30 Thus, microRNA nanomedicines may provide new therapeutic options for the treatment of many disease states, including cystic fibrosis. Herein we explored the use of cationic Imatinib Mesylate molecular weight nanoparticles in the delivery of miRNA into CF cell lines to determine if they could modulate miR-126 levels in CFBE41o- cells and impact on the associated expression of the miR-126 validated target, TOM1. There are limited data available on the physicochemical characteristics of miRNA nanoparticles. The sizes of PEI-miRNA complexes appear to be buffer-dependent, as size was significantly reduced when the complexes were prepared in a 5% (w/v) glucose solution instead of PBS (Figure 1). Others have found similar reductions in size of PEI-DNA complexes by using 5% glucose instead of salt-based solutions.

31 As for other nucleic acids, including plasmid DNA and siRNA, PEI appears to be an extremely effective complexation agent for miRNA mimics, complexing the premiR completely at low N/P ratios to create small, positively charged nanoparticles. While the chitosan-miRNA nanoparticles prepared herein were slightly larger than the chitosan-siRNA polyplexes reported in the literature,32 use of the TPP crosslinker significantly reduced the size of the miRNA nanoparticles and altered the intracellular distribution of the miRNA once internalized by the CFBE cells (Figure 2B). High content analysis (Figure 2A) confirms the effective uptake of miRNA nanomedicines into CFBE41o- cells. This technique has been successfully applied to screen delivery of plasmid DNA33 and siRNA,34 and herein we apply it for the first time to the screening of intracellular delivery for miRNA nanoparticles.

The advantages of the technique include the ability to screen a range of delivery systems in parallel and generate both qualitative images and quantitative uptake data. The high content analysis data indicated that delivery of premiRNA-PEI nanomedicines to CFBE cells was twice as effective as miRNA transfection using RiboJuice, a commercial transfection agent often used for miRNA transfection in molecular biology applications. The high content analysis uptake data also indicated that PEI was a more efficient miRNA carrier than chitosan. The findings of the high content analysis uptake study are borne out by the miR-126 assay (Figure 4) in which PEI-miRNA nanomedicines were found to be significantly more efficient than chitosan-miRNA nanomedicines at increasing miR-126 levels.

This may be explained by both the higher binding efficiency of premiRs to PEI than chitosan (data not shown) and its highly cationic nature that increases its interaction with the cell membrane, thereby enhancing transfection. This builds on and supports previous work harnessing PEI and chitosan for siRNA delivery that found PEI was significantly better at both Batimastat complexing siRNA and transfecting cells with short RNA sequences.