Identification of TLR2 as a Key Target in Neuroinflammation in Vascular Dementia

Abstract

Vascular dementia (VaD) is the second most common cause of dementia. At present, precise molecular processes of VaD are unclear. We attempted to discover the VaD relevant candidate genes, enrichment biological processes and pathways, key targets, and the underlying mechanism by microarray bioinformatic analysis. We selected GSE122063 related to the autopsy samples of VaD for analysis. We first took use of Weighted Gene Co-expression Network Analysis (WGCNA) to achieve modules related to VaD and hub genes. Second, we filtered out significant differentially expressed genes (DEGs). Third, significant DEGs then went through Geno Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. Fourth, Gene Set Enrichment Analysis (GSEA) was performed. At last, we constructed the protein-protein interaction (PPI) network. The results showed that the yellow module had the strongest correlation with VaD, and we finally identified 21 hub genes. Toll-like receptor 2 (TLR2) was the top hub gene and was strongly correlated with other possible candidate genes. In total, 456 significant DEGs were filtered out and these genes were found to be enriched in the Toll receptor signaling pathway and several other immune-related pathways. In addition, Gene Set Enrichment Analysis results showed that similar pathways were significantly over-represented in TLR2-high samples. In the PPI network, TLR2 was still an important node with high weight and combined scores. We concluded that the TLR2 acts as a key target in neuroinflammation which may participate in the pathophysiological process of VaD.

Keywords: TLR2; WGCNA; bioinformatic analysis; neuroinflammation; vascular dementia.

FIGURE 1

Results and visualization of Weighted Gene Co-expression Network Analysis (WGCNA) analysis. (A) Determination of soft threshold β. Left: scale independence; right: mean connectivity. (B) Heat map showing the TOM among all 5,000 genes involved in the WGCNA with cluster dendrogram showing on the axis. Each color represents one specific co-expression module; the above branches represent genes. The genes with strong correlations are clustered into modules, which are represented as dark sections symmetrically distributed along the diagonal in the heatmap, corresponding to the cluster dendrogram. (C) Module–trait relationships among the five gene modules. The yellow module is the most correlated module (correlation coefficient = 0.57, ***p < 0.001).

FIGURE 2

Selection of hub genes. Module membership (MM) vs. gene significance (GS) in the yellow module (correlation coefficient = 0.65, ***p < 0.001). The red dotted lines represented the thresholds of MM > 0.9 and GS > 0.3 set for hub genes and separated an area in the upper right corner. Toll-like receptor 2 (TLR2) is selected as the top hub gene.

FIGURE 3

Differentially expressed genes (DEGs) present in vascular dementia (VaD) and control groups in microarray from GSE122063 and the expression level of Toll-like receptor-2 (TLR2). (A) Volcano plot showed the distribution of the DEGs between two groups. The red dots correspond to the significantly regulated genes. (B) Violin plot of TLR2. TLR2 is upregulated in the VaD group (***p < 0.001).

FIGURE 4

Results of the Geno Ontology (GO) terms enrichment analysis of significant DEGs. (*p < 0.05). Blue bars showed the results of upregulated genes while red bars showed the results of downregulated genes.

FIGURE 5

Gene Set Enrichment Analysis (GSEA) results grouped by the expression level of TLR2. (A) BP enriched in TLR2-high group. (B) CC enriched in TLR2-high group. (C) MF enriched in TLR2-high group. (D) KEGG pathways enriched in TLR2-high group.

FIGURE 6

Construction of the protein–protein interaction (PPI) network consisting of DEGs. (A) PPI of DEGs. (B) Partial network centered on TLR2. The size and color of the nodes reflect the degree and the width and color of the edges reflect the combined scores (color: from blue to red). Larger size and bluer nodes indicated the higher degree while wider and bluer lines indicated the higher combined scores.

FIGURE 7

Potential mechanism for high expression of TLR2 to promote VaD. The network is summarized according to GSE122063 database and public KEGG pathway. Red indicates the upregulated genes.