The Imbalance between Foxp3⁺Tregs and Th1/Th17/Th22 Cells in Patients with Newly Diagnosed Autoimmune Hepatitis

Figure 2

FACS analysis of the numbers of different subsets of circulating effector CD3⁺CD4⁺T cells and ELISA analysis of serum IFN-γ, IL-17, and IL-22 in AIH patients. PBMCs were isolated from individual subjects, and PBMCs were stained in duplicate with APC-anti-CD4 and PerCP-anti-CD3 or isotype controls, fixed, and permeabilized, followed by intracellular staining with FITC-anti-IL-17 and PE-Cy7-anti-IFN-γ and PE-anti-IL-22. The frequency of CD3⁺CD4⁺IFN-γ⁺Th1, CD3⁺CD4⁺IL-17⁺Th17, and CD3⁺CD4⁺IL-22⁺Th22 cells was determined by flow cytometry analysis. The cells were gated on living lymphocytes and then gated on CD3⁺CD4⁺ cells, and at least about 30,000 events were analyzed for each sample. The numbers of each type of CD3⁺CD4⁺T cells were calculated, according to the total numbers of PBMCs and different types of CD3⁺CD4⁺T cells. (a) The flow cytometry analysis; (b–d) the numbers of CD3⁺CD4⁺IFN-γ⁺Th1, CD3⁺CD4⁺IL-17⁺Th17, and CD3⁺CD4⁺IL-22⁺Th22 cells; (e–g) serum levels of IFN-γ, IL-17, and IL-22. Data shown are representative FACS charts or the mean numbers of each type of cells per mL of peripheral blood in individual subjects from two separate experiments. The horizontal lines indicate the median values for each group. Data shown are representative charts of different subsets of CD3⁺CD4⁺T cells and serum IL-10 from individual groups of subjects ( for the HC, for the patients at 0 week, and for the patients at 8 weeks posttreatment).

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