ACE2 activator diminazene aceturate ameliorates Alzheimer's disease-like neuropathology and rescues cognitive impairment in SAMP8 mice

Figures

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  Figure 1. DIZE activated ACE2/Ang-(1-7)/MAS1 axis in the brain of SAMP8 mice. (A) The Ang- (1-7) levels in mice brain were detected by ELISA. (B) The activity of ACE2 in mice brain was assessed using a specific detection kit (#AS-72086, AnaSpec, Inc., Fremont, CA, USA) with Mc-Ala/Dnp fluorescence resonance energy transfer peptides as described. The fluorescence of Mc-Ala was monitored at excitation/emission 330 nm/390 nm. The specificity was confirmed using a specific ACE2 inhibitor DX600. (C) The Mas1 mRNA levels in mice brain were evaluated by qRT-PCR, and Gapdh was used as an internal control. (D) The protein levels of MAS1 in mice brain were detected by western blot. β-actin was used as a loading control. (E) Quantitative analysis of MAS1 protein levels. Data from panel B, C and E were expressed as a fold change relative to the vehicle-treated age-matched SAMR1 control mice. All data were analyzed by one-way ANOVA followed by Tukey’s post hoc test. Columns represent mean ± SD (n=8 per group). *P<0.05 versus age-matched vehicle-treated SAMR1 control mice. #P<0.05 versus vehicle-treated SAMP8 mice.

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  Figure 2. DIZE reduced Aβ1-42 levels in the brain of SAMP8 mice. The levels of TBS-soluble Aβ1-42 in the brain were detected by ELISA. Data were analyzed by one-way ANOVA followed by Tukey’s post hoc test. Columns represent mean ± SD (n=8 per group). *P<0.05 versus age-matched vehicle-treated SAMR1 control mice. #P<0.05 versus vehicle-treated SAMP8 mice.

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  Figure 3. DIZE ameliorated tau hyperphosphorylation in the brain of SAMP8 mice. (A) The levels of tau hyperphosphorylation at Thr205 and Ser396 sites as well as total tau in the brain were detected by western blot. β-actin was used as a loading control. (B) Quantitative analysis of tau hyperphosphorylation at Thr205/total tau ratio. (C) Quantitative analysis of tau hyperphosphorylation at Ser396/total tau ratio. Data from panel B and C were expressed as a fold change relative to the age-matched vehicle-treated SAMR1 control mice. Data were analyzed by one-way ANOVA followed by Tukey’s post hoc test. Columns represent mean ± SD (n=8 per group). *P<0.05 versus age-matched vehicle-treated SAMR1 control mice. #P<0.05 versus vehicle-treated SAMP8 mice.

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  Figure 4. DIZE attenuated neuroinflammation in the brain of SAMP8 mice. (A) The protein levels of IL-1α in the brain were investigated by ELISA. (B) The protein levels of IL-1β in the brain were investigated by ELISA. (C) The protein levels of IL-6 in the brain were investigated by ELISA. (D) The protein levels of TNF-α in the brain were investigated by ELISA. All data were analyzed by one-way ANOVA followed by Tukey’s post hoc test. Columns represent mean ± SD (n=8 per group). *P<0.05 versus age-matched vehicle-treated SAMR1 control mice. #P<0.05 versus vehicle-treated SAMP8 mice.

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  Figure 5. DIZE alleviated synaptic and neuronal losses in the brain of SAMP8 mice. (A) The protein levels of synaptophysin in the brain were detected by western blot. β-actin was used as a loading control. (B) Quantitative analysis of synaptophysin protein levels. Data were expressed as a fold change relative to the age-matched vehicle-treated SAMR1 control mice. (C) Neuronal loss in the parietal cortex of mice were detected by Nissl staining. Neurons with dark violet nucleus and intact morphology were identified as Nissl-positive neurons. Scale bar=100 μm. (D) Quantitative analysis of Nissl-positive neurons in the brain. Data were analyzed by one-way ANOVA followed by Tukey’s post hoc test. Columns represent mean ± SD (n=8 per group). *P<0.05 versus age-matched vehicle-treated SAMR1 control mice. #P<0.05 versus vehicle-treated SAMP8 mice.

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  Figure 6. DIZE rescued spatial cognitive impairment in SAMP8 mice. (A) Swimming speed of each group in the MWM test. Data were analyzed by one-way ANOVA followed by Tukey’s post hoc test. (B) Path length of each group in the hidden platform task. Data were analyzed by two-way repeated measures ANOVA followed by Bonferroni’s multiple comparisons test. Columns represent mean ± SD (n=12 per group). *P<0.05 versus age-matched vehicle-treated SAMR1 control mice. #P<0.05 versus vehicle-treated SAMP8 mice.