Fibrotic Phenotype of Peritumour Mesenteric Adipose Tissue in Human Colon Cancer: A Potential Hallmark of Metastatic Properties

Abstract

The impact of tumour associated stroma on cancer metastasis is an emerging field. However, cancer associated genes in peritumoral adipose tissue (pAT) in human colon cancer have not been explored. The aim of this study was to identify differentially expressed genes (DEGs) associated with cancer pathways in mesenteric pAT compared with adjacent adipose tissue. In total, nine patients with colon cancer pathological stage T2/T4 were employed in this study. DEGs were identified in 6 patients employing Nanostring PanCancer Pathway Panel and pathway enrichment analyses were performed. Differential expression of the 5 most up-regulated and 2 down regulated genes was validated with qRT-PCR. Results showed collagen type I alpha 1 chain (COL1A1) p = 0.007; secreted frizzled related protein (SFRP2) p = 0.057; fibroblast growth factor 7 (FGF7) not significant (ns); phospholipase A2, group IIA (PLA2G2A) ns; nerve growth factor receptor (NGFR) ns; lymphoid enhancer binding factor 1 (LEF1) p = 0.03; cadherin 1, Type 1, E-cadherin (epithelial) (CDH1) 0.09. Results have highlighted down-regulation of the Wingless/Integrated (Wnt) pathway in mesenteric pAT compared to distal adipose tissue. Highly upregulated genes in mesenteric pAT were involved in extracellular matrix (ECM)-receptor interactions and focal adhesion. Highly down regulated genes were involved in the cell cycle. Immunohistochemistry revealed differential distribution of COL1A1 showing maximum levels in tumour tissue and gradually decreasing in distant adipose tissue. COL1A1 and down regulation of Wnt pathway may have a role in local invasion and distant metastasis. COL1A1 may represent a stromal prognostic biomarker and therapeutic target in colon cancer.

Keywords: adipose tissue; colon cancer; extracellular matrix; tumour microenvironment.

Figure 1

Hierarchical clustering for the 50 genes with highest variance. The heat map diagram shows the result of the two-way hierarchical clustering of genes and samples. Each row represents one gene and each column represents one sample. The gene clustering tree is shown on the left. The colour scale shown at the top-left illustrates the relative expression level of genes across all samples: blue colour represents an expression level below mean, red colour represents expression higher than the mean.

Figure 2

Boxplots showing normalised expression values for the six most significantly differentially expressed genes.

Figure 3

Volcano plot showing the log2 fold changes (Peritumour/Distant, x-axis) and −log10 p-values for all genes.

Figure 4

Validation of the 5 most significantly up-regulated genes identified with PanCancer pathway analysis. RT-PCR confirmed higher RNA expression of COL1A1, FGF7 and SFRP2 in pAT compared to distal AT (COL1A1 n = 8, p = 0.007; FGF7 n = 9, p = 0.05; PLA2G2A n = 9, p = 0.16; NGFR n = 7, p = 0.10; SFRP2 n = 8, p = 0.05).

Figure 5

Validation with RT-PCR of 2 down-regulated genes identified with PanCancer pathway analysis (LEF1 n = 6, p = 0.03; CDH1 n = 6, p = 0.09).

Figure 6

Immunohistochemical staining for COL1A1 expression in colon cancer tissue, peritumour adipose tissue and distal adipose tissue in 5 colon cancer patients. A “mesenteric collagen architectural score” based on collagen bundles and architecture of adipose tissue lobes is reported for each patient. We hypothesize COL1A1 up-regulation is mediated by down regulation of Wingless/Integrated (Wnt) pathway, contributing to the creation of a propagating mesenteric collagen network to reach target organs and facilitate engraftment and colonisation.

Figure 6

Immunohistochemical staining for COL1A1 expression in colon cancer tissue, peritumour adipose tissue and distal adipose tissue in 5 colon cancer patients. A “mesenteric collagen architectural score” based on collagen bundles and architecture of adipose tissue lobes is reported for each patient. We hypothesize COL1A1 up-regulation is mediated by down regulation of Wingless/Integrated (Wnt) pathway, contributing to the creation of a propagating mesenteric collagen network to reach target organs and facilitate engraftment and colonisation.

Figure 7

Schematic representation of the role of Wingless/Integrated (Wnt) signalling in embryonic stem cell and pre adipocyte differentiation. Wnt signalling promotes differentiation of embryonic stem cell in mesenchymal stem cell. At the next stage, Wnt antagonists induce differentiation of pre adipocyte into mature adipocytes.