Effect of Notch1 signaling pathway on the differentiation and function of Th17 cells in peripheral blood of patients with psoriasis / 中华皮肤科杂志

ABSTRACT

Objective To evaluate the effect of Notch 1 signaling pathway on the differentiation and function of Th17 cells in peripheral blood of patients with psoriasis.Methods Peripheral blood samples were obtained from 35 patients with psoriasis and 32 healthy controls.Flow cytometry was performed to determine the proportion of Th17 cells in CD4+ T cells in peripheral blood mononuclear cells (PBMCs),real-time RT-PCR to measure the mRNA expression of retinoic acid receptor-related orphan receptor γt (RORγt),interleukin (IL)-17,Notch 1 and hairy-and-enhancer-of-split-1 (Hes-1),and enzymelinked immunosorbent assay (ELISA) to detect levels of IL-17 in the serum and culture supernatant of PBMCs stimulated with phorbol ester,calcium ionophore and brefeldin A.The correlation of Notch 1 mRNA expression with psoriasis area and severity index (PASI) was analyzed,so were its correlation with the proportion of Th17 cells,mRNA expression of RORγt,and mRNA and protein expression of IL-17.PBMCs isolated from the patients with psoriasis were divided into 5 groups to be treated with γ-secretase inhibitor N-[N-(3,5-difluorophenacetyl)-1-alanyl]-S-phenylglycine t-butyl ester (DAPT) at different concentrations of 0(control group),2.5,5.0,10.0 and 20.0μmol/L,respectively,so as to evaluate the effects of blocking the Notch1 signaling pathway by DAPT on the proportion of Th17 cells in PBMCs,levels of RORγt and IL-17.Results Compared with the healthy controls,patients with psoriasis showed a significant increase in the proportion of Th17 cells in CD4+ T cells in PBMCs (2.863％ ± 0.969％ vs.0.604％ ± 0.124％,P ＜ 0.01),mRNA expression of RORγt (5.255 ± 0.998 vs.1.530-± 0.485,P ＜ 0.01),Notch1 (6.743 ± 1.756 vs.1.731 ± 0.456,P ＜ 0.01),Hes-1 (6.384 ± 1.665 vs.1.627 ± 0.485,P ＜ 0.01) and IL-17 (6.944 ± 1.626 vs.1.698 ± 0.329,P ＜ 0.01),and serum level of IL-17 ([36.444 ± 5.936] ng/L vs.[11.762 ± 2.260] ng/L,P ＜ 0.01).Among the patients with psoriasis,the mRNA expression of Notch1 was positively correlated with PASI scores,proportion of Th17 cells,mRNA expression of RORγt and IL-17,and serum level of IL-17 (r =0.584,0.544,0.518,0.549 and 0.511,respectively,all P ＜ 0.05).There were significant differences in the proportion of Th17 cells,mRNA expression of RORγt and IL-17,and level of IL-17 in the culture supematant among the control group,2.5-,5.0-,10.0-and 20.0-μmol/L DAPT groups (F =79.527,82.239,78.086 and 80.558,respectively,all P ＜ 0.01).The above indices were significantly lower in the 2.5-,5.0-,10.0-and 20.0-μmol/L DAPT groups than in the control group (all P ＜ 0.05),and decreased along with the increase of DAPT concentrations.Conclusion Notch1 signaling pathway can promote the differentiation of Th17 cells and the expression of RORγt,IL-17 and Hes-1 in the peripheral blood of patients with psoriasis.