Is-7: Micro and Nanomaterials to Manipulate Stem Cells (Pages: 0-0)


Ferreira L *,

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The administration of some biomolecules to differentiate stem cells poses significant challenges because of their poor water solubility, short half-life, and potentially undesired side effects. One example is retinoic acid (RA), which interacts with members of the hormone receptor super-family, including the RA receptor (RAR), located in the cell nuclei. RA is rapidly metabolized by cells and has low solubility in aqueous solutions. In addition, the use of this biomolecule in an in vivo setting for the differentiation of resident stem cells remains elusive. We anticipate that nanoparticles (NPs) can be an excellent platform to ensure protection and intracellular transport of RA. Several RA NPs formulations have been described in the last years; however, no formulation was used for the intracellular delivery of RA in order to drive the differentiation of stem cells. Herein we describe a RA-containing NP formulation (RA+-NP) able to cross the cell membrane of subventricular zone (SVZ) stem/progenitor cells and release its payload intracellularly. This process drives the differentiation of stem cells into neuronal cells without affecting cell viability.We report a novel method to modulate the differentiation of SVZ cells into neurons involving the use of RA+-NPs. NPs with an average diameter of 200 nm were prepared, having a significant high RA payload (86 ± 28 μg of RA per mg of NPs) and a positive net charge (+15 mV). The NPs presented higher degradation rates at endossomal (pH = 5 - 6) than at physiological pH (pH = 7.4), likely due to the protonation of PEI amine groups (in excess relatively to the sulfate groups in DS) and the concomitant repulsion between positive charges. The RA+-NPs can be taken up rapidly by SVZ cells (first 5 hours) under the tested conditions and localize in the overall cell cytoplasm after 18 hours. RA+-NPs for concentrations below 100 μg/mL do not exert a cytotoxic effect as evaluated by TUNEL, propidium iodide uptake and active caspase 3 immunodetection. Incubation of SVZ cells with RA+-NPs at concentrations ranging between 0.1 and 1 μg/mL (corresponding to a RA payload of 1.2-12 ng/mL) induced an increase of NeuN+ cells (neurons). Importantly, RA+-NP-conditioned medium (CM) is unable to promote the differentiation of stem cells indicating that neurodifferentiation is only mediated by internalization of the RA+-NPs. In addition, RA+-NPs are pro-neurogenic for lower concentrations of RA as compared to RA dissolved in the differentiation medium, highlighting for the first time the importance of drug spatial positioning and concentration in terms of stem cell differentiation. Finally, we show that the internalization of RA+-NPs contribute for a high percentage of functional neuronal cells as shown by an increase of intracellular calcium following KCl but not histamine stimuli.