Data showed an increase of the fluorescence intensity up to about 10 μg/mL. A saturation of the signal can be observed VEGFR inhibitor for nanoparticle concentrations higher than 10 μg/mL. To prove the internalization of the carriers in the cells, images at different focal depth were recorded. Figure 6 shows that going from upper cell LY2874455 solubility dmso surface to the focus inside the cells, an increase of red diatomite fluorescence can be observed thus indicating the uptake of DNPs* by H1355 cells. Figure 5 Confocal microscopy images and cell fluorescence intensity analysis. Confocal microscopy image of H1355 cells incubated with different concentrations of DNPs* (A); scale bar corresponds to 20 μm. Cell fluorescence
intensity vs nanoparticles concentration (B); the values reported were obtained from fluorescence analysis of diatomite-TRITC images in panel (A). Figure 6 Confocal microscopy image with different focal depth of H1355 cells incubated with FK506 ic50 10 μg/mL of DNPs*. Conclusions In this work, a procedure for preparing diatomite nanoparticles with an average size of 200 nm was described. DNP morphology and surface chemical modifications were investigated by DLS, SEM and TEM, and FTIR analyses, respectively. Confocal microscopy experiments revealed an efficient nanoparticle uptake into cytoplasm of human epidermoid carcinoma cells. This preliminary study demonstrates
that the diatomite nanoparticles could represent a promising tool for the delivery of anticancer molecules such as siRNA, miRNA, and drugs inside cancer cells. Since APTES functionalization of the nanoparticles showed the possibility to efficiently bind amino-reactive groups (TRITC), the development of chemical protocols
for loading anticancer molecules represents a further step in order to finalize the use of diatomite in medical applications. Moreover, it would be expected that compared to other nanocarriers, their Morin Hydrate selective targeted functionalization will improve the delivery of anti-tumoral molecules to specific cell population. Acknowledgements The authors thank the DEREF S.p.A. for kindly providing the diatomite earth sample. The authors also thank S. Arbucci of the IGB-CNR Integrated Microscopy Facility for the assistance with confocal microscopy acquisition and Dr. P. Dardano of the IMM-CNR for the SEM analysis. This work has been partially supported by Italian National Operative Program PON01_02782 and POR Campania FSE 2007-2013, Project CRÈME. References 1. Mai WX, Meng H: Mesoporous silica nanoparticles: a multifunctional nano therapeutic system. Integr Biol 2013, 5:19–28. 10.1039/c2ib20137bCrossRef 2. Zhang H, Shahbazi MA, Mäkilä EM, da Silva TH, Reis RL, Salonen JJ, Hirvonen JT, Santos HA: Diatom silica microparticles for sustained release and permeation enhancement following oral delivery of prednisone and mesalamine. Biomaterials 2013, 34:9210–9219. 10.1016/j.biomaterials.