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The nanoparticles transported antisense oligonucleotides (ASOs), either encapsulated (CA nanocomplexes) or electrostatically bound externally (NP and ES nanocomplexes). After synthesis, the PEI\/ASO nanoconjugates were functionalized with a muscle-specific RNA aptamer. Using this combinatorial formulation methodology, we obtained nanocomplexes that were further used as nanocarriers for the delivery of RNA therapeutics (ASO), specifically into muscle cells. In particular, we performed a detailed confocal microscopy-based comparative study to analyze the overall transfection efficiency, the cell-to-cell homogeneity, and the mean fluorescence intensity per cell of micron-sized domains enriched with the nanocomplexes. Furthermore, using high-magnification electron microscopy, we were able to describe, in detail, the ultrastructural basis of the cellular uptake and intracellular trafficking of nanocomplexes by the clathrin-independent endocytic pathway. Our results are a clear demonstration that coaxial electrospraying is a promising methodology for the synthesis of therapeutic nanoparticle-based carriers. Some of the principal features that the nanoparticles synthesized by coaxial electrospraying exhibit are efficient RNA-based drug encapsulation, increased nanoparticle surface availability for aptamer functionalization, a high transfection efficiency, and hyperactivation of the endocytosis and early\/late endosome route as the main intracellular uptake mechanism.<\/jats:p>","DOI":"10.3390\/biom12081012","type":"journal-article","created":{"date-parts":[[2022,7,22]],"date-time":"2022-07-22T12:53:45Z","timestamp":1658494425000},"page":"1012","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":14,"title":["Coaxial Synthesis of PEI-Based Nanocarriers of Encapsulated RNA-Therapeutics to Specifically Target Muscle Cells"],"prefix":"10.3390","volume":"12","author":[{"given":"Raquel","family":"de la Hoz","sequence":"first","affiliation":[{"name":"Department of Chemical and Biomolecular Engineering, University of Cantabria, 39011 Santander, Spain"},{"name":"Health Research Institute Valdecilla (IDIVAL), 39011 Santander, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2636-6305","authenticated-orcid":false,"given":"Nazely","family":"Diban","sequence":"additional","affiliation":[{"name":"Department of Chemical and Biomolecular Engineering, University of Cantabria, 39011 Santander, Spain"},{"name":"Health Research Institute Valdecilla (IDIVAL), 39011 Santander, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mar\u00eda T.","family":"Berciano","sequence":"additional","affiliation":[{"name":"Health Research Institute Valdecilla (IDIVAL), 39011 Santander, Spain"},{"name":"Department of Molecular Biology, University of Cantabria, 39011 Santander, Spain"},{"name":"Centro de Investigaci\u00f3n Biom\u00e9dica en Red Sobre Enfermedades Neurodegenerativas (CIBERNED), 28029 Madrid, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0437-001X","authenticated-orcid":false,"given":"Carlos","family":"San Emeterio","sequence":"additional","affiliation":[{"name":"Research Group on Food, Nutritional Biochemistry and Health, Universidad Europea del Atl\u00e1ntico, 39011 Santander, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ane","family":"Urtiaga","sequence":"additional","affiliation":[{"name":"Department of Chemical and Biomolecular Engineering, University of Cantabria, 39011 Santander, Spain"},{"name":"Health Research Institute Valdecilla (IDIVAL), 39011 Santander, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3402-1152","authenticated-orcid":false,"given":"Miguel","family":"Lafarga","sequence":"additional","affiliation":[{"name":"Health Research Institute Valdecilla (IDIVAL), 39011 Santander, Spain"},{"name":"Centro de Investigaci\u00f3n Biom\u00e9dica en Red Sobre Enfermedades Neurodegenerativas (CIBERNED), 28029 Madrid, Spain"},{"name":"Department of Anatomy and Cellular Biology, University of Cantabria, 39011 Santander, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8369-3118","authenticated-orcid":false,"given":"Jos\u00e9 C.","family":"Rodr\u00edguez-Rey","sequence":"additional","affiliation":[{"name":"Health Research Institute Valdecilla (IDIVAL), 39011 Santander, Spain"},{"name":"Department of Molecular Biology, University of Cantabria, 39011 Santander, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0802-7975","authenticated-orcid":false,"given":"Olga","family":"Tapia","sequence":"additional","affiliation":[{"name":"Research Group on Food, Nutritional Biochemistry and Health, Universidad Europea del Atl\u00e1ntico, 39011 Santander, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2022,7,22]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"60","DOI":"10.1016\/j.chembiol.2011.12.008","article-title":"RNA-Based Therapeutics: Current Progress and Future Prospects","volume":"19","author":"Burnett","year":"2012","journal-title":"Chem. 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