Immune modulation of Stem Cell Educator therapy. Patient lymphocytes were isolated from peripheral blood by Ficoll-Hypaque technique (γ = 1.077) for flow cytometric analyses in AA patients at baseline and 4 weeks after Stem Cell Educator therapy. Isotype-matched IgG served as control. Histologic examination of alopecic skin (C and D). (A) Flow analysis of intracellular cytokines demonstrating differential effects on key interleukins at 4 weeks post-treatment. (B) Flow analysis of costimulating molecules demonstrating an increase of CD28 expression at 4 weeks post-treatment. Data are representative of preparations from all AA participants (n = 9) who received Stem Cell Educator therapy. (C) Fresh skin tissues were collected from the scalps via biopsy for immunohistochemistry testing in participants with alopecia totalis before treatment and 12 weeks after receiving Stem Cell Educator therapy. TGF-β1 staining surrounds a hair follicle of AA participants after receiving Stem Cell Educator therapy, with vertical section of hair follicle (top panels) and horizontal section of hair follicle. Isotype-matched mouse IgG1 served as a negative control for TGF-β1 immunostaining in a serial hair follicle section. Representative images were obtained from five experiments. Scale bar, 25 μm. (D) H&E staining of scalp tissues. Scale bar, 25 μm.