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. 2024 Aug 21:15:1440515.
doi: 10.3389/fphar.2024.1440515. eCollection 2024.

Transcriptome analysis of the aged SAMP8 mouse model of Alzheimer's disease reveals novel molecular targets of formononetin protection

Bo Liu  1 , Di Cui  1 , Jie Liu  1 , Jing-Shan Shi  1
Affiliations

Transcriptome analysis of the aged SAMP8 mouse model of Alzheimer's disease reveals novel molecular targets of formononetin protection

Bo Liu et al. Front Pharmacol. .

Abstract

Background: Senescence-accelerated mouse prone 8 (SAMP8) and age-matched SAMR1 mice are used to study the pathogenesis and therapeutics of Alzheimer's disease (AD); however, the molecular mechanisms are not completely understood.

Objective: This study aimed to examine the effects of the 5-month administration of formononetin in SAMP8 mice and used RNA-seq to explore the molecular targets.

Methods: SAMP8 mice were orally administered formononetin (0, 8, and 16 mg/kg) from 4 months of age, and age-matched SAMR1 mice were used as controls. Behavioral tests were performed in 9-month-old mice, followed by histopathologic analysis. Total RNA from the hippocampus was isolated and subjected to RNA-seq, RT-qPCR, and bioinformatics analysis.

Results: The 9-month-old SAMP8 mice exhibited cognition deficits, evidenced by novel object recognition, open-field test, elevated plus maze, and passive avoidance. Nissl bodies in the cortex and hippocampus were decreased. Formononetin treatments ameliorated behavioral deficits and improved morphological changes, which were evidenced by Nissl and H&E staining. RNA-seq revealed distinct gene expression patterns between SAMP8 and SAMR1 mice. Differentially expressed genes in SAMP8 mice were attenuated or normalized by formononetin. Ingenuity pathway analysis (IPA) of canonical pathway and upstream regulators revealed increases in proinflammatory factors and immune dysfunction and decreases in NRF2 and SIRT-1 signaling pathways, leading to neuroinflammation. Formononetin treatment attenuated or reversed these molecular changes. The transcriptome of SAMP8 mice was correlated with transcriptomic profiles of other AD mouse models in the GEO database.

Conclusion: Neuroinflammation and decreased antioxidant and SIRT-1 signaling contributed to cognitive deficits in aged SAMP8 mice, which are potential therapeutic targets of formononetin in combination with other therapies.

Keywords: Alzheimer’s disease; RNA-seq analysis; aged SAMP8 mouse; formononetin; ingenuity pathway analysis.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The authors declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision.

Figures

FIGURE 1
FIGURE 1
Effects of FMN on emotion and learning and memory in SAMP8 mice. (A) The activity time spent by mice in exploring the novel object. (B) The activity time spent by mice in the central region. (C) The activity time spent by mice in open arms. (D) The activity time spent by mice in the light box. Data are expressed as mean ± SD of n = 10, #p < 0.05 vs. SAMR1 mice, *p < 0.05 vs. SAMP8 mice.
FIGURE 2
FIGURE 2
Effects of FMN on neuron morphology change in SAMP8 mice. (A) Representative images showing Nissl bodies in CA1 and DG regions of the hippocampus and cerebral cortex (magnification 200×, scale bar = 50 μm); (B) quantitation of Nissl-stained neurons in the normal form in the CA1 hippocampal region. (C) Representative images showing Nissl bodies in the DG hippocampal region. (D) Quantitation of Nissl-stained neurons in the normal form in the cerebral cortex. The numbers of Nissl bodies were captured in the three fields of the hippocampus CA1 and DG regions as well as in the cerebral cortex. ( x¯ ± SD, n = 3), # p < 0.05 vs. SAMR1, * p < 0.05 vs. SAMP8 vehicle.
FIGURE 3
FIGURE 3
Differentially expressed genes. (A) Principle component analysis (PCA). (B) Differentially expressed genes (DEGs). (C) 2-D cluster heatmap of DEGs as compared to SAMR1 mice.
FIGURE 4
FIGURE 4
RT-qPCR analysis of 12 selected DEGs. (A) Upregulated genes in SAMP8 mice. (B) Downregulated genes. (C) Proinflammatory genes. Data are expressed as mean ± SD (n = 6). #Significantly different from SAMR1, p < 0.05; *Significantly different from SAMP8, p < 0.05.
FIGURE 5
FIGURE 5
Ingenuity Pathway Analysis (IPA). (A) Selected canonical pathways. (B) Selected upstream regulators based on the Z-score. Red indicates upregulation, and blue indicates downregulation, and blank indicates no change, as compared to the SAMR1 group. (C) Graphical summary of SAMP8_vs._SAMR1 key gene interactions. (D) Graphical summary of SAMP8+FMN-H_vs._SAMR1 key gene interactions.
FIGURE 6
FIGURE 6
Correlation with the GEO database analysis. The DEGs were imported into the Illumina Correlation Engine to compare the gene expression biosets in the GEO database using –log (p-value), with upregulation shown in red and downregulation shown in blue. The correlations were in the format of the GSE number. Study name in the GEO database.
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Grants and funding

The authors declare that financial support was received for the research, authorship, and/or publication of this article. This work was supported by the J-SS Tutor Studio of Pharmacology (GZS-2016(07)); Guizhou Science and Technology Department ZK (2022-596), ZK (2022-598), and ZK (2023-575); Zunyi Science and Technology Cooperation HZ (2022-245) and HZ (2020-69); and Plan Project of Science and Technology in Guizhou Province (No. Qian Ke He (2020) 040).

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