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ORIGINAL ARTICLE
Year : 2017  |  Volume : 13  |  Issue : 2  |  Page : 252-256

Dual association of serum interleukin-10 levels with colorectal cancer


1 Shiraz Institute for Cancer Research, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
2 Colorectal Research Center, Shiraz University of Medical Sciences, Shiraz, Iran

Date of Web Publication23-Jun-2017

Correspondence Address:
Abbas Ghaderi
Shiraz Institute for Cancer Research, School of Medicine, Shiraz University of Medical Sciences, Shiraz
Iran
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0973-1482.199448

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 > Abstract 

Background: Interleukin 10 (IL-10) is considered an immune modulator cytokine, showing both antitumor and pro-tumor characteristics. Its role in the pathogenesis and progression of colorectal cancer depends on microenvironmental milieu.
Materials and Methods: A case–control study with 58 newly diagnosed colorectal cancer (CRC) patients, and 30 healthy individuals was conducted to compare the serum IL-10 levels between patients and controls. Furthermore, the correlation of the cytokine levels with the pathological features and prognosis of the CRC was investigated. IL-10 levels in the sera of patients and controls were measured by Enzyme-linked immunosorbent assay.
Results: Mean serum IL-10 levels were significantly lower in CRC patients than in controls (P = 0.04). CRC patients with worse prognosis at the time of diagnosis tend to have higher levels of circulating IL-10 than those with better prognosis (P = 0.008). Receiver operating characteristics curve analysis demonstrated that IL-10 levels in the sera of CRC patients can be used as a prognostic biomarker in CRC patients (area under the curve = 0.71; P = 0.01).
Conclusions: Our results demonstrated a dual association of serum IL-10 levels in the initiation and progression of CRC. While lower IL-10 levels were associated with higher risk of the disease, its higher levels were associated with a poorer prognosis.

Keywords: Cancer immunology, colorectal cancer, interleukin 10, regulatory T cells


How to cite this article:
Abtahi S, Davani F, Mojtahedi Z, Hosseini SV, Bananzadeh A, Ghaderi A. Dual association of serum interleukin-10 levels with colorectal cancer. J Can Res Ther 2017;13:252-6

How to cite this URL:
Abtahi S, Davani F, Mojtahedi Z, Hosseini SV, Bananzadeh A, Ghaderi A. Dual association of serum interleukin-10 levels with colorectal cancer. J Can Res Ther [serial online] 2017 [cited 2019 Jan 22];13:252-6. Available from: http://www.cancerjournal.net/text.asp?2017/13/2/252/199448


 > Introduction Top


Interleukin 10 (IL-10) is an anti-inflammatory cytokine with both protumor and antitumor properties depending on the milieu. IL-10 is mostly produced by type 1 regulatory T cells, also known as inducible regulatory T (T-reg) cells. This cytokine is also produced by many other cells including monocytes, macrophages, T helper (Th) 1 and Th2 cells and CD4+ Foxp3+ CD25+ “natural” T-reg cells.[1],[2] IL-10 is considered to be an anti-inflammatory cytokine, which suppresses macrophage/T-cell cytokine expression and inhibits antigen presenting capacity of these cells.[3] Despite this, there are experimental models demonstrating the opposite, when studying IL-10 under a different circumstance.[4]

The immune system plays an important role in all neoplasms including colorectal cancer (CRC), and chronic inflammation is a leading cause of CRC in patients with inflammatory bowel disease (IBD).[5],[6] All CRCs, including those associated with IBDs and sporadic ones, exhibit tumor-elicited immune/inflammatory infiltrates and different patient outcomes for distinct immune players has been demonstrated.[7] Cytokines and chemokines produced by tumor cells and cells recruited to the tumor environment like mast cells and macrophages are the main cause of inflammation.[8] There are several studies perusing these molecule levels in the sera of CRC patients. IL-10 is one of the cytokines that its role in CRC pathogenesis and treatment is of particular interest.

