Prognostic significance of abnormal matrix collagen remodeling in colorectal cancer based on histologic and bioinformatics analysis

Abstract

As the major component of the tumor matrix, collagen greatly influences tumor invasion and prognosis. The present study compared the remodeling of collagen and collagenase in 56 patients with colorectal cancer (CRC) using Sirius red stain and immunohistochemistry, exploring the relationship between collagen remodeling and the prognosis of CRC. Weak or strong changes in collagen fiber arrangement in birefringence were observed. With the exception of a higher density, weak changes equated to a similar arrangement in normal collagen, while strong changes facilitated cross‑linking into bundles. Compared with normal tissues, collagen I (COL I) and III (COL III) deposition was significantly increased in CRC tissues, and was positively correlated with the metastasis status. In tissues without distant metastasis, collagen IV (COL IV) levels were higher than that in normal tissues, while in tissues with distant metastasis, collagen IV expression was significantly lower. Furthermore, the expression of matrix metalloproteinase (MMP)‑1, MMP‑2, MMP‑7, MMP‑9 and lysyl oxidase‑like 2 (LOXL2) was found to be elevated in the cancer stroma, which contributed to the hyperactive remodeling of collagen. The association between collagen‑related genes and the occurrence and prognosis of CRC were analyzed using biometric databases. The results indicated that patients with upregulated expression of a combination of coding genes for collagen and collagenase exhibited poorer overall survival times. The coding genes COL1A1‑2, COL3A1, COL4A3, COL4A6 and MMP2 may therefore be used as biomarkers to predict the prognosis of patients with CRC. Furthermore, the results of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis suggest that collagen may promote tumor development by activating platelets. Collectively, the abnormal collagen remodeling, including associated protein and coding genes is associated with the tumorigenesis and metastasis, affecting the prognosis of patients with CRC.

Keywords: collagen; matrix metalloproteinase; lysyl oxidase-like 2; Oncomine; TCGA; GEO.

Figure 1.

Changes in collagen fiber arrangement differ in different cases of CRC. Differences are indicted by black arrow. (A) Representative images of collagen fiber arrangement characteristics. (B) Comparison of the remodeling intensity of collagen arrangement in metastatic and non-metastatic CRC. Magnification, ×400; Scale bar, 100 µm. CRC, colorectal cancer.

Figure 2.

Comparison of collagen expression in different tissue samples. (A) Images and expression levels of COL I, COL III, and COL IV in different tissue samples. (B-I) Differences in the expression of (B) COL I, (C) COL I area, (D) COL III, (E) COL III area, (F) the ratio of COL I/COL III, (G) the ratio of COL I area/COL III area, and (H and I) COL IV. Magnification, ×200. Scale bar, 50 µm. COL, collagen; AOD, average optical density; CRC, colorectal cancer. *P<0.05; **P<0.01; ^nsP>0.05.

Figure 3.

Comparison of collagenase expression levels in different tissue samples. (A) Images of MMP-1, MMP-2, MMP-7, MMP-9 and LOXL2 in different tissue samples. (B-F) The expression levels of (B) MMP-1, (C) MMP-2, (D) MMP-7, (E) MMP-9 and (F) LOXL2 in different tissue samples. (G) Interactions between LOXL2 and MMP-1, MMP-2, MMP-7 and MMP-9. (H-J) Correlation analysis between the expression of (H) MMP-7 and MMP-1, (I) MMP-7 and MMP-2 and (J) MMP-7 and MMP-9. (K-N) Correlation analysis between the expression of (K) LOXL2 and MMP-1, (L) LOXL2 and MMP-2, (M) LOXL2 and MMP-7 and (N) LOXL2 and MMP-9. Magnification, ×200; Scale bar, 50 µm. MMP, matrix metalloproteinase; LOXL2, lysyl oxidase-like 2; AOD, average optical density; CRC, colorectal cancer. **P<0.01.

Figure 4.

mRNA expression levels of collagens and collagenases in various types of cancer. (A) High or low expression of related coding genes in different types of human cancer tissues, compared with normal tissues, using the Oncomine database (P<0.01). mRNA expression levels in CRC are highlighted in yellow. (B and C) Expression patterns of related coding genes in CRC using TCGA database (P<0.01). (D) Intersection between all related coding genes (n=14) among Oncomine and TCGA expression data. CRC, colorectal cancer; TCGA, The Cancer Genome Atlas.

Figure 5.

Functional enrichment analysis of coding genes associated with the prognosis of patients with CRC. Functional enrichment analysis including cellular component, molecular function, biological process, and KEGG analysis of 14 coding genes associated with collagen and collagenases. CRC, colorectal cancer; KEGG, Kyoto Encyclopedia of Genes and Genomes.

Figure 6.

Prognostic analysis of collagen-related coding genes in CRC. (A) OS analysis of combined expression of genes including COL1A1-2, COL3A1, COL4A1-6, LOXL2, MMP1, MMP2, MMP7 and MMP9 using PROGgenev2. (B) Time-dependent ROC analysis of the sensitivity and specificity of the six-mRNA signature (COL1A1-2, COL3A1, COL4A3, COL4A6 and MMP2). (C) Kaplan-Meier analysis of the six-mRNA signature. (D) ROC curve of the prognostic model. (E) Heatmap of the expression of the six-mRNA signature. CRC, colorectal cancer; OS, overall survival; ROC, receiver operating characteristic; AUC, area under the curve.

Figure 7.

Kaplan-Meier analysis of the association between COL1A1, COL1A2, COL3A1, COL4A3, COL4A6 and MMP2 and the OS times of patients with CRC. (A) COL1A1, (B) COL1A2, (C) COL3A1, (D) COL4A3, (E) COL4A6 and (F) MMP2. OS, overall survival; CRC, colorectal cancer.