Cytotoxic and genotoxic effects mediated by M2 muscarinic receptor activation in human glioblastoma cells

Maria Di Bari, Vanessa Tombolillo, Claudia Conte, Emila Castigli, Miriam Sciaccaluga, Egidio Iorio, Giulia Carpinelli, Ruggero Ricordy, Mario Fiore, Francesca Degrassi, Ada Maria Tata

Research output: Contribution to journalArticlepeer-review

Abstract

Glioblastomas are the most common brain tumors in humans. Previously, we demonstrated that the muscarinic receptor agonist, arecaidine propargyl ester, via M2 receptors, inhibits cell proliferation in a time and dose-dependent manner and induces a severe apoptosis in human U251 and U87 glioblastoma cell lines. In order to clarify the mechanisms causing apoptosis after arecaidine treatment, we analyzed the ability of arecaidine to induce oxidative stress. By dichloro-dihydro-fluorescein diacetate (DCFDA) staining, we demonstrated that arecaidine increased the intracellular ROS levels. ROS accumulation was completely counteracted by the ROS scavenger, N-acetyl-l-cysteine (NAC). Apoptotic cell death appeared directly correlated to ROS production since NAC was able to counteract this effect. Although there was an up-regulation of some detoxifying enzyme expression such as superoxide dismutase (MnSOD) and sirtuin-1 (SIRT1), the cytotoxic effect caused by arecaidine treatment caused DNA damage, as demonstrated by the increase of histone γ-H2AX positive cells, and chromosomal aberrations. These effects were mediated by M2 receptor activation; in fact after silencing of M2 receptors by siRNA, the increase of γ-H2AX positive cells was abolished. In conclusion, in addition to a cytostatic effect previously described, in the present study we have better characterized the mechanisms causing the cytotoxic effects and the apoptotic cell death in glioblastoma cells after M2 receptor activation. These data allow to consider this receptor a new interesting therapeutic tool for the glioblastoma treatment.

Original languageEnglish
Pages (from-to)261-270
Number of pages10
JournalNeurochemistry International
Volume90
DOIs
Publication statusPublished - Nov 1 2015

Keywords

  • Acetylcholine
  • Apoptosis
  • DNA damage
  • M2 receptors
  • ROS

ASJC Scopus subject areas

  • Cellular and Molecular Neuroscience
  • Cell Biology

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