Regardless of etiology, most CRCs arises from adenomatous polyps. The probability of an adenoma developing into cancer is due to genetic and epigenetic changes in proto-oncogenes and tumor suppressor genes. Colorectal mucosal immune/inflammatory responses can either promote or inhibit CRC.[6] There are studies reporting higher serum levels of IL-10 in higher stages of CRC, while others did not find such associations.[9] The diagnostic value of some serum markers like carcinoembryonic antigen in CRC has been investigated extensively (10)[10] while using serum IL-10 as a marker needs more investigation.

As can be concluded, IL-10 has a disputable function in cancers, including CRC. We designated a case–control study to further clarify the role of IL-10 in CRC by comparing IL-10 levels in the sera of CRC patients and healthy individuals. Furthermore, this study was aimed to correlate the IL-10 serum levels with other clinical, pathological, and biochemical features of CRC patients.


 > Materials and Methods Top


Study design and participants

A case–control study was conducted to explore the role of serum IL-10 in susceptibility of individuals to CRC, besides discovering the correlation of this cytokine levels with the pathological features and stage of the CRC. Fifty-eight patients along with thirty sex/age-matched healthy individuals were recruited from clinics affiliated with Shiraz University of Medical Sciences. Only newly diagnosed patients were included in this study and patients undergone surgery, chemotherapy, and radiotherapy were excluded from this study. In addition, patients with a history of IBD (including ulcerative colitis and Crohn disease) were excluded from this study. All participants were informed about the study, and their will to participate was taken in written informed consent before study enrollment. Declaration of Helsinki was considered in the research performance and was approved by the Ethics Committee of the Shiraz University of Medical Sciences.

Patients were interviewed to obtain a thorough medical history, including initial presentation, age, gender, and family history of CRC. A detailed evaluation of patients including physical exam, chest radiograph, biochemical assessment of liver function, and measurement of carcinoembryonic antigen (CEA) was performed before surgery. During laparotomy, an en bloc resection of the tumor, its lymphatic drainage, and locally invaded structures was performed, and a minimum of 12 lymph nodes were sampled. Location and the size of the tumor were determined. Samples were examined by an experienced pathologist and the tumor type, grade, and pathological stage were defined. The staging was determined according to the American Joint Committee on Cancer, also known as the tumor node metastasis system.

Enzyme-linked immunosorbent assay for interleukin 10 and carcinoembryonic antigen

Blood samples were drawn from the selected patients before surgery, and the IL-10 and CEA levels of their sera were measured. Blood samples of thirty healthy individuals with no signs of any malignancy were collected for controls. Levels of IL-10 in the sera were measured by a quantitative enzyme-linked immunosorbent assay (ELISA) kit (eBioscience, UK) according to the protocols described by the manufacturer. CEA levels were measured by a noncompetitive biotin-avidin-based sandwich ELISA assay (eBioscience, UK) according to the manufacturer's instruction.

Statistical methods

Shapiro-Wilk test was employed to determine if variable values were normally distributed. Variables with normal distribution were presented as mean ± standard deviation, otherwise as the median. Frequencies were presented as a percentage. Nonparametric tests, including Mann–Whitney U-test and Kruskal–Wallis test, were used to analyze the difference among groups. Pearson correlation was utilized to correlate bivariate. Receiver operating characteristic (ROC) curve was established to evaluating IL-10 levels in CRC prognostication. P< 0.05 was considered statistically significant. Statistical Package for Social Sciences (SPSS, version 13; SPSS Inc., Chicago, IL, USA) was used for data analysis.


 > Results Top


A total of 58 patients with CRC and thirty healthy individuals as controls were included in this case–control study. Mean age in CRC patients was 54.81 ± 14.87 and in controls was 53.93 ± 12.70. CRC group consisted of 31 (53%) males and 27 (46.6%) females. 18 (60%) healthy males and 12 (40%) healthy females were elected as control group.

