In vitro biocompatibility study of sub-5 nm silica-coated magnetic iron oxide fluorescent nanoparticles for potential biomedical application

Magnetic iron oxide nanoparticles (IONPs), for their intriguing properties, have attracted a great interest as they can be employed in many different biomedical applications. In this multidisciplinary study, we synthetized and characterized ultrafine 3 nm superparamagnetic water-dispersible nanoparticles. By a facile and inexpensive one-pot approach, nanoparticles were coated with a shell of silica and contemporarily functionalized with fluorescein isothiocyanate (FITC) dye. The obtained sub-5 nm silica-coated magnetic iron oxide fluorescent (sub-5 SIO-Fl) nanoparticles were assayed for cellular uptake, biocompatibility and cytotoxicity in a human colon cancer cellular model. By confocal microscopy analysis we demonstrated that nanoparticles as-synthesized are internalized and do not interfere with the CaCo-2 cell cytoskeletal organization nor with their cellular adhesion. We assessed that they do not exhibit cytotoxicity, providing evidence that they do not affect shape, proliferation, cellular viability, cell cycle distribution and progression. We further demonstrated at molecular level that these nanoparticles do not interfere with the expression of key differentiation markers and do not affect pro-inflammatory cytokines response in Caco-2 cells. Overall, these results showed the in vitro biocompatibility of the sub-5 SIO-Fl nanoparticles promising their safe employ for diagnostic and therapeutic biomedical applications.

[1]  A. Tsourkas,et al.  Size, charge and concentration dependent uptake of iron oxide particles by non-phagocytic cells. , 2008, Biomaterials.

[2]  Bengt Fadeel,et al.  Efficient internalization of silica-coated iron oxide nanoparticles of different sizes by primary human macrophages and dendritic cells. , 2011, Toxicology and applied pharmacology.

[3]  Thomas J. Raub,et al.  Characterization of the human colon carcinoma cell line (Caco-2) as a model system for intestinal epithelial permeability. , 1989, Gastroenterology.

[4]  C. Slaughter,et al.  Nucleotide and amino acid sequences of human intestinal alkaline phosphatase: close homology to placental alkaline phosphatase. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[5]  Ji-won Yang,et al.  Optical Properties of Fluorescein‐labeled Organoclay , 2010, Photochemistry and photobiology.

[6]  S. Nie,et al.  Targeted magnetic iron oxide nanoparticles for tumor imaging and therapy , 2008 .

[7]  Y. Berger,et al.  The human intestinal epithelial cell line Caco-2; pharmacological and pharmacokinetic applications , 1995, Cell Biology and Toxicology.

[8]  Thomas J Webster,et al.  Nanomedicine: what’s in a definition? , 2006, International journal of nanomedicine.

[9]  Francesco Conversano,et al.  Magnetic/Silica Nanocomposites as Dual‐Mode Contrast Agents for Combined Magnetic Resonance Imaging and Ultrasonography , 2011 .

[10]  Valérie Fessard,et al.  Toxicity, genotoxicity and proinflammatory effects of amorphous nanosilica in the human intestinal Caco-2 cell line. , 2015, Toxicology in vitro : an international journal published in association with BIBRA.

[11]  L Wang,et al.  Raman and FTIR spectroscopies of fluorescein in solutions. , 2001, Spectrochimica acta. Part A, Molecular and biomolecular spectroscopy.

[12]  Egon Matijević,et al.  Uniform inorganic colloid dispersions. Achievements and challenges , 1994 .

[13]  C. Bräuchle,et al.  Uptake kinetics and nanotoxicity of silica nanoparticles are cell type dependent. , 2013, Small.

[14]  M. Papi,et al.  Dynamic light scattering for the characterization and counting of extracellular vesicles: a powerful noninvasive tool , 2014, Journal of Nanoparticle Research.

[15]  Warren C W Chan,et al.  Nanoparticle-mediated cellular response is size-dependent. , 2008, Nature nanotechnology.

[16]  J. Santamaría,et al.  Magnetic nanoparticles for drug delivery , 2007 .

[17]  Kirsten Gerloff,et al.  Influence of simulated gastrointestinal conditions on particle-induced cytotoxicity and interleukin-8 regulation in differentiated and undifferentiated Caco-2 cells , 2013, Nanotoxicology.

[18]  R. Hong,et al.  Preparation and characterization of silica-coated Fe3O4 nanoparticles used as precursor of ferrofluids , 2009 .

[19]  Tronc,et al.  Size Tailoring of Magnetite Particles Formed by Aqueous Precipitation: An Example of Thermodynamic Stability of Nanometric Oxide Particles. , 1998, Journal of colloid and interface science.

