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Objective To evaluate the relationship of peripheral blood Th17 and regulatory T (Treg) cells with disease activity in patients with systemic sclerosis (SSc).Methods This study recruited 21 patients with active SSc,24 patients with inactive SSc and 24 normal human controls with informed consent.Peripheral blood samples were obtained from these subjects.Flow cytometry was used to detect the percentages of Th17 and Treg cells in peripheral blood CD4+ cells,a fluorescence-based quantitative PCR to determine the levels of interleukin (IL)-17A,retinoid-related orphan receptor gamma t (RoRγt),forkhead box P3 (FoxP3) mRNA in peripheral blood mononuclear cells (PMBCs),and enzyme linked immunosorbent assay to measure the serum level of IL-17.Results Increased percentage of Th17 cells in peripheral blood CD4+ cells was observed in patients with active SSc compared with those with inactive SSc and normal human controls (2.34％ ± 1.19％vs.0.68％ ± 0.39％ and 0.57％ ± 0.49％,respectively,both P ＜ 0.05).No statistical difference was noted in the percentage of Treg cells in CD4+ cells or the mRNA expression levels of FoxP3 between the patients with active SSc,inactive SSc and normal human controls (all P ＞ 0.05).There was a significant increase in the mRNA expression of IL-17A,RoRγt in PBMCs and serum levels of IL-17 in patients with active SSc compared with patients with inactive SSc and normal human controls ( 11.73 ± 0.80 vs.9.77 ± 1.30 and 10.79 ± 0.74,respectively,both P ＜ 0.05; 18.48 ± 1.09 vs.15.89 ± 1.48 and 17.77 ± 1.64,respectively,both P ＜ 0.05; 53.60± 9.90 pg/ml vs.15.18 ± 3.24 pg/ml and 15.53 ± 4.12 pg/ml,respectively,both P ＜ 0.05).The percentage of Th17 cells in CD4+ cells and serum IL-17 levels were both positively correlated with disease activity in patients with active SSc (r =0.675,0.644,respectively,both P ＜ 0.05).Conclusions Th17 cells are highly proliferative in patients with active SSc,which may be closely correlated with the activity of SSc.
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AIM:To simultaneously determine jatrorrhizine hydrochloride,palmatine hydrochloride,berberine hydrochloride、 evodiamine and rutaecarpine in Zuojin Pill(Rhizoma Coptidis,Fructus Evodiae).METHODS:A RP-HPLC method was established with gradient elution and an agilent C_ 18 column(250 mm?4.6 mm,5 ?m)was used.The mobile phase A was acetonitrile and the mobile phase B was water with 0.3% phosphoric acid-triethylamine.The detective wavelength was at 225 nm.RESULTS:Jatrorrhizine hydrochloride had a good linearity in the range of 0.013-0.065 ?g.The average recovery was 103.1% with RSD of 1.1%.Palmatine hydrochloride had a good linearity in the range of 0.013-0.102 ?g.The average recovery was 99.8% with RSD of 1.9%.Berberine hydrochloride had a good linearity in the range of 0.063-0.317 ?g.The average recovery was 99.8% with RSD of 1.4%.Evodiamine had a good linearity in the range of 0.001-0.007 ?g.The averoge recovery was 100.5% with RSD of 1.5%.Rutaecarpine had a good linearity in the range of 0.002-0.010 ?g.The average recovery was 98.9% with RSD of 1.9%.CONCLUSION:This method is simple,accurate and reproducible.It can be used to determine jatrorrhizine hydrochloride、 palmatine hydrochloride、 berberine hydrochloride、 evodiamine and rutaecarpine together with the same chromatogram condition.