Fragment 31–35 of β-amyloid peptide induces neurodegeneration in rat cerebellar granule cells via bax gene expression and caspase-3 activation: A crucial role for the redox state of methionine-35 residue

The amyloid β-peptide (AβP) is the major protein component of brain senile plaques in Alzheimer's disease. The redox state of methionine-35 residue plays a critical role in peptide neurotoxic actions. We used the fragment 31–35 of AβP [AβP(31–35)], containing a single methionine-35 residue (Met-35), to investigate the relationship between the oxidative state of Met-35 and neurotoxic and pro-apoptotic actions induced by the peptide; in rat cerebellar granule cells (CGC), we compared the effects of AβP(31–35), in which the Met-35 is present in the reduced state, with those of a modified peptide with oxidized Met-35 [AβP(31–35)Met-35OX], as well as an AβP-derivative with Met-35 substituted by norleucine [AβP(31–35)Nle-35]. AβP(31–35) induced a time-dependent decrease in cell viability. AβP(31–35)Met-35OX was significantly less potent, but still induced a significant decrease in cell viability compared to control. No toxic effects were observed after treatment with AβP(31–35)Nle-35. AβP(31–35) induced a 2-fold increase in bax mRNA levels after 4 h, whereas AβP(31–35)Met-35OX raised bax mRNA levels by 41% and AβP(31–35)Nle-35 had no effect. Finally, AβP(31–35) caused a 43% increase in caspase-3 activity after 24 h; AβP(31–35)Met-35OX caused only a 18% increase, and AβP(31–35)Nle-35 had no effect. These findings suggest that AβP(31–35)-induced neurodegeneration in CGC is mediated by a selective early increase in bax mRNA levels followed by delayed caspase-3 activation; the redox state of the single Met-35 residue is crucial in the occurrence and extent of the above phenomena.