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Mesenchymal stem cells (MSCs) are stem cells present in adult tissues. They can be cultured, have great growth capacity, and can differentiate into several cell types. The isolation of urine-derived mesenchymal stem cells (hUSCs) was recently described. hUSCs present additional benefits in the fact that they can be easily obtained noninvasively. Regarding gene delivery, nonviral vectors based on cationic niosomes have been used and are more stable and have lower immunogenicity than viral vectors. However, their transfection efficiency is low and in need of improvement.

Cationic niosomes have become important non-viral vehicles for transporting a good number of small drug molecules and macromolecules. Growing interest shown by these colloidal nanoparticles in therapy is determined by their structural similarities to liposomes. Cationic niosomes are usually obtained from the self-assembly of non-ionic surfactant molecules. This process can be governed not only by the nature of such surfactants but also by others factors like the presence of additives, formulation preparation and properties of the encapsulated hydrophobic or hydrophilic molecules. This review is aimed at providing recent information for using cationic niosomes for gene delivery purposes with particular emphasis on improving the transportation of antisense oligonucleotides (ASOs), small interference RNAs (siRNAs), aptamers and plasmids (pDNA).

Bone morphogenetic protein-7(BMP-7) plays a pivotal role in the transformation of mesenchymal stem cells (MSCs) into bone. However, its impact is hampered due to its short half-life. Therefore, gene therapy may be an interesting approach to deliver BMP-7 gene to D1-MSCs. In this manuscript we prepared and characterized niosomes based on cationic lipid 2,3-di(tetradecyloxy)propan-1-amine, combined with polysorbate 80 for gene delivery purposes. Niosomes were characterized and combined initially with pCMS-EGFP reporter plasmid, and later with pUNO1-hBMP-7 plasmid to evaluate osteogenesis differentiation. Additionally, specific blockers of most relevant endocytic pathways were used to evaluate the intracellular disposition of complexes. MSCs transfected with niosomes showed increased growth rate, enhanced alkaline phosphatase activity (ALP) and extracellular matrix deposition which suggested the formation of osteoblast-like cells. We concluded that hBMP-7-transfected MSCs could be considered not only as an effective delivery tool of hBMP-7, but also as proliferating and bone forming cells for bone regeneration.

We designed niosomes based on three lipids that differed only in the polar-head group to analyze their influence on the transfection efficiency. These lipids were characterized by small-angle X-ray scattering before being incorporated into the niosomes which were characterized in terms of pKa, size, zeta potential, morphology and physical stability. Nioplexes were obtained upon the addition of a plasmid. Different ratios (w/w) were selected to analyze the influence of this parameter on size, charge and the ability to condense, release and protect the DNA. In vitro transfection experiments were performed in HEK-293, ARPE-19 and MSC-D1 cells. Our results show that the chemical composition of the cationic head-group clearly affects the physicochemical parameters of the niosomes and especially the transfection efficiency. Only niosomes based on cationic lipids with a dimethyl amino head group (lipid 3) showed a transfection capacity when compared with their counterparts amino (lipid 1) and tripeptide head-groups (lipid 2). Regarding cell viability, we clearly observed that nioplexes based on the cationic lipid 3 had a more deleterious effect than their counterparts, especially in ARPE-19 cells at 20/1 and 30/1 ratios. Similar studies could be extended to other series of cationic lipids in order to progress in the research on safe and efficient non-viral vectors for gene delivery purposes.

The transfection of human NTera2/D1 teratocarcinoma-derived cell line (or NT2 cells) represents a promising strategy for the delivery of exogenous proteins or biological agents into the central nervous system (CNS). The development of suitable nonviral vectors with high transfection efficiencies requires a profound knowledge of the whole transfection process. In this work, we elaborated and characterized in terms of size and zeta potential three different nonviral vectors: lipoplexes (144 nm; -29.13 mV), nioplexes (142.5 nm; +35.4 mV), and polyplexes (294.8 nm; +15.1 mV). We compared the transfection efficiency, cellular uptake, and intracellular trafficking of the three vectors in NT2 cell line. Lipoplexes exhibited the highest percentages of EGFP positive cells. The values obtained with polyplexes were lower compared to lipoplexes but higher than the percentages obtained with nioplexes. Cellular uptake results had a clear correlation with respect to the corresponding transfection efficiencies. Regarding the endocytosis mechanism, lipoplexes enter in the cell, mainly, via clathrin-mediated endocytosis (CME) while polyplexes via caveolae-mediated endocytosis (CvME). Nioplexes were discarded for this experiment due to their low cellular uptake. By simulating an artificial endosome, we demonstrated that the vectors were able to release the DNA cargo once inside the late endosome. The data collected from this assay showed that at 6 h the genetic material carried by polyplexes was still located in the late endosome, while DNA carried by lipoplexes was already in the nucleus. This result indicates a faster intracellular traffic of the lipid-based vectors. Overall, our work gives new insights into the transfection process of NT2 cells by different nonviral vectors as a first step in the development of ex vivo gene therapy platform.