Polymer-coated hexagonal upconverting nanoparticles: chemical stability and cytotoxicity.

Large (120 nm) hexagonal NaYF4:Yb, Er nanoparticles (UCNPs) were synthesized by high-temperature coprecipitation method and coated with poly(ethylene glycol)-alendronate (PEG-Ale), poly (N,N-dimethylacrylamide-co-2-aminoethylacrylamide)-alendronate (PDMA-Ale) or poly(methyl vinyl ether-co-maleic acid) (PMVEMA). The colloidal stability of polymer-coated UCNPs in water, PBS and DMEM medium was investigated by dynamic light scattering; UCNP@PMVEMA particles showed the best stability in PBS. Dissolution of the particles in water, PBS, DMEM and artificial lysosomal fluid (ALF) determined by potentiometric measurements showed that all particles were relatively chemically stable in DMEM. The UCNP@Ale-PEG and UCNP@Ale-PDMA particles were the least soluble in water and ALF, while the UCNP@PMVEMA particles were the most chemically stable in PBS. Green fluorescence of FITC-Ale-modified UCNPs was observed inside the cells, demonstrating successful internalization of particles into cells. The highest uptake was observed for neat UCNPs, followed by UCNP@Ale-PDMA and UCNP@PMVEMA. Viability of C6 cells and rat mesenchymal stem cells (rMSCs) growing in the presence of UCNPs was monitored by Alamar Blue assay. Culturing with UCNPs for 24 h did not affect cell viability. Prolonged incubation with particles for 72 h reduced cell viability to 40%-85% depending on the type of coating and nanoparticle concentration. The greatest decrease in cell viability was observed in cells cultured with neat UCNPs and UCNP@PMVEMA particles. Thanks to high upconversion luminescence, high cellular uptake and low toxicity, PDMA-coated hexagonal UCNPs may find future applications in cancer therapy.

Cytotoxicity Evaluation of Photosensitizer-Conjugated Hexagonal Upconverting Nanoparticles.

In this report, we synthesized hexagonal NaYF4:Yb,Er upconverting nanoparticles (UCNPs) of 171 nm in size with a narrow particle size distribution. To address their colloidal stabi-lity in aqueous media and to incorporate a photosensitizer that can produce reactive singlet oxygen (1O2) to kill tumor cells, UCNPs were conjugated with 6-bromohexanoic acid-functionalized Rose Bengal (RB) and coated with PEG-alendronate (PEG-Ale). The particles were thoroughly characterized by transmission electron microscopy, dynamic light scattering, ATR FTIR, X-ray photoelectron spectroscopy, thermogravimetric analysis, and spectrofluorometry, and 1O2 formation was detected using a 9,10-diphenylanthracene spectrophotometric probe. Cytotoxicity determination on rat mesenchymal stem cells by using the MTT assay showed that neutralization of the large positive surface charge of neat UCNPs with PEG-Ale and the bound RB sensitizer significantly reduced the concentration-dependent cytotoxicity. The presented strategy shows great potential for the use of these particles as a novel agent for the photodynamic therapy of tumors.

Chemical and Colloidal Stability of Polymer-Coated NaYF4:Yb,Er Nanoparticles in Aqueous Media and Viability of Cells: The Effect of a Protective Coating.

