The production of amyloid beta peptide is a critical requirement for the viability of central neurons - PubMed (original) (raw)

The production of amyloid beta peptide is a critical requirement for the viability of central neurons

Leigh D Plant et al. J Neurosci. 2003.

Abstract

The amyloid beta peptide (Abeta) is a product of the sequential gamma- and beta-secretase cleavage of amyloid precursor protein. Inhibitors of secretase enzymes have been proposed as a potential therapeutic strategy in the treatment of Alzheimer's disease. Here, we investigate the effect of inhibiting these key enzymes on the viability of a range of cell types. Treatment of rat cortical neurons for 24 hr with secretase inhibitors or an antibody that binds Abeta resulted in a marked reduction in cell viability, as measured by MTT reduction. Incubation with secretase inhibitors caused similar effects on other neuronal cell types (rat cerebellar granule neurons and the human SH-SY5Y cell line). Interestingly, rat astrocytes and a number of non-neuronal cell lines investigated (HEK293, DDT1-FM2, and human teratorhabdoid tumor cells) were unaffected by incubation with secretase inhibitors. The coincubation of Abeta1-40 prevented the toxicity of secretase inhibitors in neuronal cells. Abeta1-40 was protective in a concentration-dependent manner, and its effects were significant at concentrations as low at 10 pm. Importantly, the protective effects of Abeta were Abeta size-form specific, with the Abeta1-42 size form affording limited protection and the Abeta25-35 size form having very little protective effect. The present study demonstrates that inhibition of beta-or gamma-secretase activity induces death in neuronal cells. Importantly, this toxicity, which our data suggest is a consequence of a decline in neuronal Abeta levels, was absent in non-neuronal cells. This study further supports a key physiological role for the enigmatic Abeta peptide.

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Figures

Figure 1.

Figure 1.

Effect of γ-secretase inhibition on cell morphology. Phase-contrast photomicrographs of cortical neurons after 24 hr incubation with 1 n

m

Aβ1–40 (A), 24 hr incubation with 10 μ

m

γ-IV (B), and 24 hr coincubation of 1 n

m

Aβ1–40 and 10 μ

m

γ-IV (C). Scale bar, 100 μm.

Figure 2.

Figure 2.

Effect of secretase inhibition on cell viability. A, Incubation for 24 hr with γ- or β-secretase inhibitors on the viability of cortical neurons (n = 9 for each condition). B, Time dependence of γ-IV toxicity on cortical cultures compared with vehicle-treated controls. Values are the mean of four experiments for each time point. C, Effect of 24 hr incubation with secretase inhibitors on cell death in cerebellar granule neurons (open bars; n = 9), the SH-SY5Y cell line (hatched bars; _n_=9), and rat astrocytes cultures (filled bars; n = 9). D, Effect of 48 hr incubation with secretase inhibitors on the viability of non-neuronal cell lines DDT1-FM2 (open bars; n = 4), HEK293 (hatched bars; n = 4), and human teratorhabdoid tumor cells (filled bars; _n_=4). γ-IX, γ-secretase inhibitor.

Figure 3.

Figure 3.

Effect of secretase inhibition on Aβ levels in neurons. Fluorescence photomicrographs of cerebellar granule neurons under control conditions (A), after 24 hr incubation with 10 μ

m

γ-IV (B), and after 24 hr incubation with 100 n

m

βSI (C). Scale bar, 30 μm.

Figure 4.

Figure 4.

Effect of amyloid proteins on γ-secretase inhibitor neurotoxicity. A, Effect of 24 hr incubation with γ-secretase inhibitor on the viability of cortical neurons (open bars; n = 9), cerebellar granule neurons (striped bars; n = 9), and the SH-SY5Y cell line (hatched bars; n = 9) in the presence of different concentrations of Aβ1–40. B, Effect of 24 hr incubation with γ-secretase inhibitor (10 μ

m

) on the viability of cortical neurons (open bars; n = 9), cerebellar granule neurons (striped bars; n = 9), and the SH-SY5Y cell line (hatched bars; n = 9) in the presence of different Aβ size forms at a concentration of 1 n

m

in each case. C, Toxicity of the monoclonal antibody 3D6 (1 μg/ml, 24 hr) on cortical neurons. Toxicity was reversed by coincubation with Aβ1–40 (1 n

m

) but not Aβ25–35 (1 n

m

). Aβ peptides alone had no effect on cell survival. In each case, n = 4.

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