title: Fullerenol cytotoxicity in kidney cells is associated with cytoskeleton disruption, autophagic vacuole accumulation, and mitochondrial dysfunction.
abstract: Water soluble fullerenes, such as the hydroxylated fullerene, fullerenol (C₆₀OHx), are currently under development for diagnostic and therapeutic biomedical applications in the field of nanotechnology. These molecules have been shown to undergo urinary clearance, yet there is limited data available on their renal biocompatibility. Here we examine the biological responses of renal proximal tubule cells (LLC-PK1) exposed to fullerenol. Fullerenol was found to be cytotoxic in the millimolar range, with viability assessed by the sulforhodamine B and trypan blue assays. Fullerenol-induced cell death was associated with cytoskeleton disruption and autophagic vacuole accumulation. Interaction with the autophagy pathway was evaluated in vitro by Lysotracker Red dye uptake, LC3-II marker expression and TEM. Fullerenol treatment also resulted in coincident loss of cellular mitochondrial membrane potential and ATP depletion, as measured by the Mitotracker Red dye and the luciferin-luciferase assays, respectively. Fullerenol-induced ATP depletion and loss of mitochondrial potential were partially ameliorated by co-treatment with the autophagy inhibitor, 3-methyladenine. In vitro fullerenol treatment did not result in appreciable oxidative stress, as measured by lipid peroxide and glutathione content. Based on these data, it is hypothesized that cytoskeleton disruption may be an initiating event in fullerenol cytotoxicity, leading to subsequent autophagy dysfunction and loss of mitochondrial capacity. As nanoparticle-induced cytoskeleton disruption, autophagic vacuole accumulation and mitochondrial dysfunction are commonly reported in the literature, the proposed mechanism may be relevant for a variety of nanomaterials.
authors: Nanotechnology Characterization Lab (NCL), Advanced Technology Program, SAIC-Frederick, Inc., NCI-Frederick, Frederick, MD 21702, USA.
authors: Denise N
authors: DN
authors: Johnson-Lyles
authors: Kimberly
authors: K
authors: Peifley
authors: Stephen
authors: S
authors: Lockett
authors: Barry W
authors: BW
authors: Neun
authors: Matthew
authors: M
authors: Hansen
authors: Jeffrey
authors: J
authors: Clogston
authors: Stephan T
authors: ST
authors: Stern
authors: Scott E
authors: SE
authors: McNeil
mesh_terms: D000818:Animals; D001343:Autophagy / Q000187:drug effects* / Q000502:physiology; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D003599:Cytoskeleton / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D003603:Cytotoxins / Q000633:toxicity; D004305:Dose-Response Relationship, Drug; D037741:Fullerenes / Q000633:toxicity*; D007668:Kidney / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D018374:LLC-PK1 Cells; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D013552:Swine; D014617:Vacuoles / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology
chemical_list: D003603:Cytotoxins; D037741:Fullerenes; C108127:fullerenol