[20]  J. Millán,et al.  Accelerated Fat Absorption in Intestinal Alkaline Phosphatase Knockout Mice , 2003, Molecular and Cellular Biology.

[21]  Vibha Rani,et al.  Nanotechnology: Emerging Tool for Diagnostics and Therapeutics , 2011, Applied biochemistry and biotechnology.

[22]  Thomas D. Schmittgen,et al.  Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. , 2001, Methods.

[23]  Cui Tang,et al.  Effects of particle size and surface charge on cellular uptake and biodistribution of polymeric nanoparticles. , 2010, Biomaterials.

[24]  D. Fischer,et al.  Surface-modified biodegradable albumin nano- and microspheres. II: effect of surface charges on in vitro phagocytosis and biodistribution in rats. , 1998, European journal of pharmaceutics and biopharmaceutics : official journal of Arbeitsgemeinschaft fur Pharmazeutische Verfahrenstechnik e.V.

[25]  V. Fessard,et al.  Genotoxicity of a freshwater cyanotoxin, cylindrospermopsin, in two human cell lines: Caco‐2 and HepaRG , 2009, Environmental and molecular mutagenesis.

[26]  L. Ren,et al.  One-step synthesis of monodisperse, water-soluble ultra-small Fe3O4 nanoparticles for potential bio-application. , 2013, Nanoscale.

[27]  Ralph Weissleder,et al.  Noninvasive detection of clinically occult lymph-node metastases in prostate cancer. , 2003, The New England journal of medicine.

[28]  V. Rotello,et al.  The role of surface functionality in determining nanoparticle cytotoxicity. , 2013, Accounts of chemical research.

[29]  T. Tamura,et al.  Grain-oriented calcium hydroxyapatite ceramic and film prepared by magnetic alignment , 2007 .

[30]  G. Bellomo,et al.  Cytoskeleton as a target in menadione‐induced oxidative stress in cultured mammalian cells. I. Biochemical and immunocytochemical features , 1990, Journal of cellular physiology.

[31]  Takeshi Kobayashi,et al.  Cancer hyperthermia using magnetic nanoparticles , 2011, Biotechnology journal.

[32]  Qiang Yang,et al.  A Facile Approach for Fabricating Fluorescent Cellulose , 2010 .

[33]  Patrick Boisseau,et al.  Nanomedicine, Nanotechnology in medicine , 2011 .

[34]  T. Xia,et al.  Understanding biophysicochemical interactions at the nano-bio interface. , 2009, Nature materials.

[35]  Christoph Alexiou,et al.  Magnetic Drug Targeting—Biodistribution of the Magnetic Carrier and the Chemotherapeutic agent Mitoxantrone after Locoregional Cancer Treatment , 2003 .

[36]  A. C. Hunter,et al.  Nanomedicine: current status and future prospects , 2005, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[37]  C. Berry Possible exploitation of magnetic nanoparticle–cell interaction for biomedical applications , 2005 .

[38]  Morteza Mahmoudi,et al.  Assessing the in vitro and in vivo toxicity of superparamagnetic iron oxide nanoparticles. , 2012, Chemical reviews.

[39]  Zhen Cheng,et al.  Effects of nanoparticle size on cellular uptake and liver MRI with polyvinylpyrrolidone-coated iron oxide nanoparticles. , 2010, ACS nano.

[40]  Ali Khademhosseini,et al.  Micro‐ and Nanoengineering of Biomaterials for Healthcare Applications , 2013, Advanced healthcare materials.

[41]  K. Ng,et al.  Solid-state synthesis of monocrystalline iron oxide nanoparticle based ferrofluid suitable for magnetic resonance imaging contrast application , 2007 .

[42]  Rodolfo Miranda,et al.  Engineering Iron Oxide Nanoparticles for Clinical Settings , 2014, Nanobiomedicine.

[43]  H. Harris,et al.  Sequence and characterization of the human intestinal alkaline phosphatase gene. , 1988, The Journal of biological chemistry.

[44]  Albert P. Philipse,et al.  Magnetic silica dispersions: preparation and stability of surface-modified silica particles with a magnetic core , 1994 .

[45]  G. Wang,et al.  Facile synthesis of Fe3O4/SiO2 composite nanoparticles from primary silica particles , 2008 .

[46]  Tiago R. Oliveira,et al.  Application of hyperthermia induced by superparamagnetic iron oxide nanoparticles in glioma treatment , 2011, International journal of nanomedicine.

[47]  Ralph Weissleder,et al.  Nanoparticle imaging of integrins on tumor cells. , 2006, Neoplasia.

