Caspase-6 Knockout in the 5xFAD Model of Alzheimer's Disease Reveals Favorable Outcome on Memory and Neurological Hallmarks

Abstract

Alzheimer's disease (AD) is a progressive neurodegenerative disorder and is the most common form of dementia in the elderly. Caspases, a family of cysteine proteases, are major mediators of apoptosis and inflammation. Caspase-6 is considered to be an up-stream modulator of AD pathogenesis as active caspase-6 is abundant in neuropil threads, neuritic plaques, and neurofibrillary tangles of AD brains. In order to further elucidate the role of caspase-6 activity in the pathogenesis of AD, we produced a double transgenic mouse model, combining the 5xFAD mouse model of AD with caspase-6 knock out (C6-KO) mice. Behavioral examinations of 5xFAD/C6-KO double transgenic mice showed improved performance in spatial learning, memory, and anxiety/risk assessment behavior, as compared to 5xFAD mice. Hippocampal mRNA expression analyses showed significantly reduced levels of inflammatory mediator TNF-α, while the anti-inflammatory cytokine IL-10 was increased in 5xFAD/C6-KO mice. A significant reduction in amyloid-β plaques could be observed and immunohistochemistry analyses showed reduced levels of activated microglia and astrocytes in 5xFAD/C6-KO, compared to 5xFAD mice. Together, these results indicate a substantial role for caspase-6 in the pathology of the 5xFAD model of AD and suggest further validation of caspase-6 as a potential therapeutic target for AD.

Keywords: Alzheimer’s disease; caspase-6; knock-out mice; neurodegeneration.

Figure 1

Behavioral amelioration in 5xFAD/C6-KO female mice in the Y-maze and elevated plus maze.

Figure 2

Caspase-6 KO in 5xFAD mice improves cognitive performance in the Morris water maze test.

Figure 3

Caspase-6 KO in 5xFAD mice improves searching pattern in the probe test.

Figure 4

Synaptophysin is increased in 5xFAD/C6-KO double transgenic mice. Syn: Synaptophysin.

Figure 5

Altered gene expression in hippocampus of 5xFAD/C6-KO mice. Mice hippocampi were taken for RT-PCR analysis of several genes: mRNA quantification for female and male TNF-α (A,B), IL-10 (C,D); and Caspase-3 (Casp3; E,F) respectively. Data are mean ± SEM. *p < 0.05, **p < 0.01, and ****p < 0.0001. Two-tailed t-test between marked groups. NS: Not significant.

Figure 6

Caspase-6 KO in 5xFAD mice reduces amyloid-β plaque load. Representative images of hippocampal slices stained with ThioS for male (A–B) and female (C–D) transgenic mice and their respective quantification (n = 5–7). Scale bar = 200μm. Data are mean ± SEM. **p< 0.01 and ***p < 0.001. Two-tailed t-test.

Figure 7

Low IBA1 and GFAP expression in 5xFAD caspase-6 KO mice. Representative images of hippocampal slices stained with GFAP (A–D) or IBA1 (E–H). Quantification of GFAP and IBA1 are mean intensity of n = Scale bar = 25 μm. Data are mean ±SEM. *p < 0.05, **p < 0.01, and ***p < 0.001. Two-tailed t-test.