Upconverting nanoparticles (UCNPs) are of particular interest in nanomedicine for in vivo deep-tissue optical cancer bioimaging due to their efficient cellular uptake dependent on polymer coating. In this study, particles, ca. 25 nm in diameter, were prepared by a high-temperature coprecipitation of lanthanide chlorides. To ensure optimal dispersion of UCNPs in aqueous milieu, they were coated with three different polymers containing reactive groups, i.e., poly(ethylene glycol)-alendronate (PEG-Ale), poly(N,N-dimethylacrylamide-co-2-aminoethylacrylamide)-alendronate (PDMA-Ale), and poly(methyl vinyl ether-co-maleic acid) (PMVEMA). All the particles were characterized by TEM, DLS, FTIR, and spectrofluorometer to determine the morphology, hydrodynamic size and ξ-potential, composition, and upconversion luminescence. The degradability/dissolution of UCNPs in water, PBS, DMEM, or artificial lysosomal fluid (ALF) was evaluated using an ion-selective electrochemical method and UV-Vis spectroscopy. The dissolution that was more pronounced in PBS at elevated temperatures was decelerated by polymer coatings. The dissolution in DMEM was relatively small, but much more pronounced in ALF. PMVEMA with multiple anchoring groups provided better protection against particle dissolution in PBS than PEG-Ale and PDMA-Ale polymers containing only one reactive group. However, the cytotoxicity of the particles depended not only on their ability to rapidly degrade, but also on the type of coating. According to MTT, neat UCNPs and UCNP@PMVEMA were toxic for both rat cells (C6) and rat mesenchymal stem cells (rMSCs), which was in contrast to the UCNP@Ale-PDMA particles that were biocompatible. On the other hand, both the cytotoxicity and uptake of the UCNP@Ale-PEG particles by C6 and rMSCs were low, according to MTT assay and ICP-MS, respectively. This was confirmed by a confocal microscopy, where the neat UCNPs were preferentially internalized by both cell types, followed by the UCNP@PMVEMA, UCNP@Ale-PDMA, and UCNP@Ale-PEG particles. This study provides guidance for the selection of a suitable nanoparticle coating with respect to future biomedical applications where specific behaviors (extracellular deposition vs. cell internalization) are expected.

PMVEMA-coated upconverting nanoparticles for upconversion-linked immunoassay of cardiac troponin.

Surface engineering of upconverting nanoparticles (UCNPs) is crucial for their bioanalytical applications. Here, an antibody specific to cardiac troponin I (cTnI), an important biomarker for acute myocardial infection, was covalently immobilized on the surface of UCNPs to prepare a label for the detection of cTnI biomarker in an upconversion-linked immunoassay (ULISA). Core-shell UCNPs (NaYF4:Yb,Tm@NaYF4) were first coated with poly(methyl vinyl ether-alt-maleic acid) (PMVEMA) and then conjugated to antibodies. The morphology (size and uniformity), hydrodynamic diameter, chemical composition, and amount of coating on the of UCNPs, as well as their upconversion luminescence, colloidal stability, and leaching of Y3+ ions into the surrounding media, were determined. The developed ULISA allowed reaching a limit of detection (LOD) of 0.13 ng/ml and 0.25 ng/ml of cTnI in plasma and serum, respectively, which represents 12- and 2-fold improvement to conventional enzyme-linked immunosorbent based on the same immunoreagents.

Poly(N,N-dimethylacrylamide)-coated upconverting NaYF4:Yb,Er@NaYF4:Nd core-shell nanoparticles for fluorescent labeling of carcinoma cells.

Upconverting luminescent lanthanide-doped nanoparticles (UCNP) belong to promising new materials that absorb infrared light able to penetrate in the deep tissue level, while emitting photons in the visible or ultraviolet region, which makes them favorable for bioimaging and cell labeling. Here, we have prepared upconverting NaYF4:Yb,Er@NaYF4:Nd core-shell nanoparticles, which were coated with copolymers of N,N-dimethylacrylamide (DMA) and 2-(acryloylamino)-2-methylpropane-1-sulfonic acid (AMPS) or tert-butyl [2-(acryloylamino)ethyl]carbamate (AEC-Boc) with negative or positive charges, respectively. The copolymers were synthesized by a reversible addition-fragmentation chain transfer (RAFT) polymerization, reaching Mn ~ 11 kDa and containing ~ 5 mol% of reactive groups. All copolymers contained bisphosphonate end-groups to be firmly anchored on the surface of NaYF4:Yb,Er@NaYF4:Nd core-shell nanoparticles. To compare properties of polymer coatings, poly(ethylene glycol)-coated and neat UCNP were used as a control. UCNP with various charges were then studied as labels of carcinoma cells, including human hepatocellular carcinoma HepG2, human cervical cancer HeLa, and rat insulinoma INS-1E cells. All the particles proved to be biocompatible (nontoxic); depending on their ξ-potential, the ability to penetrate the cells differed. This ability together with the upconversion luminescence are basic prerequisites for application of particles in photodynamic therapy (PDT) of various tumors, where emission of nanoparticles in visible light range at ~ 650 nm excites photosensitizer.

Poly(ethylene glycol)-Alendronate-Coated Magnetite Nanoparticles Do Not Alter Cardiovascular Functions and Red Blood Cells' Properties in Hypertensive Rats.