[48]  C. Thompson,et al.  Assessment of Cr(VI)-Induced Cytotoxicity and Genotoxicity Using High Content Analysis , 2012, PloS one.

[49]  L. Prodi,et al.  Proper design of silica nanoparticles combines high brightness, lack of cytotoxicity and efficient cell endocytosis. , 2013, Nanoscale.

[50]  P. Choyke,et al.  Synthesis and characterization of ultra-small superparamagnetic iron oxide nanoparticles thinly coated with silica , 2008, Nanotechnology.

[51]  M. Papi,et al.  Plasma Protein Corona Reduces the Haemolytic Activity of the Graphene Oxide Nano and Micro Flakes , 2015 .

[52]  G. Nienhaus,et al.  Engineered nanoparticles interacting with cells: size matters , 2014, Journal of Nanobiotechnology.

[53]  A. Stammati,et al.  The Caco-2 cell line as a model of the intestinal barrier: influence of cell and culture-related factors on Caco-2 cell functional characteristics , 2005, Cell Biology and Toxicology.

[54]  K. Niikura,et al.  Surface engineering of nanoparticles for therapeutic applications , 2014 .

[55]  Ajay Kumar Gupta,et al.  Synthesis and surface engineering of iron oxide nanoparticles for biomedical applications. , 2005, Biomaterials.

[56]  S. Dou,et al.  One-pot aqueous synthesis of cysteine-capped CdTe/CdS core–shell nanowires , 2014, Journal of Nanoparticle Research.

[57]  Jing Bai,et al.  Cellular uptake of nanoparticles by membrane penetration: a study combining confocal microscopy with FTIR spectroelectrochemistry. , 2012, ACS nano.

[58]  J. Klostergaard,et al.  Magnetic nanovectors for drug delivery. , 2012, Nanomedicine : nanotechnology, biology, and medicine.

[59]  M. Bañobre‐López,et al.  Magnetic nanoparticle-based hyperthermia for cancer treatment. , 2013, Reports of practical oncology and radiotherapy : journal of Greatpoland Cancer Center in Poznan and Polish Society of Radiation Oncology.

[60]  A. Bretscher,et al.  Villin: the major microfilament-associated protein of the intestinal microvillus. , 1979, Proceedings of the National Academy of Sciences of the United States of America.

[61]  A. Lu,et al.  Magnetic nanoparticles: synthesis, protection, functionalization, and application. , 2007, Angewandte Chemie.

[62]  Á. Jos,et al.  Microcystin-LR induces toxic effects in differentiated and undifferentiated Caco-2 cells , 2010, Archives of Toxicology.

[63]  M. E. Khosroshahi,et al.  Evaluation of cell viability and T2 relaxivity of fluorescein conjugated SPION-PAMAM third generation nanodendrimers for bioimaging. , 2016, Materials science & engineering. C, Materials for biological applications.

[64]  A. Vrij,et al.  Synthesis and characterization of colloidal dispersions of fluorescent, monodisperse silica spheres , 1992 .

[65]  C. Alexiou,et al.  In vitro and in vivo investigations of targeted chemotherapy with magnetic nanoparticles , 2005 .

[66]  Christian Plank,et al.  Magnetically enhanced nucleic acid delivery. Ten years of magnetofection—Progress and prospects , 2011, Advanced Drug Delivery Reviews.

[67]  H. Hofmann,et al.  Superparamagnetic nanoparticles for biomedical applications: Possibilities and limitations of a new drug delivery system , 2005 .

[68]  Sangsig Kim,et al.  Sub 5 nm magnetite nanoparticles: Synthesis, microstructure, and magnetic properties , 2007 .

[69]  G. Pojana,et al.  Coating-dependent induction of cytotoxicity and genotoxicity of iron oxide nanoparticles , 2015, Nanotoxicology.

[70]  A. Athanassiou,et al.  Toxicity Assessment of Silica Coated Iron Oxide Nanoparticles and Biocompatibility Improvement by Surface Engineering , 2014, PloS one.

[71]  W. Rubas,et al.  A human colonic cell line sharing similarities with enterocytes as a model to examine oral absorption: advantages and limitations of the Caco-2 model. , 1997, Critical reviews in therapeutic drug carrier systems.

[72]  Jiaqi Lin,et al.  Penetration of lipid membranes by gold nanoparticles: insights into cellular uptake, cytotoxicity, and their relationship. , 2010, ACS nano.

[73]  Masashige Shinkai Functional magnetic particles for medical application. , 2002, Journal of bioscience and bioengineering.

[74]  Yoo-Hun Suh,et al.  Nanotechnology, nanotoxicology, and neuroscience , 2009, Progress in Neurobiology.