Evaluation of a five-gene signature associated with stromal infiltration for diffuse large B-cell lymphoma

Figure 1 The top 20 differentially expressed genes in diffuse large B-cell lymphoma. A: A total of 31 cases in the normal lymphocytes group and 30 cases in the diffuse large B-cell lymphoma group were subjected to analysis of the differentially expressed genes; B: The principal components analysis plot for 31 normal lymphocyte samples and 30 diffuse large B-cell lymphoma samples; C: The Spearman correlation matrix analysis of all samples was conducted by pairwise correlation coefficient analysis. A2M: Alpha-2-macroglobulin; CTSB: Cathepsin B; FN1: Fibronectin; MMP9: Matrix metallopeptidase 9; PC: Principal components; SPARC: Secreted protein acidic and cysteine-rich.

Figure 2 The five hub genes were confirmed as overexpressed in diffuse large B-cell lymphoma. A: The protein-protein interaction was confirmed in STRING, and the hub genes were obtained; B-F: The expressions of alpha-2-macroglobulin (A2M) (B), fibronectin (FN1) (C), secreted protein acidic and cysteine-rich (SPARC) (D), cathepsin B (CTSB) (E), and matrix metallopeptidase 9 (MMP9) (F) were subjected to GEPIA to analyze the expression in diffuse large B-cell lymphoma and normal lymphocytes within The Cancer Gene Atlas dataset. ^aP < 0.001 vs the diffuse large B-cell lymphoma tumor samples.

Figure 3 The alteration of hub genes had a potential association with the development of diffuse large B-cell lymphoma. A: The gene alterations were conducted in the cBioportal and included missense mutation, amplification, deep deletion, and messenger ribonucleic acid high; B-F: The expressions of alpha-2-macroglobulin (A2M) (B), cathepsin B (CTSB) (C), fibronectin (FN1) (D), matrix metallopeptidase 9 (MMP9) (E), secreted protein acidic and cysteine-rich (SPARC) (F) were subjected to GEPIA to analyze the gene expression in different stages of DLBCL. ^bP < 0.05 vs stage 1 group.

Figure 4 Overexpression of five hub genes was confirmed in diffuse large B-cell lymphoma. A-E: Correlation analyses between alpha-2-macroglobulin (A2M) (A), cathepsin B (CTSB) (B), fibronectin (FN1) (C), matrix metallopeptidase 9 (MMP9) (D), secreted protein acidic and cysteine-rich (SPARC) (E) expression, and tumor purity were conducted by the Estimation of Stromal and Immune cells in Malignant Tumor tissues using Expression data Score (ESTIMATE) method; F: Expression correlation analysis between CD19 and the five hub genes was conducted in GEPIA. P < 0.05 was considered as statistically different.

Figure 5 Stromal infiltration analysis of the five hub genes. A-E: Alpha-2-macroglobulin (A2M) (A), cathepsin B (CTSB) (B), fibronectin (FN1) (C), matrix metallopeptidase 9 (MMP9) (D), and secreted protein acidic and cysteine-rich (SPARC) (E) expression levels were confirmed by correlation analysis with the stromal score. P < 0.05 was considered as statistically different.

Figure 6 The five hub genes were closely associated with infiltration level of cancer-associated fibroblasts with the MCP-Counter method. A-E: The alpha-2-macroglobulin (A2M) (A), cathepsin B (CTSB) (B), fibronectin (FN1) (C), matrix metallopeptidase 9 (MMP9) (D), and secreted protein acidic and cysteine-rich (SPARC) (E) gene expressions were evaluated for correlation with infiltration levels of cancer-associated fibroblasts (CAFs) in diffuse large B-cell lymphoma, and the correlation analysis was conducted with the Spearman method. P < 0.05 was considered as statistically different.

Figure 7 Validation of the positive correlation between the five genes and the infiltration level of cancer-associated fibroblasts. A-E: Alpha-2-macroglobulin (A2M) (A), cathepsin B (CTSB) (B), fibronectin (FN1) (C), matrix metallopeptidase 9 (MMP9) (D), and secreted protein acidic and cysteine-rich (SPARC) (E) were subjected to analysis for correlation with the infiltration level of cancer-associated fibroblasts (CAFs) by the tumor immune dysfunction and exclusion (TIDE) portal. P < 0.05 was considered as statistically different.

Figure 8 The top two genes, fibronectin and secreted protein acidic and cysteine-rich, were expressed in a remarkably correlated manner. A: A total of 48 diffuse large B-cell lymphoma samples were applied to TIMER to determine the expression correlation between fibronectin (FN1) and secreted protein acidic and cysteine-rich (SPARC); B: The FN1 protein levels were positively correlated with the SPARC messenger ribonucleic acid levels. The correlation analysis was conducted by the Spearman method, and P < 0.05 was considered as statistically different.