A comparative study of neurotoxic potential of synthesized polysaccharide-coated and native ferritin-based magnetic nanoparticles

Abstract

Aim: To analyze the neurotoxic potential of synthesized magnetite nanoparticles coated by dextran, hydroxyethyl starch, oxidized hydroxyethyl starch, and chitosan, and magnetic nanoparticles combined with ferritin as a native protein.

Methods: The size of nanoparticles was analyzed using photon correlation spectroscopy, their effects on the conductance of planar lipid membrane by planar lipid bilayer technique, membrane potential and acidification of synaptic vesicles by spectrofluorimetry, and glutamate uptake and ambient level of glutamate in isolated rat brain nerve terminals (synaptosomes) by radiolabeled assay.

Results: Uncoated synthesized magnetite nanoparticles and nanoparticles coated by different polysaccharides had no significant effect on synaptic vesicle acidification, the initial velocity of L-[(14)C]glutamate uptake, ambient level of L-[(14)C]glutamate and the potential of the plasma membrane of synaptosomes, and conductance of planar lipid membrane. Native ferritin-based magnetic nanoparticles had no effect on the membrane potential but significantly reduced L-[(14)C]glutamate transport in synaptosomes and acidification of synaptic vesicles.

Conclusions: Our study indicates that synthesized magnetite nanoparticles in contrast to ferritin have no effects on the functional state and glutamate transport of nerve terminals, and so ferritin cannot be used as a prototype, analogue, or model of polysaccharide-coated magnetic nanoparticle in toxicity risk assessment and manipulation of nerve terminals by external magnetic fields. Still, the ability of ferritin to change the functional state of nerve terminals in combination with its magnetic properties suggests its biotechnological potential.

Figure 1

Dynamic light scattering histogram: assessment of the size of magnetite nanoparticles (5 mg/mL) coated by dextran in the standard salt solution. The measurements were performed in 2 minutes.

Figure 2

Membrane potential of the synaptosomes after the addition of synthesized magnetite nanoparticles (MNP). The suspension of the synaptosomes was equilibrated with potential-sensitive dye Rhodamine 6G (0.5 µM); when the steady level of the dye fluorescence had been reached, MNP (2 mg/mL) were added (arrows: 1 – uncoated MNP; 2 – MNP coated by dextran; 3 – coated by hydroxyethyl starch; 4 – coated by oxidized hydroxyethyl starch; and 5 – coated by chitosan). Trace represents four experiments performed with different preparations.

Figure 3

Lack of the influence of synthesized magnetite nanoparticles (MNP) coated by dextran on the conductance of bilayer lipid membrane. MNP (arrow) were added from the cis-side of membrane at a final concentration of 0.1 mg/mL.

Figure 4

Acidification of synaptosomes in the presence of synthesized magnetite nanoparticles (MNP). The synaptosomes were equilibrated with acridine orange (5 µM); when the steady level of the dye fluorescence had been reached, MNP (2 mg/mL) were added (arrows: 1 – uncoated MNP; 2 – MNP coated by dextran; 3 – coated by hydroxyethyl starch; 4 – coated by oxidized hydroxyethyl starch; and 5 – coated by chitosan). Trace represents four experiments performed with different preparations.

Figure 5

Time course of uptake of L-[¹⁴C]glutamate by control synaptosomes (solid line); synaptosomes in presence of dextran-coated magnetite nanoparticles (MNP) (2 mg/mL) (dashed line). Uptake was initiated by the addition of L-[¹⁴C]glutamate to synaptosomes, after incubation the samples were rapidly sedimented and radioactivity was determined as described in Materials and methods. Data are presented as mean ± standard error of the mean of three independent experiments.

Figure 6

Comparison of the effects of synthesized magnetite nanoparticles (MNP) coated by dextran (2 mg/mL) (synthesized nanoparticle) and ferritin (100 μg/mL) (native nanoparticle) on the initial velocity of L-[¹⁴C]glutamate uptake by synaptosomes (A), the ambient level of L-[¹⁴C]glutamate at 5 minutes time point (B), synaptic vesicle acidification (C), and the membrane potential (D). Data are presented as mean ± standard error of the mean of three independent experiments. *Р≤0.05 as compared to control.