Table 1.
Gene names, accession numbers, primer sequences, and annealing temperatures used in qRT-PCR.
Fig 1.
Datasets selection procedure for bioinformatic analysis.
Totally, two datasets out of the 741 records identified from the gene expression omnibus (GEO) database meet the selection criteria.
Fig 2.
Identifying DEGs between oxaliplatin-resistant and oxaliplatin-sensitive HT-29 cells.
Diagrams A and B display the volcano plots related to DEGs for dataset GS4E2387 (A) and GSE76092 (B). The red and blue points represent up- and down-regulated genes, respectively, based on│Log2 FC│≥ 1.5 and adjusted P-value < 0.05. Diagrams C and D demonstrate the Venn diagrams of the 18 up- and 36 down-regulated overlapping DEGs between two data series, respectively.
Fig 3.
Heatmap related to the log2 FC values of common DEGs between GSE42387 and GSE76092.
Each column and row represents a root sample and a gene, respectively. Log2 FC values are shown in each pixel. Red and blue pixels indicate log2 FC ≥ 1.5 and log2 FC ≤ -1.5, respectively. FC: Fold change; DEG: Differentially expressed gene.
Fig 4.
Hierarchical clustering heatmap related to overlapping DEGs.
Heatmap related to the expression profile of 57 common DEGs between three resistant and three sensitive HT-29 cells from the microarray (A) GSE42387 and (B) GSE76092 datasets. Red and blue pixels indicate the over- and under-expression of NNMT, SLC14A1, and TFF2 genes in oxaliplatin-resistant cells, respectively. The above-mentioned genes exhibit reverse expression pattern in two datasets to down-regulate in GSE42387 and up-regulate in GSE76092. DEG: Differentially expressed gene.
Fig 5.
PPI networks of common hub genes in GSE42387 and GSE76092 datasets constructed using the cytoHubba plug-in in Cytoscape based on IMEx database.
(A) Up-regulated and (B) down-regulated common DEGs. The nodes and edges represent genes and their interaction, respectively. The red and blue nodes represent the up- and down-regulated DEGs, respectively. The node size and color indicate the degree value. DEG: Differentially expressed gene.
Fig 6.
Venn diagrams of overlapping hub genes between GSE42387 and GSE76092.
Totally, 25 overlapping hub genes are identified including (A) 12 down-regulated and (B) 13 up-regulated genes. DEG: Differentially expressed gene.
Table 2.
The overlapping hub genes in the PPI networks of down-regulated differentially expressed genes (DEGs) from GSE76092 and GSE42387.
Table 3.
The overlapping hub genes in the PPI networks of up-regulated differentially expressed genes (DEGs) from GSE76092 and GSE42387.
Table 4.
The top 10 uncommon hub genes in the PPI networks of up- and down-regulated differentially expressed genes (DEGs) from GSE42387.
Table 5.
The top 10 uncommon hub genes in the PPI networks of up and down-regulated differentially expressed genes (DEGs) from GSE76092.
Table 6.
Modules identified from the PPI networks of down- and up-regulated differentially expressed genes (DEGs) from GSE42387 and GSE76092 by MCOD.
Table 7.
Functional enrichment analysis of common up-regulated differentially expressed genes (DEGs) in GSE76092 and GSE42387.
Table 8.
Functional enrichment analysis of common down-regulated differentially expressed genes (DEGs) in GSE76092 and GSE42387.
Table 9.
Pathway analysis of common down-regulated differentially expressed genes (DEGs) in GSE76092 and GSE42387.
Fig 7.
Gene Ontology and pathway analysis of GSE42387 DEGs.
The top 15 GO enrichment terms and pathways are shown. (A) Biological process, (B) molecular function, (C) cellular component, and (D) KEGG pathway of up-regulated (red column) and down-regulated (blue column) DEGs. KEGG: Kyoto Encyclopedia of Genes and Genomes; DEGs: Differentially expressed genes. P value is shown in S2 Table.
Fig 8.
Gene Ontology and pathway analysis of GSE76092 DEGs.
The top 15 GO enrichment terms and pathways are shown. (A) Biological process, (B) molecular function, (C) cellular component, and (D) KEGG pathway of up-regulated (red column) and down-regulated (blue column) DEGs. KEGG: Kyoto Encyclopedia of Genes and Genomes; DEGs: Differentially expressed genes. P value is indicated in S2 Table.
Fig 9.
Characterizing oxaliplatin-resistant HCT116 cells.
A) Dose-response curves of cell viability in the parental HCT116 cells, HCT116/OX-R4.3, and HCT116/OX-R10 cells treated with various concentrations of oxaliplatin. B) The survival fraction of cells treated with 0.1–60 μM oxaliplatin. C) Representative phase contrast and fluorescence images related to acridine orange- and SA-β-gal—stained parental HCT116, HCT116/OX-R4.3, and HCT116/OX-R10 cells (×20 magnification, Scale bar: 50 μm). Yellow arrows and red circle indicate enlarged multinucleated cells. D) Quantifying the relative percentage of SA-β-gal-positive cells. E) Cell cycle analysis of the parental and resistant HCT116 cells by flow cytometry. Data indicate that the means ± SD of three independent experiment and statistical analysis are performed by one-way ANOVA test. *p < 0.05 and **p < 0.01. HCT116/OX-R: Oxaliplatin-resistant HCT116 cells; SA-β-gal: Senescence-associated beta-galactosidase; OX: Oxaliplatin.
Fig 10.
Validating in silico analysis by quantitative real-time PCR.
Relative mRNA expression levels related to six selected hub genes (USH1C, TGM2, HMGA2, FXYD3, LGALS4, and ECI2) in the parental HCT116 cells and two oxaliplatin-resistant sub-lines, HCT116/OX-R4.3 and HCT116/OX-R10 cells. Data are represented as means ± standard error mean (SEM). Statistical analysis is performed by the student’s t-test compared to the control group; *p < 0.05, and ***p < 0.001. HCT116/OX-R: Oxaliplatin-resistant HCT116 cells; USH1C: Usher syndrome 1C; TGM2: Transglutaminase 2; HMGA2: High mobility group A2 protein; FXYD3: FXYD domain-containing ion transport regulator 3; LGALS4: Galectin-4; ECI2: Enoyl-coA delta isomerase 2.