Different patterns of immune cell activation and tissue destruction in patients with steroid-refractory aGVHD responding differently to MSC therapy. (A) Graphical depiction of the appearance of characteristic immune populations and degree of epithelial cell damage in patients with aGVHD either responding (top) or refractory (bottom) to second-line immunosuppressive therapy. Both patient groups were treated consecutively with first-line immunosuppressive and second-line MSC therapy. HSCT patients with steroid-refractory aGVHD either responsive or nonresponsive to MSC-based second-line therapy groups (GVHD-CR and GVHD-NR, respectively) initially showed high frequencies of circulating CD163⁺CD11b⁺ monocytes and CXCR3⁺CCR9⁺CCR10⁺ effector T cells shortly after introduction of immunosuppressive therapy. Patients nonresponsive to MSC-based second-line therapy further showed increased frequencies of CD163⁺CD11b^– DCs, CD56⁺ DCs, and plasmablasts, which persisted over time. In patients responsive to MSC-based second-line therapy, who showed complete resolution of all clinical aGVHD symptoms, CXCR3⁺CCR9⁺CCR10⁺ effector T cells, along with CXCR3⁺CCR9⁺CCR10⁺ Tregs, decreased over time. These populations remained high in the group of patients nonresponsive to MSC-based second-line therapy, indicative of escalating immune reactivity leading to progressive tissue damage in aGVHD target organs, loss of epithelial barrier function, and concurrent infectious complications. (B) Endoscopy image showing the macroscopic appearance of the colon of one of the patients nonresponsive to MSC-based second-line therapy. The biopsy was taken after MSC therapy was initiated.