Differential mucosal expression of Th17-related genes between the inflamed colon and ileum of patients with inflammatory bowel disease

Abstract

Background: Immunological and genetic findings implicate Th17 effector cytokines in the pathogenesis of inflammatory bowel disease (IBD). Expression of Th17 pathway-associated genes is mainly studied in colonic disease. The present study assessed the mRNA expression levels of Th17 effector cytokines (IL17A, IL17F, IL21, IL22 and IL26) and genes involved in differentiation (IL6, IL1B, TGFB1, IL23A and STAT3) and recruitment of Th17 cells (CCR6 and CCL20) by quantitative real-time PCR analysis of colonic and ileal biopsies from 22 healthy control subjects, 26 patients with Crohn's disease (CD) and 12 patients with ulcerative colitis (UC). Inflammation was quantified by measuring expression of the inflammatory mediators IL8 and TNF.

Results: Evaluation of mRNA expression levels in colonic and ileal control samples revealed that TNF, TGFB1, STAT3 and CCR6 were expressed at higher levels in the ileum than in the colon. Expression of all the Th17 pathway-associated genes was increased in inflamed colonic samples. The increased expression of these genes was predominantly observed in samples from UC patients and was associated with more intense inflammation. Although increased expression of IL17A, IL17F, IL21 and IL26 was detected in inflamed ileal samples, expression of the indispensable Th17 cell differentiation factors TGFB1 and IL23A, the signaling molecule STAT3 and the Th17 recruitment factors CCR6 and CCL20 were unchanged.

Conclusions: Our findings suggest that immune regulation is different in colonic and ileal disease, which might have important consequences for therapeutic intervention.

Figure 1

mRNA expression levels of proinflammatory and Th17-pathway-associated genes in colon and ileum of healthy controls. The relative transcription levels are determined by SYBR Green qPCR using the geometric mean of GAPDH, HPRT and SDHA as an endogenous control in colonic and ileal samples from healthy controls. The data are expressed as geometric means. The geometric mean of the colonic controls was set as 1. Data are presented on a log scale. (a) proinflammatory cytokines, (b) Th17 effector cytokines, (c) Th17 differentiation factors, (d) Th17 recruitment factors. *P < 0.05, **P < 0.01, ***P < 0.001

Figure 2

mRNA expression levels of proinflammatory and Th17-pathway-associated genes in inflamed colonic and ileal samples of IBD patients. Fold changes in transcription levels are determined by SYBR Green qPCR using the geometric mean of GAPDH, HPRT and SDHA as an endogenous control in inflamed colonic UC, colonic CD and ileal CD samples. The fold changes shown are relative to control samples. The data are presented as box plots with medians and interquartile ranges. (a) proinflammatory cytokines, (b) Th17 effector cytokines, (c) Th17 differentiation factors, (d) Th17 recruitment factors. *P < 0.05, **P < 0.01, ***P < 0.001, ^#P < 0.0001

Figure 3

mRNA expression levels of RORC in inflamed colonic and ileal samples of IBD patients. The relative transcription levels of RORC are determined by SYBR Green qPCR using the geometric mean of GAPDH, HPRT and SDHA as an endogenous control in (a) colonic and (b) ileal samples from healthy controls, UC patients and CD patients. The geometric mean of the control samples was set as 1. Data are presented on a log scale. **P < 0.01