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. 2000 Dec 15;20(24):9119-25.
doi: 10.1523/JNEUROSCI.20-24-09119.2000.

Delaying caspase activation by Bcl-2: A clue to disease retardation in a transgenic mouse model of amyotrophic lateral sclerosis

S Vukosavic  1 , L Stefanis, V Jackson-Lewis, C Guégan, N Romero, C Chen, M Dubois-Dauphin, S Przedborski
Affiliations

Delaying caspase activation by Bcl-2: A clue to disease retardation in a transgenic mouse model of amyotrophic lateral sclerosis

S Vukosavic et al. J Neurosci. .

Abstract

Molecular mechanisms of apoptosis may participate in motor neuron degeneration produced by mutant copper/zinc superoxide dismutase (mSOD1), the only proven cause of amyotrophic lateral sclerosis (ALS). Consistent with this, herein we show that the spinal cord of transgenic mSOD1 mice is the site of the sequential activation of caspase-1 and caspase-3. Activated caspase-3 and its produced beta-actin cleavage fragments are found in apoptotic neurons in the anterior horn of the spinal cord of affected transgenic mSOD1 mice; although such neurons are few, their scarcity should not undermine the potential importance of apoptosis in the overall mSOD1-related neurodegeneration. Overexpression of the anti-apoptotic protein Bcl-2 attenuates neurodegeneration and delays activation of the caspases and fragmentation of beta-actin. These data demonstrate that caspase activation occurs in this mouse model of ALS during neurodegeneration. Our study also suggests that modulation of caspase activity may provide protective benefit in the treatment of ALS, a view that is consistent with our recent demonstration of caspase inhibition extending the survival of transgenic mSOD1 mice.

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Figures

Fig. 1.
Fig. 1.
Caspase-1 and caspase-3 mRNA levels are altered in the spinal cords of transgenic mSOD1 mice. A, B, Trend toward increased levels of caspase-1 mRNA in transgenic mSOD1 mice at the beginning of symptoms and at end-stage. A, C, Significant increased caspase-3 mRNA levels in transgenic mSOD1 mice at the beginning of symptoms and at end-stage; *p < 0.05 higher than age-matched nontransgenic controls, Newman–Keulspost hoc test. N, Nontransgenic;A, asymptomatic; O, onset of symptoms;E, end-stage.
Fig. 2.
Fig. 2.
Activation of caspase-1 and caspase-3 in the spinal cords of transgenic mSOD1 mice. A, B, By 12 weeks of age the levels of pro-caspase-1 (45 kDa; black bars) decrease, whereas those of its cleaved fragment (20 kDa; white bars) increase in transgenic mSOD1 mice. A, C, A few weeks later the same phenomenon occurs for pro-caspase-3 (32 kDa;black bars) and its cleaved fragment (17 kDa;white bars); *p < 0.05 and †p < 0.01 different from age-matched nontransgenic controls, Newman–Keuls post hoc test.N, Nontransgenic; A, asymptomatic (at 4, 6, 8, 12 weeks of age); O, onset of symptoms;E, end-stage; C, cerebellum.
Fig. 3.
Fig. 3.
Increased caspase-1- and caspase-3-like activities in the spinal cords of transgenic mSOD1 mice. Caspase-1-like activity (black bars) increases before caspase-3-like activity (white bars), but both peak at the beginning of symptoms. Data are mean ± SEM for 5–15 mice per group; *p < 0.05 and **p < 0.01 higher than age-matched nontransgenic controls, Newman–Keulspost hoc test. N, Nontransgenic;A, asymptomatic (at 4, 6, 8, 12 weeks of age);O, onset of symptoms; E, end-stage;C, cerebellum.
Fig. 4.
Fig. 4.
In nontransgenic controls a large number of cells from the lumbar segment are strongly immunoreactive for pro-caspase-1 (A) and pro-caspase-3 (D); those cells have a neuronal morphology (see higher magnification inB and E). In end-stage transgenic mSOD1, there is a dramatic loss of pro-caspase-1-positive (C) and pro-caspase-3-positive (F) neurons in the lumbar segment. Conversely, specific immunostaining for CM1 (G; seearrow) and fractin (H; seearrow) is seen only in symptomatic transgenic mSOD1 mice within apoptotic cells (see insets;arrowheads indicate apoptotic chromatin clumps) and colocalizes with the neuronal marker neurofilament (I), but not with the glial marker GFAP (J). Scale bars: A, C, D, F–H, 100 μm; B, E, I, J, 20 μm.
Fig. 5.
Fig. 5.
Bcl-2 delays caspase activation. Western blot analyses (A–D) show that transgenic mSOD1/Bcl-2 mice that are aged-matched (AM) with end-stage (E) transgenic mSOD1 exhibit significantly lower levels of cleaved caspase-1 (A, C) and caspase-3 (B, D) than end-stage transgenic mSOD1. However, end-stage transgenic mSOD1/Bcl-2 mice exhibit levels of cleaved caspase-1 (A, C) and caspase-3 (B, D) comparable with end-stage transgenic mSOD1. A similar situation is found for the activity of caspase-1 and caspase-3 (E); *p < 0.05 higher and †p < 0.05 lower than end-stage transgenic mSOD1 and mSOD1/Bcl-2 mice, Newman–Keuls post hoc test.
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