As the levels of IL-10 and CEA in patients and controls were not normally distributed (Shapiro–Wilk test, P= 0.00), nonparametric tests were employed to compare these values. Patients were more likely to have a lower average IL-10 level than control subjects [Figure 1]; 0.45240 vs. 0.81215 pg/ml; P= 0.04], but CEA levels were obviously higher in CRC group (3.388500 vs. 2.075450 ng/ml; P= 0.000).
Figure 1: Scatter dot plot, presenting interleukin-10 levels in sera of control subjects (Ctrl) and patients with colorectal cancer. Serum interleukin-10 levels were significantly lower in colorectal cancer patients (P < 0.05).The middle horizontal line presents the median and the upper and lower ones presents interquartile range

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CRCs patients were divided into two groups: Patients with poor outcome, and others. The criteria used for prognostication are: The number of lymph nodes involved (four or more involved for worse prognosis), tumor penetration through bowel wall (T3 and T4 considered for poorer prognosis), histopathological grade of the tumor (poorly differentiated considered with worse prognosis), perforation and adherence of tumor to adjacent organs (if present patient has poorer prognosis), vascular invasion (positive for worse prognosis), and CEA levels (>5.0 ng/ml increases the chance for recurrence) before surgery are considered. CRC patients having at least one of the above criteria were considered as patients with worse outcome (38 patients, 65.5%). We compared the IL-10 levels in the sera of these two groups of patients. IL-10 levels were significantly higher in sera of CRC patients with poor prognosis (1.2755 vs. 0.4277 pg/ml; P= 0.008).

ROCs curve analysis was employed to evaluate the effectiveness of serum IL-10 levels in CRC patients' prognostication. The area under the curve for serum IL-10 levels was 0.71 [Figure 2]; P= 0.01]. This means that IL-10 levels in the sera of CRC patients can be used as a prognostic factor. We presume the 0.3600 pg/ml value as the optimal cutoff point, with maximum specificity and sensitivity (75% and 66%, respectively).
Figure 2: Receiver operating characteristics curve for serum interleukin-10 levels in colorectal cancer patients' prognostication. The area under the curve of 0.71 shows that serum interleukin-10 levels can be used for outcome prediction

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We compared serum IL-10 levels in different clinicopathological groups of patients. IL-10 levels in the sera of patients with tumor invasion to vasculature were significantly higher than those without invasion (2.9383 vs. 0.6335 pg/ml; P= 0.041). We also observed that in CRC patients with CEA levels higher than 5 ng/ml, serum IL-10 levels were significantly higher (1.5189 vs. 0.6313 pg/ml; P= 0.002). There was no other statistically significant correlation between serum IL-10 levels and other clinicopathological feature [Table 1].
Table 1: Association of serum interleukin 10 levels with clinicopathological variables

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 > Discussion Top


In this study, we observed that IL-10 in the sera of CRC patients was significantly lower than the control group but higher in patients with poor prognosis. There are studies consistent with our results, showing lower levels of serum IL-10 were associated with neoplasia,[11] however others fail to find such associations.[12],[13] In a gene therapy study, Erdman et al. demonstrated IL-10-mediatiated inhibition of tumor growth in mice with CRC.[14] In preclinical models, IL-10 promotes Natural Killer cells cytotoxicity [4] and it seems at the early stages of immune response to malignant cells, IL-10 activates the innate arm of the immune system, and this may improve the following adaptive immune responses.[15] Lower levels of circulating IL-10 observed in the CRC patients can cause a defective innate immune response to neoplastic cells and consequently their escape from adaptive immunity.

Another explanation for the results observed may be interrelated to the role of IL-10 in cyclooxygenase 2 (COX-2) expression. COX-2 is overexpressed at sites of adenomatous polyps and carcinomas in the large bowel, and may be an early event in colorectal carcinogenesis.[16],[17] COX-2 contributes to tumorigenesis by inhibition of apoptosis, increased angiogenesis, conversion of procarcinogens to carcinogens and immunosuppression.[17] COX-2 mRNA expression is regulated at different levels by IL-10. IL-10 regulates COX-2 expression at the transcriptional level by inhibiting nuclear factor kappa B signaling which is involved in the COX-2 expression.[18],[19] In addition, it induces instability in the COX-2 mRNA resulting in its degradation and by this way controls COX-2 gene expression at posttranscriptional level.[20] The results observed in this study may be due to the inhibitory role of IL-10 in COX-2 expression, and lower levels of IL-10 in the sera may contribute to colorectal carcinogenesis by means of COX-2 overexpression.