In this study, magnetite nanoparticles were prepared and coated with poly(ethylene glycol) terminated by alendronate to ensure firm binding to the iron oxide surface. Magnetic nanoparticles, designated as magnetite coated with poly(ethylene glycol)-alendronate (Fe3O4@PEG-Ale), were characterized in terms of number-average (Dn) and hydrodynamic (Dh) size, ζ-potential, saturation magnetization, and composition. The effect of particles on blood pressure, vascular functions, nitric oxide (NO), and superoxide production in the tissues of spontaneously hypertensive rats, as well as the effect on red blood cell (RBC) parameters, was investigated after intravenous administration (1 mg Fe3O4/kg of body weight). Results showed that Fe3O4@PEG-Ale particles did negatively affect blood pressure, heart rate and RBC deformability, osmotic resistance and NO production. In addition, Fe3O4@PEG-Ale did not alter functions of the femoral arteries. Fe3O4@PEG-Ale induced increase in superoxide production in the kidney and spleen, but not in the left heart ventricle, aorta and liver. NO production was reduced only in the kidney. In conclusion, the results suggest that acute intravenous administration of Fe3O4@PEG-Ale did not produce negative effects on blood pressure regulation, vascular function, and RBCs in hypertensive rats.

Colloidally Stable P(DMA-AGME)-Ale-Coated Gd(Tb)F3:Tb3+(Gd3+),Yb3+,Nd3+ Nanoparticles as a Multimodal Contrast Agent for Down- and Upconversion Luminescence, Magnetic Resonance Imaging, and Computed Tomography.

Multimodal imaging, integrating several modalities including down- and up-conversion luminescence, T 1- and T 2(T 2*)-weighted MRI, and CT contrasting in one system, is very promising for improved diagnosis of severe medical disorders. To reach the goal, it is necessary to develop suitable nanoparticles that are highly colloidally stable in biologically relevant media. Here, hydrophilic poly(N,N-dimethylacrylamide-N-acryloylglycine methyl ester)-alendronate-[P(DMA-AGME)-Ale]-coated Gd(Tb)F3:Tb3+(Gd3+),Yb3+,Nd3+ nanoparticles were synthesized by a coprecipitation method in ethylene glycol (EG) followed by coating with the polymer. The particles were tho-roughly characterized by a dynamic light scattering (DLS), transmission electron microscopy (TEM), Fourier-transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), X-ray energy dispersive spectroscopy (EDAX), selected area electron diffraction (SAED), elemental ana-lysis and fluorescence spectroscopy. Aqueous particle dispersions exhibited excellent colloidal stability in water and physiological buffers. In vitro toxicity assessments suggested no or only mild toxicity of the surface-engineered Gd(Tb)F3:Tb3+(Gd3+),Yb3+,Nd3+ particles in a wide range of concentrations. Internalization of the particles by several types of cells, including HeLa, HF, HepG2, and INS, was confirmed by a down- and up-conversion confocal microscopy. Newly developed particles thus proved to be an efficient contrast agent for fluorescence imaging, T 1- and T 2(T 2*)-weighted magnetic resonance imaging (MRI), and computed tomography (CT).

Doxorubicin-Conjugated Iron Oxide Nanoparticles: Surface Engineering and Biomedical Investigation.

Development of therapeutic systems to treat glioblastoma, the most common and aggressive brain tumor, belongs to priority tasks in cancer research. We have synthesized colloidally stable magnetic nanoparticles (Dh =336 nm) coated with doxorubicin (Dox) conjugated copolymers of N,N-dimethylacrylamide and either N-acryloylglycine methyl ester or N-acryloylmethyl 6-aminohexanoate. The terminal carboxyl groups of the copolymers were reacted with alendronate by carbodiimide formation. Methyl ester groups were then transferred to hydrazides for binding Dox by a hydrolytically labile hydrazone bond. The polymers were subsequently bound on the magnetic nanoparticles through bisphosphonate terminal groups. Finally, the anticancer effect of the Dox-conjugated particles was investigated using the U-87 glioblastoma cell line in terms of particle internalization and cell viability, which decreased to almost zero at a concentration of 100 µg of particles per ml. These results confirmed that poly(N,N-dimethylacrylamide)-coated magnetic nanoparticles can serve as a solid support for Dox delivery to glioblastoma cells.