The exacerbation of GVHD in Siglec-G–deficient hosts is dependent on the intensity of conditioning in experimental MHC-mismatched allo-HCT. (A) Siglec-G expression on splenic DCs tested by FACS with anti–Siglec-G antibody in naïve mouse and at 24 hours after TBI; the representative histogram (upper) and summarized data from 3 experiments (lower) (n = 2 to 4 per group). (B) The absence of Siglec-G decreased phosphorylated and total of SHP-1. Immunoblot analysis of phosphorylated(p)- and total SHP-1 protein in lysates of splenocytes from WT-B6 or Siglec-G^−/− animals after irradiation. The bar graph shows the ratio pSHP-1/total SHP-1. (C-H) Siglec-G deficiency exacerbates GVHD. WT-B6 and Siglec-G^−/− mice were lethally irradiated with 13 Gy and infused with 2 to 2.5 × 10⁶ CD90⁺ T cells along with 5 × 10⁶ TCD-BM cells from either syngeneic B6 or allogeneic MHC-mismatched BALB/c donors. (C) Survival following 13 Gy TBI (n = 6 to 11 per group, pooled from 2 experiments); P = .007 when B6 vs Siglec-G^−/− B6 mice were used as recipients. (D) Histopathologic analysis of GI tracts (small and large intestine) by hematoxylin and eosin stain (left) and GVHD score on days 7 and 14 after allo-HCT (n = 3 to 6 per group, pooled from 2 experiments). Original magnification ×200. (E) Donor T-cell (H-2k^d+CD4⁺CD8⁺) expansion in the spleen on days 7 and 14 after allo-HCT (n = 4 to 6 per group, pooled from 2 experiments). (F-H) Serum HMGB-1, IFN-γ, and TNF-α levels on day 7 after allo-HCT (n = 3 to 6 per group, pooled from 2 experiments). The bar shows the mean ± standard error of the mean (SEM).