的程序，将 CaP-SBF 纳米颗粒的物理化学和生物学特性与在纯水中（CaP-water）制成的纳米颗粒进行比较。然后将 CaP-SBF 和 CaP-water 溶液调节至两种不同的 pH 值 7.4 和 8.0，与质粒 DNA（pDNA）混合，并以不同的量添加到人胚肾（HEK 293T）细胞中。转染效率和细胞活力通过报告基因（荧光素酶和增强型绿色荧光蛋白）表达和 Resazurin 还原测定在体外进行研究，分别在与纳米颗粒孵育 24 和 48 小时后进行。我们的结果表明，在生理 pH（7.4）下，具有高 CaP 含量的 CaP-SBF/DNA 复合物具有相当高的体外转染效率。此外，SBF 溶液具有降低 CaP 颗粒随时间快速生长的能力，导致 CaP-SBF/DNA 复合物的转染效率高于在水中（CaP-water/DNA）制成的复合物。
