Chronic inflammation confers to the metabolic reprogramming associated with tumorigenesis of colorectal cancer

Abstract

It's well known that microenvironment inflammatory signals could promote cancer development and progression. In colorectal cancer (CRC), chronic inflammation is a major driving mechanism for the development of CRC in patients having long-standing inflammatory bowel disease (IBD). Though it has been addressed that cancer cells ferment much of their glucose supply into lactate regardless of the presence of oxygen, it is unclear whether cell metabolism has been reprogramed during the process from IBD to CRC. Herein, with dextran sulfate sodium (DSS)-induced mouse colitis model, we found that inflammation upregulated key glycolytic enzymes expression via activation of STAT3/c-Myc signaling pathway. Interestingly, during the whole phase of chronic inflammation, the key metabolic enzymes demonstrated increased expression constantly, indicating the metabolic reprogramming was induced by long-term inflammatory signal. Moreover, either the inhibition of STAT3 signaling or c-Myc activity could block the glycolytic enzymes expression induced by interleukin 6 (IL-6). Thus, we presented the view that inflammation could induce the metabolic reprogramming and promote the progression from chronic colitis to colorectal cancer.

Keywords: Chronic inflammation; IL-6; STAT3; c-Myc; metabolic reprogramming.

Figure 1.

Upregulation of glycolytic enzymes expression in DSS-induced acute colitis model. (A) Body weight change percentage was calculated in control WT (gray) and DSS-treated group, namely 2.5% DSS-induced mice (red) and 4% DSS (blue). Colonic length ratio, and photographs of large intestine, oriented from cecum to rectum, was presented. Data expressed as mean ± SEM. Statistical analysis was performed using 2-way ANOVA with Bonferroni posttest to calculate p values. Significant resistance to colitis, as established by percent of weight loss (p < 0.01) and colonic weight/length ratio (p < 0.001) were observed in DSS-treated mice, including 2.5% and 4%, as compared with WT mice. *p < 0.05, **p < 0.01, ***p < 0.001. (B) Representative H&E-stained colon sections (magnification *200). (C) Histology score. Values were expressed as means ± SD (n = 10). *p < 0.05, **p < 0.01, ***p < 0.001 vs. control group. (D) Inmmunohistochemical staining showing expression of the key enzymes in colon mucosa of DSS-induced colitis and control groups. (E and F) The mRNA and protein expression of the key enzymes of aerobic glycolysis. (G) Quantitative analysis of PKM2 protein level.

Figure 2.

The key metabolic enzymes expression significantly increased in DSS-induced chronic colitis model. (A) Representative H&E-stained colon sections (magnification *200). (B) Histology score. Values were expressed as means ± SD (n = 10). *p < 0.05, **p < 0.01, ***p < 0.001 vs. control group. (C) Inmmunohistochemical staining showing expression of the key enzymes in colon mucosa of each DSS-induced colitis groups and control group. (D and E) The mRNA and protein expression of the key enzymes of aerobic glycolysis.

Figure 3.

Pro-inflammatory cytokine IL-6 promoted the expression and activity of glycolytic enzymes in CRC and normal cells. (A) Both CRC and normal cells proliferation and / or viability using MTT assay. Error bars represent cell numbers ± SD for triplicate experiments. (B) Colony- formation assay with HIEC cells treated with IL-6 and 5-Fu as indicated. (C) Cell apoptosis was detected of HIEC cells treated with IL-6 and 5-Fu using FITC-Annexin V / PI staining kit and analyzed using flow cytometry. Representative flow cytometry results are shown. (D and E) mRNA and protein expression levels of key glycolytic enzymes in HIEC cell tread with IL-6. (F and G) Enzyme activity analysis of LDH-A and PKM2 in HIEC, SW480 and SW620 cells respectively. (H) Detection of lactate production with IL-6 treatment. Data was shown as mean ± SE of the above independent experiments.

Figure 4.

Protein expression analysis of the key glycolytic enzymes after inhibition of stat3/c-Myc signaling. (A) The expression of phos-STAT3, c-Myc, LDHA and HK2 were inhibited by STAT3 inhibitor S3I-201. (B) HT-29 cells first treated with IL-6 in advance to activate c-Myc and phos-STAT3 expression. Then glycolytic enzymes protein expression was detected after S3I-201 treatment. (C and D) c-Myc expression was inhibited by Myc inhibitor JQ1 in HIEC, then analyzed the key enzymes expression with JQ1 only or combined with IL-6 respectively. (E) S3I-201 blocked c-Myc and glycolytic enzymes expression within a short time after cells pre-treated IL-6 in 12 hours. (F) A scheme of the mechanism involved in the inflammation-induced metabolic reprogramming.