Guliyeva Ulker, Oner Filiz, Ozsoy Sule, Haziroğlu Rifki
Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Hacettepe University, Ankara, Turkey.
Eur J Pharm Biopharm. 2006 Jan;62(1):17-25. doi: 10.1016/j.ejpb.2005.08.006. Epub 2005 Oct 27.
The potential of chitosan as a polycationic gene carrier for oral administration has been explored since 1990s. Chitosan has been shown to effectively bind DNA in saline or acetic acid solution and protect DNA from nuclease degradation. In this study, pDNA (plasmid DNA) was encapsulated in chitosan microparticles. Chitosan-DNA microparticles were prepared using a complex coacervation process and stability of plasmid DNA was investigated in this complex. The chitosan-DNA microparticles could protect the encapsulated plasmid DNA from nuclease degradation. Release of pDNA from microparticles was studied in simulated gastric, simulated intestinal medium and acidic PBS (phosphate buffer saline) (pH 4.5) buffer at 37 degrees C, and released pDNA was assayed spectrophotometrically. In vitro release of pDNA from chitosan microparticles was dependent on pH, as the pH of the release medium increased release profile decreased. In in vivo-animal studies blue color was observed with X-gal (4-chloro-5-bromo-3-indolyl-beta-galactosidase) staining of histological stomach and small intestine sections after oral administration of pDNA-chitosan microparticles as an indicator of exogeneous gene expression.
自20世纪90年代以来,壳聚糖作为口服给药的聚阳离子基因载体的潜力就已得到探索。壳聚糖已被证明能在盐水或醋酸溶液中有效结合DNA,并保护DNA免受核酸酶降解。在本研究中,将pDNA(质粒DNA)包裹于壳聚糖微粒中。采用复合凝聚法制备壳聚糖-DNA微粒,并研究了该复合物中质粒DNA的稳定性。壳聚糖-DNA微粒能够保护包裹的质粒DNA免受核酸酶降解。在37℃下,于模拟胃液、模拟肠液培养基和酸性PBS(磷酸盐缓冲盐水)(pH 4.5)缓冲液中研究了pDNA从微粒中的释放情况,并用分光光度法测定释放出的pDNA。壳聚糖微粒中pDNA的体外释放取决于pH值,随着释放介质pH值升高,释放曲线下降。在体内动物研究中,口服pDNA-壳聚糖微粒后,对组织学上的胃和小肠切片进行X-gal(4-氯-5-溴-3-吲哚基-β-半乳糖苷酶)染色,观察到蓝色,作为外源基因表达的指标。