Expression Levels Of Inhibitory Molecules On Regulatory T Cells And Serum Adipocytokines Correlate With Progression Of NSCLC

Objectives To detect the expression levels of CTLA-4, LAG-3, PD-1, LepR and CD39on CD4+T cells and CD4+CD25+FoxP3+Treg cells in peripheral blood or tumor tissuesfrom NSCLC patients and to discover their potential internal relationships with diseasestatus of NSCLC; To assay the serum adipokines concentrations of Leptin, adiponectin,resistin, PAI-1and soluble LepR, and investigate the relationships between Treg cells andLeptin/soluble LepR in NSCLC. Methods88objects including newly-admitted53NSCLC patients,17disease control patients and18healthy controls were studied.Mononuclear cells were obtained from anticoagulant fresh peripheral blood of allsubjects and13cases of tumor and tumor-adjacent tissues of NSCLC surgery patients.CD4+T, CD4+CD25+FoxP3+Treg cells and the five kinds of inhibitory molecules/receptorwere assessed by flowcytometry. To detect the serum concentrtions of TGF-β, IL-10,leptin, Adiponectin, resistin, PAI-1and soluble LepR By ELISA. ResultsCD4+CTLA-4+（2.79%±2.43%vs1.75%±0.52%,1.56%±0.96%）and CD4+PD-1+T（12.94%±5.96%vs10.83%±3.68%,8.96%±2.27%）cells were significantly higher inthe peripheral blood of NSCLC patients than those in disease or healthy controls; Theratio of CD4+PD-1+T cells significantly increased in patients with stages Ⅲ and ⅣNSCLC as compared with patients with stagesⅠand ⅡNSCLC（13.21%%±5.96%vs11.06%±3.42%）; Considerable CD4+CTLA-4+T cells（5.07%±2.11%vs3.02%±1.35%）and CD4+PD-1+T cells（40.20%+18.84%vs27.82%±16.59%）were identified in tumortissues which were less detected in tumor-adjacent tissues; more CD4+CTLA-4+T（5.07%±2.11%vs3.13%±1.01%）, CD4+LAG-3+T（7.86%±3.24%vs2.65%±1.48%）and CD4+PD-1+T（40.20%±18.84%vs15.79%±5.69%）cells were verified in tumortissues than in their matched peripheral blood. The percentages of Treg（9.12%±3.57%vs6.43%±2.48%）, CTLA-4+Treg （6.01%±4.49%vs2.53%±2.04%）, LAG-3+Treg（4.89%±4.80%vs1.79%±2.18%） and PD-1+Treg （20.14%±11.57%vs11.68%±5.86%）cells were higher in NSCLC patients than in healthy controls. The ratiosof Treg （9.78%±2.95%vs8.26%±2.51%） and CTLA-4+Treg （7.15%±2.83%vs5.41%±3.01%） cells also significantly increased in patients with stages Ⅲ and ⅣNSCLC; The percentages of Treg（36.92%±15.11%vs18.87%±8.17%）, CTLA-4+Treg（13.27%±6.63%vs9.31%±5.15%）, LAG3+Treg（8.99%±3.78%vs6.66%±3.66%）and PD-1+Treg（54.42%±16.96%vs44.25%±13.56%）cells were identified in tumortissues than in tumor-adjacent tissues, and than in peripheral blood[Treg （36.92%±15.11%vs8.78%±2.95%)],[CTLA-4+Treg （13.27%±6.63%vs6.50%±3.61%）],[LAG3+Treg （8.99%±3.78%vs5.04%±3.80%）],[PD-1+Treg（54.42%±16.96%vs35.36%±16.39%）], simultaneously. Serum concentrations ofTGF-β[（20.24±6.08vs14.85±6.17,16.21±5.09）ng/ml] and IL-10[（3.31±1.65vs3.05±1.63,0.57±0.36）pg/ml] were significantly higher in NSCLC patients than indisease controls and healthy controls, and there was no significantly negative correlationrelationship between the percentage of Treg cells and TGF-β or IL-10level in peripheralblood, respectively. There were no statistical differences in age and gender between3groups, adjusted by BMI, serum adiponectin concentration in NSCLC patients wassignificantly higher than in healthy controls[（6.08±3.56vs3.87±2.40）ng/ml], resistin[（4.26±3.22vs7.71±5.43,8.93±6.67） ng/ml] and soluble LepR[（223.23±107.77vs363.55±85.09）pg/ml] concentrations were significantly lower inNSCLC patients than in disease controls and healthy controls, there were no significantdifferrences statistically in leptin and PAI-1level between3groups. The percentage ofLepR+Treg cells in peripheral blood and tumor tissues of NSCLC patients weresignificantly higher than in healthy controls（3.81%±2.52%vs0.89%±0.97%） and inmatched tumor-adjacent tissues（6.45%±3.12%vs1.99%±1.61%）, respectively. Theremight exist significantly negative correlation relationship between serum Leptin leveland Tregs percentage, existing significnatly positive correlation relationship betweenserum LepR level and Tregs percentage, and signifianctly negative correlationrelationship between serum Leptin concentration and Lep+Tregs percentage, however.However, these statistically significant correlation relationships aboved were not foundin subjects of disease controls or healthy controls. Conclusions Increased expressionsof inhibitory molecules/receptor, including CTLA-4, LAG-3, PD-1and LepR, on CD4+Tand Treg cells in peripheral blood and tumor tissues of NSCLC patients, may be one ofthe mechanisms related to immune escape of tumor cells, progression of disease and poorprognosis. The serum concentrations changes of adiponectin, resistin and soluble LepRindicate that these adipokines may be involved the occurrence and progression ofNSCLC. The correlations between Tregs and leptin, Tregs and leptin receptor,LepR+Tregs and leptin confirmed that the number and ratio of Treg cells could beregulated by leptin system; adjusting Leptin/LepR-Tregs axis is expected to be animmunomodulatory treatment strategy to targeted Treg cells, and may contribute toNSCLC patients.