Control experiments were performed in parallel: BLOCK-iT Alexa Fluor Red Fluorescent Oligo was incubated alone with MCF-7 to assess unspecific fluorescence, and it was also delivered with commercially available transfection medium recommended for dsRNA transfection to assess efficient delivery of the fluorescent oligo. As it can be observed in Figure 7, both lecithin dispersions at pH 5.0 and pH 7.0 are able to efficiently deliver oligos in MCF-7 cells. Fluorescent Inhibitors,research,lifescience,medical siRNA mainly
located in the cytoplasm of the cells near the nucleus (Figures 7(c) and 7(d)). In contrast, fluorescently labeled naked siRNA was not detected by fluorescence microscopy (Figure 7(b)) neither within Inhibitors,research,lifescience,medical cells nor in the extracellular medium, suggesting that siRNA is degraded or removed by washing the cells when the incubation period is finished. Figure 7 Fluo-siRNA uptake by MCF-7 cells transfected with lecithin dispersions in pH 5.0 and pH 7.0 buffers. Control dsRNA:Lipofectamine (a), dsRNA alone (b), dsRNA:lecithin 25mM pH 5.0 (c), and dsRNA:lecithin 25mM pH 7.0 (d) at N/P 8000 were … 4. Conclusions
In the present work, a siRNA lecithin-based delivery system capable to improve the disadvantages that nonviral carriers normally present, like Inhibitors,research,lifescience,medical poor cellular uptake or high cytotoxicity, was readily obtained. It was not necessary to add other components like cationic lipids or cationic surfactants, of recognized toxicity, so as to improve siRNA loading capacity. In this case, the efficiency in loading was reached by means of the optimization of the critical parameters in the elaboration, such as pH and ionic strength. It was proposed that in the Inhibitors,research,lifescience,medical Inhibitors,research,lifescience,medical case of nanoparticles obtained at lower pH, an important electrostatic interaction between the oligonucleotide and the positively
charged head of the amphiphile is responsible for the formation of isolated spherical particles, while at higher pH, the interactions between charged groups of lecithin and siRNA are less relevant. When assessed in parallel with the commercial transfection reagent Lipofectamine, Sitaxentan lecithin dispersions at pH 5.0 and pH 7.0 were both able to efficiently deliver oligos in MCF-7 cells, in PLX4032 manufacturer contrast to naked siRNA. Moreover, fluorescent siRNA mainly located near its target, surrounding the nucleus of the cells. Neither other components like lipids for cell transfection nor molecular modifications were necessary. If the absence of toxicity and the significant cellular uptake exhibited are considered along with the ease of preparation, critical issues for the rest of nanocarriers that have been proposed for siRNA delivery, the present oligo delivery system represents a promising one for further investigation.