On the other hand, there is evidence demonstrating IL-10 has a beneficial effect on the generation of CD8+ T-cell memory [21] and promotes the antitumor CD8+ T-cell effector function in situ.[22] Emmerich et al. demonstrated that IL-10 directly activates tumor-resident CD8+ T-cells and high doses of IL-10 increase the production of interferon-γ and granzymes in the peripheral blood [23] all of which can lead to a potent antitumor effect and lower levels of IL-10 causes a defective CD8+ response to colorectal neoplastic cells, and hence their progression to colorectal carcinoma. Moreover, Th 17 lymphocytes (a subgroup of CD4+ T-cells), together with the related cytokines, such as IL-6 and IL-17A, participate in human CRCs and numerous studies show a key role for these pro-inflammatory cytokines in enhancing the survival and growth of CRC cells.[24] It has been shown that IL-10 can suppress Th17 immune responses [25] and Huber et al. demonstrated that Tr1 cells could inhibit Th17 cells through an IL-10-dependent manner in vivo.[26] It might be possible that lower levels of IL-10 augment the impact of Th17 cells' on CRCs.

IL-10 levels in the sera of patients with tumor vascular invasion or CEA higher than 5 ng/ml were significantly higher than others. No other statistically significant correlation was observed between IL-10 levels in the sera of CRC patients and their age, gender, positive family history, stage of tumor, depth of tumor invasion into bowel wall, number of lymph nodes involved, distant metastasis, histopathological grade and mucin production of the tumor, perineural invasion, and tumor place in large bowel and size. There is evidence that CEA induces the release of IL-10.[27] When classifying the CRC patients into two groups for prognosis according to the criteria suggested by the previous studies,[10],[28],[29],[30],[31] IL-10 levels were significantly higher in patients with worse prognosis and circulating IL-10 levels can be prognostic in CRC patients, along with number of lymph nodes involved, penetration of tumor through bowel wall, bowel perforation, vascular invasion, and serum CEA levels.[10],[28],[29],[30],[31] There are studies in favor of our results, claiming that higher levels of circulating IL-10 are associated with worse outcome in CRC patients. In a study conducted by O'Hara et al. demonstrated higher levels of IL-10 in sera of advanced CRC patients.[31] Galizia et al. investigated the prognostic role of IL-10 serum levels in CRC patients, undergoing surgery and demonstrated that circulating IL-10 levels were among the variables affecting disease-free survival rate.[12],[13] In a meta-analysis performed by Zhao et al. a tight correlation was reported between high levels IL-10 in serum and poor prognosis in cancer patients.[32]

CRC tumors show the characteristics of chronic inflammation and in response to chronic inflammatory condition, epithelial cells, tumoral cells and immune cells in tumor microenvironment secrete cytokines, chemokines and growth factors, recruiting more inflammatory cells from circulation into the tumor site.[33] Interactions between neoplastic cells and other cells in tumor microenvironment contribute to cancer progression, and inflammatory infiltrates in the tumor microenvironment are essential for tumor metastasis.[34] As the tumor progresses, more IL-10+ tumor-associated macrophages and monocytes and other immune cells accumulate in the tumor microenvironment.[35] This fact is probably the reason of higher IL-10 levels in more progressed colorectal tumors.


 > Conclusion Top


We observed the association of the serum IL-10 levels in the initiation of CRC and its progression is different. Our results are consistent with the previous publications which showed a possible dual role of IL-10 in CRC. Further studies are required to discover the exact molecular and cellular mechanisms underlying this property of IL-10.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

 
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