Interaction of Fullerene Nanoparticles With Biomembranes: From the Partition in Lipid Membranes to Effects on Mitochondrial Bioenergetics

Abstract

Partition and localization of C60 and its derivative C60(OH)18-22 in lipid membranes and their impact on mitochondrial activity were studied, attempting to correlate those events with fullerene characteristics (size, surface chemistry, and surface charge). Fluorescence quenching studies suggested that C60(OH)18-22 preferentially populated the outer regions of the bilayer, whereas C60 preferred to localize in deeper regions of the bilayer. Partition coefficient values indicated that C60 exhibited higher affinity for dipalmitoylphosphatidylcholine and mitochondrial membranes than C60(OH)18-22. Both fullerenes affected the mitochondrial function, but the inhibitory effects promoted by C60 were more pronounced than those induced by C60(OH)18-22 (up to 20 nmol/mg of mitochondrial protein). State 3 and p-trifluoromethoxyphenylhydrazone-uncoupled respirations are inhibited by both fullerenes when glutamate/malate or succinate was used as substrate. Phosphorylation system and electron transport chain of mitochondria are affected by both fullerenes, but only C60 increased the inner mitochondrial membrane permeability to protons, suggesting perturbations in the structure and dynamics of that membrane. At concentrations of C60(OH)18-22 above 20 nmol/mg of mitochondrial protein, the activity of FoF1-ATP synthase was also decreased. The evaluation of transmembrane potential showed that the mitochondria phosphorylation cycle decreased upon adenosine diphosphate addition with increasing fullerenes concentration and the time of the repolarization phase increased as a function of C60(OH)18-22 concentration. Our results suggest that the balance between hydrophilicity and hydrophobicity resulting from the surface chemistry of fullerene nanoparticles, rather than the cluster size or the surface charge acquired by fullerenes in water, influences their membrane interactions and consequently their effects on mitochondrial bioenergetics.

Toxicity of polyhydroxylated fullerene to mitochondria

“…Functional mitochondria display a transmembrane electric potential across the inner mitochondrial membrane, which is generated by the translocation of protons from the matrix to the intermembrane space. Therefore, it is expectable that proton abundance in this mitochondrial compartment could attract the negative charged fullerenol, favoring the interaction of the nanoparticles and mitochondrial membrane [50]. Our membrane fluidity studies have proved that fullerenol perturbed the mitochondrial membrane in polar protein regions.…”

“…In isolated mitochondria, respiration state 4 is related to the proton leak, while state 3 is connected with the ETC and substrate transport [57]. Fullerenol stimulated the respiration of state 4, which is generally interpreted as resulting from an increase of the inner mitochondrial membrane permeability to protons [50]. Accordingly, the loss of membrane potential is responsible for the increase of O 2 consumption [58].…”

Effect of fullerene on the dispersibility of nanostructured lipid particles and encapsulation in sterically stabilized emulsions

“…These aggregates assembled into rather big dark coloured lumps made of both fullerenes and lipids together. Spontaneous aggregation and clustering of fullerenes in aqueous media has been discussed by several researchers [7,14,41,42]. Agglomeration and surface 6 adhesion has also been demonstrated with surfactant micellar systems, cellular membranes as well as model lipid bilayers [6,8,12,14].…”

“…Stabilization may also occur due to the spontaneous penetration of fullerenes in lipid bilayers, which ideally form a part of liposomes (vesicles) [8,46] (Figure 5a). Fullerene insertion in bilayers was previously investigated by several researchers [6,[8][9][10][12][13][14]. Aforementioned low (0.1%) lipid dispersions were stable, but 5% lipid dispersions prepared using various fullerene concentrations faced severe destabilization which can be attributed partially to the following aspects: 1) self-aggregation of fullerenes in aqueous media [7,42,47], 2) adhesion of fullerenes to lipid membranes [8,10,46], 3) penetration of fullerenes [8] and/or small clusters [10] in membranes, 4) apparent high viscosity of liquid crystalline phase and 5) layer-by-layer growth of above aggregates during cooling and/or vigorous shaking where initial clusters act as templates (or nuclei) for the adhesion of additional layers [10].…”

“…Considerable theoretical (simulation-based) [9][10][11][12][13] and experimental [8,14] efforts have been taken to investigate fullerene interactions with biological as well as artificial membrane systems. These reports mainly involved studies with planar lipid bilayers, spherical micelles or vesicles.…”