| Objective:To detect the Th17and Treg cells in the peripheral blood of primary esophageal carcinoma (PEC) patients and explore the relationship between Th17/Treg cells and clinical stages in PEC. Thereby to provide the experimental basis for the early diagnosis of PEC and the study on the Th17/Treg-targeting immunotherapy against PEC.Methods:Peripheral blood samples from87patients with PEC and39healthy volunteers were collected. Peripheral blood mononuclear cells (PBMC) were prepared by conventional method and flow cytometry was used to detect the percentage of Th17and Treg cells. Real-time PCR was used to detect the mRNA expression of proliferation and differentiation-related factors in Thl7and Treg cells, including IL-1R, IL-6, IL-17, IL-21, IL-22, IL-23, TGF-β, RORc and Foxp3. SPSS16.0statistical software and multiple linear regression analysis were taken to assess the correlation between the ratio of Th17and Treg cells and the clinical pathological stages, as well as other clinical indexes such as age, gender, site of tumor, differentiation degree, invasion depth and distant metastasis in patients with PEC.Results:1. Analysis with flow cytometry showed that①the number of Th17cells in peripheral blood of PEC patients was significantly higher than that in the normal control group (4.598±0.776%vs3.017±0.613%, P<0.001), and related with clinical stages(P=0.018) and lymph node metastasis(P>=0.026), but has nothing to do with gender, age, tumor differentiation, tumor size, and infiltration depth (P>0.05).②The number of Treg cells was significantly higher than that of normal control group (8.912±1.772%vs4.732±0.884%, P<0.001), and related with clinical stages (P=0.037) and differentiation degree (P=0.029), but has nothing to do with gender, age, tumor size, invasion depth, lymph node metastasis (P>0.05).③Ratio of Treg and Th17cells was significantly higher than that of normal control group (1.885±0.417vs1.568±0.338, P<0.001), and related with lymph node metastasis (P=0.032).2. Real-time PCR analysis showed that, compared with the normal control group, the mRNA expression level of IL-1R, IL-6, IL-17, IL-21, IL-22, IL-23, TGF-β, RORc and Foxp3, was significantly increased (P<0.05).3. The multiple linear regression analysis showed that, in the peripheral blood of PEC patients, the number of Th17and Treg cells was closely related with the TNM stages (t=3.794, P=0.018; t=2.961, P=0.037).Phase I and II were defined as the early stage, meanwhile, phase III and IV were defined as the advanced stage, in which the number of Th17and Treg cells and the ratio of Treg and Th17were significant higher compared with the control group (P<0.001). Compared with the advanced stage, Th17and Treg percent were also increased in the early stage (Thl7, early stage3.996±0.683%vs advanced stage4.887±0.793%, P<0.01; Treg, early stage7.395±1.690%vs advanced stage9.896±1.276%, P<0.01); but there was no statistical difference about the ratio of Treg/Th17between the early and advanced stages(1.820±0.298vs1.926±0.356, P>0.05). Synchronically increased Th17and Treg cells in a positive linear correlation(r=0.5832, P<0.01).Conclusions:1. In peripheral blood of PEC patients, the number of Th17and Treg cells, the ratio of Treg and Th17, and the mRNA expression of proliferation and differentiation-related factors were increased significantly compared with the normal group.2. There was an intimate correlation between the proportion of Th17and Treg cells in peripheral blood and the TNM stages in PEC. Both of them maintain a positive linear correlation with the TNM stage.3. Th17and Treg cells play an important role in the development of PEC. Above two kinds of immune cells may be the indexes for early diagnosis in PEC and may be new targets for immunotherapy against PEC. |
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