ROCK2 inhibition with belumosudil reduces collagen, T cells, and IL-17–producing cells in the oral buccal mucosa. (A) Representative PSR (picrosirius red) staining paired with polarized microscopy of OM sections before and after treatment with belumosudil showing changes in collagen type I (red) and collagen type III (green). White dashed line denotes epithelial-submucosal border. (B) Changes in percentage area of collagen types I and III in the OM (n = 16; paired parametric t test). (C) Representative multiplex immunofluorescence staining for DAPI (blue), CD3 (cyan), CD4 (red), and IL-17 (green) in the OM before and after treatment with belumosudil with close-up view of marked region in yellow. (D) Changes in CD3⁺ T, CD4⁺, IL-17⁺, CD3⁺CD4⁺ T, CD3⁺CD4⁻ (CD8) T, Th17, CD3⁻CD4⁺, and CD3⁻ IL-17⁺ cells after belumosudil treatment (n = 14; Wilcoxon signed-rank test). (E) Representative multiplex immunofluorescence staining for DAPI (blue), Vim (green), CD68 (red), α-SMA (magenta) in the OM before and after treatment with belumosudil with close-up view of marked region in yellow. (F) Changes in macrophages (CD68⁺) and activated macrophages (CD68⁺Vim⁺ cells) after belumosudil treatment (n = 8; Wilcoxon signed-rank test). (G) Scatterplots showing negative correlation between macrophages and activated macrophages with collagen I. P value <.05 is considered significant. Scale bar of 100 μm was used. α-SMA, α-smooth muscle actin; DAPI, 4′,6-diamidino-2-phenylindole; Vim, vimentin.