Host ASMase regulates graft-versus-host–associated morbidity, mortality, and target organ injury. Lethally irradiated (1100 cGy) C57BL/6asmase+/+ or C57BL/6asmase−/− mice received intravenous injection of LP TCD-BM cells (5 × 106) with or without splenic T cells (3 × 106). (A) Kaplan-Meier survival and (B) clinical GVHD score derived from weekly assessment of 5 clinical parameters (weight loss, hunched posture, decreased activity, fur ruffling, and skin lesions) are shown representing 6 to 8 BM control and 13 or 14 BM + T-cell recipients per group compiled from 2 experiments. Statistical analysis is as follows: (A) BM (asmase+/+ hosts) versus BM + T (asmase+/+ hosts), P < .001; BM + T (asmase+/+ hosts) versus BM + T (asmase−/− hosts), P < .001. (B) BM (asmase+/+ hosts) versus BM + T (asmase+/+ hosts)j P < .005; BM + T (asmase+/+ hosts) versus BM + T (asmase−/− hosts)j P < .005. (C) C57BL/6asmase+/+ or C57BL/6asmase−/− mice received transplants as described in Figure 1 and were killed 21 days thereafter for histopathologic analysis. Representative 5-μm hematoxylin and eosin–stained liver sections reveal increased lymphocyte infiltration (arrows) around the central and portal veins and destruction of hepatic architecture in asmase+/+ hosts receiving allogeneic T cells compared with asmase−/− littermates (left panels). Images were acquired using Zeiss Plan-NEOFLUAR 5×/0.3 numeric aperture (NA) dry lens (Carl Zeiss Inc) and QImaging camera model Retiga EX, and were processed with Improvision Volocity software (PerkinElmer) and Adobe Photoshop Version 7.0 software (Adobe Systems). Right panels reveal higher magnification images of typical endotheliitis observed around a portal vein in asmase+/+ (top panels) but not asmase−/− (bottom panels) hosts. Images were acquired as in left panels, using a Zeiss Plan-NEOFLUAR 40×/1.3 NA oil DIC lens. (D) Representative 5-μm TUNEL-stained sections of proximal jejunum crypts displaying epithelial apoptosis. Images were acquired as in panel C, using a Zeiss Plan-NEOFLUAR 40×/1.3 NA oil DIC lens. Small arrows indicate cells containing condensed or fragmented brown nuclei contrasting with the blue stain of nonapoptotic nuclei, quantified in panel E. Large arrows indicate areas of inflammatory cell infiltration. Crypt apoptosis (E) was scored in 200 crypts per point. Data (mean ± SEM) are collated from 2 experiments. Frequency histograms of apoptotic cells in the villus lamina propria (F) represent data from 150 villae per point, collated from 2 experiments. (G) C57BL/6 recipient hosts received marrow transplants as detailed earlier in the Figure 1 legend, and skin (tongue and ear) was harvested 21 days thereafter. Alternatively, skin was harvested 14 days after transplantation of 10 × 106 TCD-BM cells with or without 0.5 × 106 T cells from B10.BR (H2k) donors. Skin GVHD score was determined by the number of dyskeratotic apoptotic keratinocytes per millimeter of epidermis (mean ± SEM) as assessed in blinded fashion on hematoxylin and eosin–stained sections. Data represent 4 to 14 mice per group compiled from 3 independent experiments.
