The Changes Of CD4~+CD25~+ Regulatory T Cells Of Non Small-cell Lung Cancer Patients And The Effects Of CpG ODN

[Objective] To observe the changes of the proportion of CD4+CD25+ regulatory T cells and the levels of TGF-βand IFN-γin the peripheral blood of NSCLC patients and to study the influencing factors related to the changes.[Methods] A total of 30 NSCLC patients diagnosed pathologically without another treatments except for the operation were involved in the study. And as the control group,30 healthy volunteers were selected. The peripheral blood mononuclear cell (PBMC) isolated from the peripheral blood of the NSCLC patients and the healthy volunteers were analyzed for the proportion of CD4+CD25+ T cells with flow cytometry, the mRNA expression of Foxp3 with real-time PCR and the levels of TGF-βand IFN-γwith ELISA.[Results] The proportion of CD4+CD25+ T cells, the mRNA expression of Foxp3 and the levels of TGF-βin the group of NSCLC patients were 42.14±14.32%,0.96±0.44 and 67.24±8.79 pg/ml respectively and were statistically higher than those of the healthy volunteers, which were 8.42±2.69%,0.66±0.09 and 33.58±4.74 pg/ml respectively. But there was no statistical difference in the different subgroups of pathological classification, clinical stage and pathological rank in NSCLC patients. There was no significant difference in the levels of IFN-γbetween the NSCLC patients and the healthy volunteers, which were 30.40±9.52 pg/ml and 33.33±20.02 pg/ml respectively. In the NSCLC patients, the level of TGF-βwas linear correlated with CD4+CD25+ T cells proportion (partial regression coefficient=0.598, P=0.046).[Conclusions] The proportion of CD4+CD25+ Foxp3+ T cells and the level of TGF-p in the PBMC of NSCLC patients were higher than those in the healthy volunteers. There was linear correlation between the proportion of CD4+CD25+ Foxp3+ T cells and the level of TGF-p in the NSCLC patients.[Objective] To observe the expression of Foxp3 gene in tumor microenvironment and metastatic/non-metastatic lymph nodes of NSCLC patients.[Methods] The NSCLC patients involved in the study were the same as in part one. The paraffin sections of lung cancer and lymph nodes were taken from the resected lung specimens. The expression of Foxp3 in lung cancer tissue, intersection of cancer and normal lung tissue and metastatic/non-metastatic lymph nodes were detected with immunohistochemistry. The levels of TGF-βand IFN-γin lung cancer tissue homogenate were tested with ELISA.[Results] The optimistic density of positive Foxp3 expression in lung cancer tissue and intersection of cancer and normal lung tissue in NSCLC patients was 14.06±4.14 and 13.29±3.81 respectively, and there was no statistical difference between the two tissues (P>0.05). The expression of Foxp3 in cancer tissue and intersectional part of adenosquamous carcinoma were higher than those of squamous carcinoma (20.50 vs 12.33±2.59,19.90 vs 11.52±2.25, P<0.05), while there was no statistical difference either between squamous carcinoma and adenocarcinoma or between adenosquamous carcinoma and adenocarcinoma (P>0.05).The OD values of Foxp3 expression in cancer tissue of clinical stageⅡ,ⅢandⅣpatients were 15.22±3.07,16.58±2.95 and 18.17±1.82, respectively, which were statistically higher than that of stage I patients (9.35±0.81, P<0.05). The OD values of Foxp3 expression in intersection part of clinical stageⅡ,ⅢandⅣpatients were 13.98±3.61,15.48±2.73 and 17.33±1.99, respectively, which were also statistically higher than that of stageⅠpatients (9.25±0.71, P<0.05). But there was no statistical difference in the Foxp3 expression among the subgroups of stageⅡ,Ⅲ andⅣ(P>0.05). There was no statistical difference in the Foxp3 expression of cancer tissue and intersection part among the subgroups of rankⅡ,Ⅱ-ⅢandⅢ(P>0.05). The Foxp3 expression of metastatic lymph nodes was statistically higher than that of non-metastatic lymph nodes (16.28±3.31 vs 8.62±0.74, P<0.05). There was no statistical difference in Foxp3 expression among the different subgroups of pathological classification, clinical stage and pathological rank, except the Foxp3 expression in metastatic lymph nodes of adenosquamous carcinoma was statistically higher than that of squamous carcinoma (19.60 vs 11.88±0.85, P<0.05). There was no statistical difference in the levels of TGF-βand IFN-γin lung cancer tissue homogenate among the different subgroups of pathological classification, clinical stage and pathological rank(P>0.05).[Conclusions] The expression of Foxp3 in metastatic lymph nodes of NSCLC patients was statistically higher than that of non-metastatic lymph nodes and the Foxp3 expression was much higher in tumor microenvironment of middle-late NSCLC patients.[Objective] To study the effects of CpG ODN on the proportion of CD4+CD25+ regulatory T cells, Foxp3 expression and the levels of TGF-βand IFN-γin the peripheral blood of NSCLC patients.[Methods] NSCLC patients involved in the study were the same as in part one. The PBMC of NSCLC patients were randomly divided into treated group (CpG ODN 2006) and placebo group (CpG ODN 1612). The treated group and placebo group were given with either 1μM CpG ODN 2006 or CpG ODN 1612 stimulation. The proportion of CD4+CD25+ T cells, the mRNA expression of Foxp3 and the levels of TGF-βand IFN-γwere measured and compared before and after treatment.[Results] After the treatment with CpG ODN 2006, the proportion of CD4+CD25+ T cell was reduced from 44.16±11.61% to 36.52±10.78%(P< 0.05), Foxp3 gene expression was degraded from 1.15±0.48 to 0.65±0.17 (P< 0.05), the level of TGF-p was decreased from 68.25±9.25 pg/ml to 58.22±4.16 pg/ml (P< 0.05). There was no statistical difference in these parameters before and after treatment in CpG ODN 1612 group (40.11±16.76% vs 40.92±16.93%,0.77±0.33 vs 0.78±0.21,65.84±9.13pg/ml vs 58.96±4.73pg/ml, P>0.05). There was no statistical difference in the levels of IFN-y before and after treatment in both groups.[Conclusions] CpG ODN 2006 could down-regulate the proportion of CD4+CD25+ Foxp3+ regulatory T cells and the level of TGF-p in the peripheral blood of NSCLC patients, while it had little effect on the level of IFN-y.