MSC treatment mediated immune regulation. (A-D) Tregs and related cytokines. (A) CD25 and FoxP3 in CD4 T cells of splenocytes in ICR control mice (ICR), NOD/Ltj untreated mice (Untreated), BALB/c BMMSC-infused NOD/Ltj mice (MSC Treated), and CXCR4-blocked BALB/c BMMSC-infused NOD/Ltj mice (C-b MSC Treated). (B) Tregs in spleen of NOD/Ltj untreated mice were far less than control ICR mice (P = 2.597 × 10−7, n = 6), and allogeneic BMMSC partly restored Treg cells (P = 4.790 × 10−9, n = 6), but CXCR4 blocking reduced MSC-mediated Treg generation compared with normal BMMSCs (P = 2.397 × 10−6, n = 6). (E-J) Th1, Th2, and related cytokines, intracellular staining for IFN-γ and IL-4 in CD4+ splenocytes. (E) Higher numbers of Th1 cells were observed in NOD/Ltj untreated mice than control ICR mice (F; P = 7.518 × 10−5, n = 6), and there was no change of Th1 response 1 week after allogeneic BMMSC or CXCR4-block BMMSC infusion. Th2 responses in the spleen of NOD/Ltj untreated mice were less than that of control ICR mice (G; P = 6.917 × 10−6, n = 6). Allogeneic BMMSC could partly restore Th2 (P = .0008, n = 6), and blocking of CXCR4 abolished this effect (P = .323, n = 6). (K-O) Th17, Tfh, and related cytokines. Flow cytometry for CXCR5 (Tfh) and IL-17 (Th17) expression in CD4 splenocytes. (K) Th17 were significantly higher in NOD/Ltj untreated mice than in control ICR mice (L; P = .0004, n = 6). Allogeneic BMMSCs mitigated the percentage of Th17 (P = 1.386 × 10−7, n = 6), and CXCR4 blocking resulted in less suppression (P = .025, n = 6). (M) MSC treatment suppressed Tfh response. Allogeneic BMMSC regulated T-cell cytokines IFN-γ (H) and IL-17 (O), regulatory cytokines IL-10 (D), IL-13 (I), and IL-4 (J), and other cytokines TGF-β (C) and IL-6 (N) production in serum and/or salivary gland homogenates. Blocking of CXCR4 of BMMSCs resulted in impairment of immunoregulatory activities. (P-T) Plasma cells and autoantibodies, CD19 and CD138 gated splenocytes in the 4 groups. (P) There was no significant difference of plasma cells in these 4 groups (Q). Both allogeneic BMMSC and CXCR4 blocked BMMSC transplantation decreased the SS-related autoantibodies: antinucleic antibody (R), anti–α-fodrin (S), and anti–SSA/Ro (T) in serum.
