The effects of amyloid precursor protein C-105 expression and ganglioside GM1 on PC12 cell vulnerability to the calcium ionophore A23 187 and hydrogen peroxide
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Abstract
Specific mutations in the gene for the beta amyloid precursor protein (APP) cause
Alzheimer's disease (AD). A key neuropathological hallmark of AD is extracellular
neuritic plaques. The core component of plaques is AB, a 39-43 amino acid peptide
derived from APP. APP C-100 and APP C-105 are C-terminal fragments of APP, 100
and 105 amino acids long, respectively. APP C-100 is a normal metabolite of APP. AI3 is
located at the N-terminus of the APP C-100 sequence.
To determine whether APP C-105 expression alters cellular vulnerability to calcium
and hydrogen peroxide, rat PC12 cells were modified to overexpress APP C-105.
Permanent transfectants (clones) were selected, then characterized by standard molecular
biological techniques. DNA and mRNA corresponding to APP C-105 were detected in
APP C-105 transfectants, but not in wild type controls. Aggregated APP C-105 was
detected in cell lysates and conditioned media from APP C-105 transfectants, but was
absent or detected at lower concentrations in vector-transfected and wild type controls.
Cell survival as a function of concentration was determined for A23 l 87, a Ca^2+
ionophore, and hydrogen peroxide in APP C-105 transfectants and vector-transfected
controls. Cells were exposed to A23 l 87 or hydrogen peroxide for 24 hours in RP:MI
media containing 3 μM insulin, and survival was quantitated using the tetrazolium dye,
MTT. APP C-105 expression significantly increased PC 12 cell vulnerability to A23 l 87,
and significantly decreased vulnerability to hydrogen peroxide.
Other experiments were performed with GM 1 ganglioside, which is known to protect
cells against numerous insults. When wild type PC 12 cells or APP C-10 5 transfectants
were exposed to a toxin and GMl concurrently for 24 hours, GMl produced
concentration-dependent inhibition of A23 l 87 toxicity in wild type PC 12 cells but was
ineffective against hydrogen peroxide in both wild type PC12 and APP C-105-transfected
clones.
The current study has demonstrated that expression of APP C-105 protects
PC 12 cells against hydrogen peroxide, but exacerbates the effect of calcium influx.
In conjunction with other reports, this study indicates that APP C-105 is an
important regulator of cellular homeostasis. Therefore, the pattern of APP
processing may alter vulnerability to neurotoxic insults.
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