title: The selection of serum-independent PC12 cells for a more-reliable manganese cytotoxicity test.
abstract: A major issue concerning the protocols of heavy metal cytotoxicity tests with PC12 cells was the hypothesis that serum in the culture medium might sequester the metal, thus altering the results obtained. However, serum withdrawal impairs the viability of PC12 cells themselves, thus impeding cytotoxicity testing in the absence of serum. In this study, we repeatedly selected undifferentiated, totally non-adherent PC12 cells in Petri dishes. Surprisingly, we discovered that these cells could survive and proliferate in serum-free medium. Moreover, features such as NGF-responsiveness, resazurin reduction potential, doubling rate, protein content, and basal caspase-3 enzyme activity, were equivalent to those exhibited by standard PC12 cultures. Further experiments aimed at fully characterising these serum-independent PC12 cells are in progress. These cells enabled cytotoxicity experiments to be conducted with manganese, both in serum-supplemented and in serum-deprived medium. The results demonstrated that serum removal decreased the LC50 of manganese from 250microM to 32microM, without affecting the internalisation of the metal. The data exclude an early competitive effect of serum on metal internalisation; rather, they suggest a late protective mechanism mediated by serum against the cytotoxic effect of the already-internalised metal.
authors.affiliation: Dipartimento di Fisica, Università di Parma, Parma, Italy.
authors.forename: Enrico
authors.initials: E
authors.lastname: Bignetti
authors.forename: Daniela
authors.initials: D
authors.lastname: Franchi
authors.forename: Alessandra
authors.initials: A
authors.lastname: Gorreri
authors.forename: Antonella
authors.initials: A
authors.lastname: Fusari
authors.forename: Giovanni
authors.initials: G
authors.lastname: Del Bono
authors.forename: Antonio
authors.initials: A
authors.lastname: Ubaldi
authors.forename: Gaetano
authors.initials: G
authors.lastname: Aiello
mesh_terms: D000310:Adrenal Gland Neoplasms; D000826:Animal Testing Alternatives / Q000379:methods; D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D016895:Culture Media, Serum-Free; D008345:Manganese / Q000633:toxicity*; D016716:PC12 Cells / Q000187:drug effects / Q000502:physiology*; D010673:Pheochromocytoma; D051381:Rats
chemical_list: D016895:Culture Media, Serum-Free; D008345:Manganese