Identification of core genes in prefrontal cortex and hippocampus of Alzheimer's disease based on mRNA-miRNA network

Abstract

Alzheimer's disease (AD) is a neurodegenerative disorder with cognitive impairment and abnormal mental behaviour. There is currently no effective cure. The development of early diagnostic markers and the mining of potential therapeutic targets are one of the important strategies. This study aimed to explore potential biomarkers or therapeutic targets related to AD in the hippocampus and prefrontal cortex, two brain regions highly related to AD. Differentially expressed genes and miRNAs between AD patients and healthy controls were obtained from the Gene Expression Omnibus database. The mRNA-miRNA network was constructed and key genes involved in AD were screened out by protein-protein interaction analysis, and were subsequently verified by independent datasets and qPCR in an AD mouse model. Our findings showed that six hub genes including CALN1, TRPM7, ATR, SOCS3, MOB3A and OGDH were believed to be involved in the pathogenesis of AD. Western blot analysis further determined that CALN1, ATR and OGDH were the possible biomarkers and therapeutic targets for AD. In addition, 6 possible miRNAs biomarkers have also been verified by qPCR on AD animal models. Our findings may benefit clinical diagnosis and early prevention of AD.

Keywords: Alzheimer's disease; bioinformatics; hub genes; mRNA; miRNA.

FIGURE 1

Identification of DEGs and DEMs. (A and C) DEGs in prefrontal cortex (A) and hippocampus (C). (B and D) The DEMs in prefrontal cortex (B) and hippocampus (D). DEGs or DEMs with |log2 Fold Change| ≥ 0.2 (A) or 1 (B–D) and p value < 0.05 were considered as significantly different. Red and green represent up‐regulated and down‐regulated expression, respectively

FIGURE 2

Analysis of the target genes of DEMs. (A and B) Network of the 4 up‐regulated miRNAs (A) and the 2 down‐regulated miRNAs (B) with their target genes in prefrontal cortex. (C and D) Network of the top 10 up‐regulated miRNAs (C) or the top 10 down‐regulated miRNAs (D) and their target genes

FIGURE 3

The functional enrichment analysis of DEGs. (A–C and E–G) GO enrichments analysis of DEGs at three aspects of biology: BP (A, E), MF (B, F) and CC (C, G) in prefrontal cortex and hippocampus as indicated in graph. (D and H) KEGG pathways enrichment analysis of DEGs in prefrontal cortex (D) and hippocampus (H). The size of the dot represents the number of genes, and the red represents the smaller p‐value, the blue represents the larger p‐value

FIGURE 4

The functional enrichment analysis of DEM target genes. (A–C and E–G) GO enrichments analysis of DEMs target genes at three aspects of biology: BP (A, E), MF (B, F) and CC (C, G) in prefrontal cortex and plasma as indicated in graph. (D and H) KEGG pathways enrichment analysis of DEM target genes in prefrontal cortex (D) and plasma (H). The size of the dot represents the number of genes, and the red represents the smaller p‐value, the blue represents the larger p‐value.

FIGURE 5

The mRNAs and miRNAs network. (A, B, F and G) The common genes between up‐regulated DEGs and down‐regulated DEM target genes (A and F) or between down‐regulated DEGs and up‐regulated DEM target genes (B and G) in prefrontal cortex and hippocampus as indicated in graph. (C, D, H and I) The network between up‐regulated common genes and their corresponding down‐regulated DEMs (C and H) or between down‐regulated common genes and their corresponding up‐regulated DEMs (D and I). (E and J) Hub genes in prefrontal cortI(E) and hippocampus (J) were identified by Cytohubba plug‐ins in Cytoscape. And the darker the nodes, the higher the score

FIGURE 6

Verify the expression of hub genes in normal controls and diseased samples. The boxplots were used to show the expression level of these genes. (A) CALN1, (B) GAN13, (C) FLCN, (D) MOB3A, (E) TRPM7, (F) ATR, (G) DDB1, (H) SOCS3, (I) PTPN1, (J) OGDH, (K) EID2. (L) qPCR analysis in an APP/PS1 model mice of AD. Data are represented as mean ± SD. Student t‐test was used for two group comparisons. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001

FIGURE 7

Verify the protein level of hub genes and potential miRNAs level in APP/PS1 mice. (A and B) Western blot analysis of candidate molecules involved in AD. (A) representative images of western blot. (B) the statistical analysis. (C) the qPCR analysis of key miRNAs. Data are represented as mean ± SD. Student t‐test was used for two group comparisons. *p < 0.05, **p < 0.01, ***p < 0.001.