MSG resident and blood CD4 Treg cells increased in frequency after belumosudil therapy. (A-B) Percentages of immune cells (CD3 T, CD4 T, CD8 T, NK, CD8 Treg cells, gdT, NKT, MAIT, ILCs, IFN-γ) in PBMCs before and after treatment with belumosudil measured by spectral flow cytometry. (C) Percentage of IL-17 before and after treatment with belumosudil from PBMCs (n = 10). (D) Gating strategy used for CD4 Treg cells (CD3⁺CD4⁺FOXP3⁺). (E) Percentage of CD4 Treg cells before and after treatment with belumosudil from PBMCs (n = 10). (F) Quantification of CD4 Treg cells in the OM. (G) Scatterplot showing significant positive correlation between CD4 Treg cells and macrophages in the OM. (H) Representative multiplex immunofluorescence staining of DAPI (blue), CD3 (red), CD4 (green), and FOXP3 (white) in labial MSGs. (I) Quantification of CD4 Treg cells and ratios to other T-cell populations in the MSGs. (J) Quantification of CD4 Treg cells in the skin. P value <.05 is considered significant. Wilcoxon signed-rank test was performed. Scale bar of 50 μm was used. FOXP3, forkhead box P3; gdT, γδ T cells; ILC, innate lymphoid cell; MAIT, mucosal-associated invariant T cell.