title: siRNA Inhibition of Endocytic Pathways to Characterize the Cellular Uptake Mechanisms of Folate-Functionalized Glycol Chitosan Nanogels.
abstract: Glycol chitosan nanogels have been widely used in gene, drug, and contrast agent delivery in an effort to improve disease diagnosis and treatment. Herein, we evaluate the internalization mechanisms and intracellular fate of previously described glycol chitosan nanogels decorated with folate to target the folate receptor. Uptake of the folate-decorated nanogel was impaired by free folate, suggesting competitive inhibition and shared internalization mechanisms via the folate receptor. Nanogel uptake was shown to occur mainly through flotillin-1 and Cdc42-dependent endocytosis. This was determined by inhibition of uptake reduction observed upon siRNA depletion of these two proteins and the pathways that they regulate. The data also suggest the involvement of the actin cytoskeleton in nanogel uptake via macropinocytosis. After 7 h of incubation with HeLa cells, approximately half of the nanogel population was localized in endolysosomal compartments, whereas the remaining 50% of the material was in undefined regions of the cytoplasm. Glycol chitosan nanogels may thus have potential as drug delivery vectors for targeting different intracellular compartments.
authors: †Institute for Biotechnology and Bioengineering, Centre of Biological Engineering, Campus de Gualtar, University of Minho, Braga 4710-05, Portugal.
authors: Paula
authors: P
authors: Pereira
authors: †Institute for Biotechnology and Bioengineering, Centre of Biological Engineering, Campus de Gualtar, University of Minho, Braga 4710-05, Portugal.
authors: Sílvia S
authors: SS
authors: Pedrosa
authors: ‡Cardiff School of Pharmacy and Pharmaceutical Sciences, Cardiff University, Redwood Building, Cardiff CF10 3NB, Wales.
authors: Jennifer M
authors: JM
authors: Wymant
authors: ‡Cardiff School of Pharmacy and Pharmaceutical Sciences, Cardiff University, Redwood Building, Cardiff CF10 3NB, Wales.
authors: Edward
authors: E
authors: Sayers
authors: §Institute of Molecular and Cell Biology, Rua Campo Alegre, Porto 4099-003, Portugal.
authors: Alexandra
authors: A
authors: Correia
authors: §Institute of Molecular and Cell Biology, Rua Campo Alegre, Porto 4099-003, Portugal.|∥Abel Salazar Biomedical Sciences Institute, University of Porto, Rua de Jorge Viterbo Ferreira No. 228, Porto 4050-313, Portugal.
authors: Manuel
authors: M
authors: Vilanova
authors: ‡Cardiff School of Pharmacy and Pharmaceutical Sciences, Cardiff University, Redwood Building, Cardiff CF10 3NB, Wales.
authors: Arwyn T
authors: AT
authors: Jones
authors: †Institute for Biotechnology and Bioengineering, Centre of Biological Engineering, Campus de Gualtar, University of Minho, Braga 4710-05, Portugal.
authors: Francisco M
authors: FM
authors: Gama
mesh_terms: D015153:Blotting, Western; D048271:Chitosan / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D004705:Endocytosis / Q000502:physiology; D005434:Flow Cytometry; D005492:Folic Acid / Q000737:chemistry*; D006367:HeLa Cells; D006801:Humans; D000080385:Nanogels; D011092:Polyethylene Glycols / Q000737:chemistry*; D011094:Polyethyleneimine / Q000737:chemistry*; D034741:RNA, Small Interfering / Q000235:genetics*
chemical_list: D000080385:Nanogels; D034741:RNA, Small Interfering; C118638:glycol-chitosan; C413692:polyethylene glycol polyethyleneimine nanogel; D011092:Polyethylene Glycols; D011094:Polyethyleneimine; D048271:Chitosan; D005492:Folic Acid
keywords: endocytic pathways; folate; glycol chitosan nanogel; intracellular localization; siRNA transfection