Time course assessment of methylmercury effects on C6 glioma cells: submicromolar concentrations induce oxidative DNA damage and apoptosis.
Organic mercury is a well-known neurotoxicant although its mechanism of action has not been fully clarified. In addition to a direct effect on neurons, much experimental evidence supports an involvement of the glial component. We assessed methylmercury hydroxide (MeHgOH) toxicity in a glial model, C6 glioma cells, exposed in the 10(-5)-10(-8) M range. The time course of the effects was studied by time-lapse confocal microscopy and supplemented with biochemical data. We have monitored cell viability and proliferation rate, reactive oxygen species (ROS), mitochondrial transmembrane potential, DNA oxidation, energetic metabolism and modalities of cell death. The earliest effect was a measurable ROS generation followed by oxidative DNA damage paralleled by a partial mitochondrial depolarization. The effect on cell viability was dose dependent. TUNEL, caspase activity and real-time morphological observation of calcein-loaded cells showed that apoptosis was the only detectable mode of cell death within this concentration range. N-acetyl-cysteine (NAC) or reduced glutathione (GSH) completely prevent the apoptotic effect of MeHgOH. The lowest effective MeHgOH concentration was 10(-7) M for ROS and DNA OH-adducts generation. The effect of submicromolar concentrations of MeHgOH on C6 cells could be relevant in the developmental neurotoxicity caused by low dose, long-term exposures, such as those of food origin. In addition, we have shown that the same concentrations are effective in the induction of DNA oxidative damage, with further potential pathogenetic implications.
Belletti
Silvana
S
Department of Experimental Medicine, Histology Section, University of Parma, Via Volturno 39, I-43100 Parma, Italy.
Orlandini
Guido
G
Vettori
Maria Vittoria
MV
Mutti
Antonio
A
Uggeri
Jacopo
J
Scandroglio
Renato
R
Alinovi
Rossella
R
Gatti
Rita
R
D000080242:8-Hydroxy-2'-Deoxyguanosine; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002455:Cell Division / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004249:DNA Damage; D003849:Deoxyguanosine / Q000031:analogs & derivatives*; D005452:Fluoresceins; D005910:Glioma; D020287:In Situ Nick-End Labeling; D008564:Membrane Potentials / Q000502:physiology; D008767:Methylmercury Compounds / Q000633:toxicity*; D018613:Microscopy, Confocal; D008928:Mitochondria / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects*; D051381:Rats; D013997:Time Factors; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000187:drug effects / Q000378:metabolism
D005452:Fluoresceins; D008767:Methylmercury Compounds; C029569:diacetyldichlorofluorescein; D000080242:8-Hydroxy-2'-Deoxyguanosine; D000255:Adenosine Triphosphate; C020771:methylmercury hydroxide; D003849:Deoxyguanosine