Barbara Adinolfi1, Mario Pellegrino2, Sara Tombelli3, Cosimo Trono4, Ambra Giannetti5, Claudio Domenici6, Greta Varchi7, Giovanna Sotgiu8, Marco Ballestri9, Francesco Baldini10. 1. Istituto di Fisica Applicata "Nello Carrara", Consiglio Nazionale delle Ricerche, Via Madonna del Piano 10, 50019 Sesto Fiorentino, FI, Italy. Electronic address: b.adinolfi@ifac.cnr.it. 2. Istituto Nazionale di Ottica, Consiglio Nazionale delle Ricerche, Via G. Moruzzi 1, 56124 Pisa, PI, Italy. Electronic address: mario.pellegrino@ino.it. 3. Istituto di Fisica Applicata "Nello Carrara", Consiglio Nazionale delle Ricerche, Via Madonna del Piano 10, 50019 Sesto Fiorentino, FI, Italy. Electronic address: s.tombelli@ifac.cnr.it. 4. Istituto di Fisica Applicata "Nello Carrara", Consiglio Nazionale delle Ricerche, Via Madonna del Piano 10, 50019 Sesto Fiorentino, FI, Italy. Electronic address: c.trono@ifac.cnr.it. 5. Istituto di Fisica Applicata "Nello Carrara", Consiglio Nazionale delle Ricerche, Via Madonna del Piano 10, 50019 Sesto Fiorentino, FI, Italy. Electronic address: a.giannetti@ifac.cnr.it. 6. Istituto di Fisiologia Clinica, Consiglio Nazionale delle Ricerche, Via G. Moruzzi 1, 56124 Pisa, PI, Italy. Electronic address: domenici@ifc.cnr.it. 7. Istituto per la Sintesi Organica e la Fotoreattività, Consiglio Nazionale delle Ricerche, Via P. Gobetti 101, 40129 Bologna, BO, Italy. Electronic address: greta.varchi@isof.cnr.it. 8. Istituto per la Sintesi Organica e la Fotoreattività, Consiglio Nazionale delle Ricerche, Via P. Gobetti 101, 40129 Bologna, BO, Italy. Electronic address: giovanna.sotgiu@isof.cnr.it. 9. Istituto per la Sintesi Organica e la Fotoreattività, Consiglio Nazionale delle Ricerche, Via P. Gobetti 101, 40129 Bologna, BO, Italy. Electronic address: marco.ballestri@isof.cnr.it. 10. Istituto di Fisica Applicata "Nello Carrara", Consiglio Nazionale delle Ricerche, Via Madonna del Piano 10, 50019 Sesto Fiorentino, FI, Italy. Electronic address: f.baldini@ifac.cnr.it.
Abstract
Polymethylmethacrylate core-shell fluorescent nanoparticles promote, in human lung A549 cancer cells, the internalization of a molecular beacon (MB) specific for survivin mRNA, an anti-apoptotic protein overexpressed in cancer cells. AIMS: To design an effective drug delivery system, the knowledge of the uptake mechanism and of the nanoparticles (NPs) and MB fate is required. MATERIALS AND METHODS AND KEY FINDINGS: Experiments with dextran as marker for endocytosis showed that in the presence of NPs the number of endocytic vesicles per cell doubled and their mean size significantly (p < 0.001) increased with respect to controls in absence of NPs, indicating an involvement of NPs in the endocytotic process. By using LysoTracker™ Deep Red, as marker of lysosomes, we found that nanoparticles co-localize with lysosomes. Moreover, a cellular release of nanoparticles detected in the culture medium, suggested a role of lysosomal exocytosis in nanoparticle elimination. The MB fluorescence in proximity of the labeled Endoplasmic Reticulum was indicative that the opening of the MB occurs in proximity of its target mRNA. SIGNIFICANCE: The results show the involvement of endocytotic pathway in the uptake of NPs, which are an appropriate delivery system capable of being eliminated by cells. Furthermore the data confirm that the MB can be considered an effective tool for the intracellular sensing.
Polymethylmethacrylate core-shell fluorescent nanoparticles promote, in humanlung A549 cancer cells, the internalization of a molecular beacon (MB) specific for survivin mRNA, an anti-apoptotic protein overexpressed in cancer cells. AIMS: To design an effective drug delivery system, the knowledge of the uptake mechanism and of the nanoparticles (NPs) and MB fate is required. MATERIALS AND METHODS AND KEY FINDINGS: Experiments with dextran as marker for endocytosis showed that in the presence of NPs the number of endocytic vesicles per cell doubled and their mean size significantly (p < 0.001) increased with respect to controls in absence of NPs, indicating an involvement of NPs in the endocytotic process. By using LysoTracker™ Deep Red, as marker of lysosomes, we found that nanoparticles co-localize with lysosomes. Moreover, a cellular release of nanoparticles detected in the culture medium, suggested a role of lysosomal exocytosis in nanoparticle elimination. The MB fluorescence in proximity of the labeled Endoplasmic Reticulum was indicative that the opening of the MB occurs in proximity of its target mRNA. SIGNIFICANCE: The results show the involvement of endocytotic pathway in the uptake of NPs, which are an appropriate delivery system capable of being eliminated by cells. Furthermore the data confirm that the MB can be considered an effective tool for the intracellular sensing.