Th22 cell accumulation is associated with colorectal cancer development

Abstract

Aim: To investigate the expression of Th22 cells and related cytokines in colorectal cancer (CRC) tissues, and the probably mechanism.

Methods: CRC tumor and paratumor tissues were collected to detect the expression levels of Th22 cells and of related cytokines by immunohistochemistry, flow cytometry and real-time quantitative polymerase chain reaction (RT-qPCR). Interleukin (IL)-22 alone or with a STAT3 inhibitor was co-cultured with RKO cells in vitro to study the effects of IL-22 on colon cancer cells. IL-22 alone or with a STAT3 inhibitor was injected into a BALB/c nude mouse model with subcutaneously transplanted RKO cells to study the effects of IL-22 on colon cancer growth.

Results: The percentage of Th22 cells in the CD4(+) T subset was significantly higher in tumor tissues compared with that in paratumor tissues (1.47% ± 0.083% vs 1.23% ± 0.077%, P < 0.05) as determined by flow cytometry. RT-qPCR analysis revealed that the mRNA expression levels of IL-22, aryl hydrocarbon receptor, CCL20 and CCL22 were significantly higher in tumor tissues compared with those in paratumor tissues. CCL27 mRNA also displayed a higher expression level in tumor tissues compared with that in paratumor tissues; however, these levels were not significantly different (2.58 ± 0.93 vs 2.30 ± 0.78, P > 0.05). IL-22 enhanced colon cancer cell proliferation in vitro and displayed anti-apoptotic effects; these effects were blocked by adding a STAT3 inhibitor. IL-22 promoted tumor growth in BALB/c nude mice; however, this effect was reversed by adding a STAT3 inhibitor.

Conclusion: Th22 cells that accumulate in CRC may be associated with the chemotactic effect of the tumor microenvironment. IL-22 is associated with CRC development, most likely via STAT3 activation.

Keywords: Colorectal cancer; Interleukin-22; STAT3; Th22 cells; Tumor microenvironment.

Figure 1

Immunohistochemistry staining of tissues. A: Immunohistochemistry (IHC) staining of normal colon tissues; B: IHC staining of colon cancer tissues.

Figure 2

Expression of Th22 cells and related cytokines in colorectal cancer. A: Gated on FSC/SSC and CD4⁺ subset, the proportion of Th22 cells in the CD4⁺ subset is presented in quadrant Q1; B: Average proportion of Th22 cells in tumor and paratumor tissues; C: Expression levels of interleukin-22, AHR, CCL20, CCL22 and CCL27 in tumor and paratumor tissues were measured by RT-qPCR. The relative expression levels were normalized to the level of β-actin mRNA for each sample. Each bar represents the mean ± SE (n = 50), ^aP < 0.05, tumor vs paratumor. NS: Not significant.

Figure 3

Effects of interleukin-22 on colon cancer cells. A: Flow cytometry to measure colon cancer cell proliferation in the presence of interleukin (IL)-22 or IL-22 + S3I-201; B: Flow cytometry for colon cancer cell apoptosis in the presence of IL-22 or IL-22 + S3I-201; C: Average proportion of proliferating colon cancer cells in the presence of IL-22 or IL-22 + S3I-201; D: Average proportion of apoptotic colon cancer cells in the presence of IL-22 or IL-22 + S3I-201. Each bar represents the mean ± SE (n = 18). ^aP < 0.05 vs the control medium.

Figure 4

Effects of interleukin-22 on colon cancer in vivo. A: Tumor tissues were obtained from BALB/c nude mice; B: Tumor growth curves for interleukin (IL)-22, IL-22 + S3I-201 and control group mice. The tumor volume was calculated as follows: (major circumference × minor circumference²)/2. Each plot represents the mean ± SE (n = 7). ^aP < 0.05 vs the control.