Treg/Th17 cell balance and phytohaemagglutinin activation of T lymphocytes in peripheral blood of systemic sclerosis patients

摘要

AIM To investigate T-cell activation, the percentage of peripheral T regulatory cells(Tregs), Th17 cells and the circulating cytokine profile in systemic sclerosis(SSc).METHODS We enrolled a total of 24 SSc patients and 16 healthy controls in the study and divided the patients as having diffuse cutaneous SSc(dc SSc, n = 13) or limited cutaneous SSc(lc SSc, n = 11). We performed a further subdivision of the patients regarding the stage of the disease-early, intermediate or late. Peripheral venous blood samples were collected from all subjects. We performed flow cytometric analysis of the activationcapacity of T-lymphocytes upon stimulation with PHA-M and of the percentage of peripheral Tregs and Th17 cells in both patients and healthy controls. We used ELISA to quantitate serum levels of human interleukin(IL)-6, IL-10, tissue growth factor-β1(TGF-β1), and IL-17 A.RESULTS We identified a decreased percentage of CD3+CD69+ cells in PHA-stimulated samples from SSc patients in comparison with healthy controls(13.35% ± 2.90% vs 37.03% ± 2.33%, P < 0.001). However, we did not establish a correlation between the down-regulated CD3+CD69+ cells and the clinical subset, nor regarding the stage of the disease. The activated CD4+CD25+ peripheral lymphocytes were represented in decreased percentage in patients when compared to controls(6.30% ± 0.68% vs 9.36% ± 1.08%, P = 0.016). Regarding the forms of the disease, dc SSc patients demonstrated lower frequency of CD4+CD25+ T cells against healthy subjects(5.95% ± 0.89% vs 9.36% ± 1.08%, P = 0.025). With regard to Th17 cells, our patients demonstrated increased percentage in comparison with controls(18.13% ± 1.55% vs 13.73% ± 1.21%, P = 0.031). We detected up-regulated Th17 cells within the lc SSc subset against controls(20.46% ± 2.41% vs 13.73% ± 1.21%, P = 0.025), nevertheless no difference was found between dc SSc and lc SSc patients. Flow cytometric analysis revealed an increased percentage of CD4+CD25-Foxp3+ in dc SSc patients compared to controls(10.94% ± 1.65% vs 6.88% ± 0.91, P = 0.032). Regarding the peripheral cytokine profile, we detected raised levels of IL-6 [2.10(1.05-4.60) pg/m L vs 0.00 pg/m L, P < 0.001], TGF-β1(19.94 ± 3.35 ng/m L vs 10.03 ± 2.25 ng/m L, P = 0.02), IL-10(2.83 ± 0.44 pg/m L vs 0.68 ± 0.51 pg/m L, P = 0.008), and IL-17 A [6.30(2.50-15.60) pg/m L vs 0(0.00-0.05) pg/m L, P < 0.001] in patients when compared to healthy controls. Furthermore, we found increased circulating IL-10, TGF-β, IL-6 and IL-17 A in the lc SSc subset vs control subjects, as it follows: IL-10(3.32 ± 0.59 pg/m L vs 0.68 ± 0.51 pg/m L, P = 0.003), TGF-β1(22.82 ± 4.99 ng/m L vs 10.03 ± 2.25 ng/m L, P = 0.031), IL-6 [2.08(1.51-4.69) pg/m L vs 0.00 pg/m L, P < 0.001], and IL-17 A [14.50(8.55-41.65) pg/m L vs 0.00(0.00-0.05) pg/m L, P < 0.001]. Furthermore, circulating IL-17 A was higher in lc SSc as opposed to dc SSc subset(31.99 ± 13.29 pg/m L vs 7.14 ± 3.01 pg/m L, P = 0.008). Within the dc SSc subset, raised levels of IL-17 A and IL-6 were detected vs healthy controls: IL-17 A [2.60(0.45-9.80) pg/m L vs 0.00(0.00-0.05) pg/m L, P < 0.001], IL-6 [2.80(1.03-7.23) pg/m L vs 0.00 pg/m L, P < 0.001]. Regarding the stages of the disease, TGF-β1 serum levels were increased in early stage against late stage, independently from the SSc phenotype(30.03 ± 4.59 ng/m L vs 13.08 ± 4.50 ng/m L, P = 0.017).CONCLUSION It is likely that the altered percentage of Th17 and CD4+CD25-Fox P3+ cells along with the peripheral cytokine profile in patients with SSc may play a key role in the pathogenesis of the disease.