Identification of CXCL10 and CXCL11 as the candidate genes involving the development of colitis-associated colorectal cancer

Abstract

Background: Ulcerative colitis (UC) is a well-known risk factor for developing colitis-associated colorectal cancer (CAC). However, the molecular mechanism of the pathogenesis of CAC remains unclear. This study aimed to explore candidate genes involved in the tumorigenesis of CAC. Methods: GSE75214 and the Cancer Genome Atlas Program (TCGA) dataset were used to analyze the differentially expressed genes (DEGs) in UC and colorectal cancer (CRC), respectively. Survival-hub genes were identified from these DEGs by sequentially constructing a protein-protein interaction network, selecting hub genes, and conducting survival analysis. Regulatory signatures were also predicted on these genes through the online database. Apc ^min/+ and UC mice models were used to validate the expression of the above-predicted molecules. Gene set enrichment analysis and CIBERSORT were performed to explore the enriched molecular pathways and associated tissue-infiltrating immune cells of genes. Results: Here, 376 common DEGs were identified from the GSE75214 and TCGA datasets. Through survival-hub gene selection and in vivo experiments, we confirmed that CXCL10 and CXCL11 were significantly upregulated in UC and CRC. We also proved that miR-34a-5p and miR-203a-5p were potential regulators of CXCL10 and CXCL11. Meanwhile, CXCL10 and CXCL11 may activate the JAK-STAT signaling pathway via the interaction with cytokine receptors in UC. Furthermore, CXCL10 and CXCL11 were positively associated with the tissue infiltration of proinflammatory M1 macrophages in UC and CRC. Conclusion: CXCL10 and CXCL11 may act as the candidate genes involved in the tumorigenesis of CAC and potential therapeutic targets to prevent the development of CAC from UC.

Keywords: TCGA; colitis-associated colorectal cancer; colorectal cancer; differentially expressed genes; ulcerative colitis.

FIGURE 1

Workflow of processing the datasets. Abbreviation: UC, ulcerative colitis; TCGA, The Cancer Genome Atlas; CRC, colorectal cancer; TFs, transcription factors; miRNA, microRNA.

FIGURE 2

The common differentially expressed genes (DEGs) between ulcerative colitis (UC) and colorectal cancer (CRC). (A) heatmap of the top 100 DEGs in the UC dataset. (B) heatmap of top 100 DEGs in the CRC dataset. (C) common upregulated DEGs in UC and CRC. (D) common downregulated DEGs in UC and CRC. Abbreviation: DEGs, Differentially expressed genes; UC, ulcerative colitis; CRC, colorectal cancer.

FIGURE 3

Functional enrichment analysis and protein–protein interaction (PPI) network of common differentially expressed genes (DEGs). (A) gene ontology enrichment analysis of common DEGs. (B) Kyoto Encyclopedia of Genes and Genomes enrichment analysis of common DEGs. (C) PPI network of common DEGs. The red node represented upregulated genes; the green node stands for downregulated genes. (D) hub genes identified from the PPI network. Abbreviation: DEGs, differentially expressed genes; BP, biological process; CC, cellular components; MF, molecular function; PPI, protein–protein interaction.

FIGURE 4

Clinical significance of hub genes in the Cancer Genome Atlas Program cohorts. (A) expression analysis of hub genes in different tumor stages. (B) overall survival analysis of CXCL11. (C–E) progression-free interval analysis of CXCL10, CXCL11, and IL1A, respectively.

FIGURE 5

Regulatory signatures of survival-associated hub genes. (A) miRNA–gene interaction network; (B) transcription factors–gene interaction network. The red node represented survival-related hub genes. The green node indicated miRNAs and transcription factors.

FIGURE 6

The expression of survival-hub genes and interacted regulatory signatures in vivo. (A) cytokine array to detect CXCL10 and CXCL11 in ulcerative colitis (UC) mice and controls (n = 3, per group), respectively. (B) cytokine array to detect CXCL10 and CXCL11 in APC ^min/+ mice and wild type (WT) (n = 3, per group), respectively. (C) cytokine array to detect IL1A in UC mice and controls (n = 3, per group). (D) cytokine array to detect IL1A in APC ^min/+ mice and WT (n = 3, per group). (E) qRT-PCR for miR-34a-5p and miR-203a-3p in UC mice and controls (n = 6, per group), respectively. (F) qRT-PCR for miR-34a-5p and miR-203a-3p in APC ^min/+ mice and WT (n = 6, per group), respectively. (G) qRT-PCR for miR-210-3p in UC mice and controls (n = 6, per group). (H) qRT-PCR for miR-210-3p in APC ^min/+ mice and WT (n = 6, per group). (I) Western blot for YY1 expression in different groups. Ctrl, control mice for UC model. WT, wild type mice. *, p < 0.05; **, p < 0.01, ***, p < 0.001. Abbreviation: Ctrl, control; UC, ulcerative colitis; WT, wild type.

FIGURE 7

Immune infiltration and gene set enrichment analysis (GSEA) of survival-hub genes. (A) correlation analysis between CXCL10 and immune cell infiltration in the Cancer Genome Atlas Program (TCGA) and GSE75214 datasets, respectively. (B) correlation analysis between CXCL11 and immune cell infiltration in the TCGA and GSE75214 datasets, respectively. (C) GSEA of CXCL10 and CXCL11 in TCGA cohorts. (D) GSEA of CXCL10 and CXCL11 in the GSE75214 dataset. Abbreviation: CRC, colorectal cancer; UC, ulcerative colitis; KEGG, Kyoto Encyclopedia of Genes and Genomes; NK, natural killer.