Figure 5.
VIP synthesis by donor pDCs limits GVHD in allo-HSCT recipients. (A-B) Percentage survival of B10.BR mice transplanted with 5 × 103 FACS-sorted HSCs and 1 × 106 MACS-enriched splenic T cells from C57BL/6J mice with the addition of 5 × 104 FACS-purified pDCs from either wild-type or VIP-KO mice. Survival and mean GVHD score of each group are shown. Biological replicates involved 18 independent experiments with n = 83 mice in wild-type pDCs, HSC, and 1MT group; n = 41 mice in the VIP-KO pDCs, HSC, and 1 × 106 group; n = 61 in the HSC and 1 × 106 T-cell group. (C-D) Survival and mean GVHD scores of mice that received 5 ×103 HSCs, 2 × 106 splenic T cells, and 5 × 104 FACS-purified wild-type or VIP-KO pDCs. N = 16 mice per group. (E) 7 days after B10.BR mice received 1 × 106 T cells, mice were euthanized, and jejunum, duodenum, and ileum were harvested, formalin-fixed, paraffin-embedded, 10-μm sections stained with hematoxylin and eosin, and histological GVHD scores were determined by a pathologist masked to treatment group. Bar indicates 50 μm at ×200 magnification. (F) Each dot represents a segment of small intestine. Four biological replicates and 12 technical replicates were analyzed for each group. Bar indicates 50 μm at ×200 magnification. (G-H) Histology score of colons in recipients of 1 × 106 T cells of recipients. Bar indicates 50 μm at ×200 magnification. (I-J) Seven days after the B10.BR mice received 2 × 106 T cells, colons were collected and stained with hematoxylin and eosin. Apoptotic crypts are counted by a pathologist. N = 8 in each group. Bar indicates 100 μm at ×40 magnification. (K-L) Seven days after allo-HSCT, formalin-fixed, paraffin-embedded sections of small intestine from recipients of 2 × 106 T cells were stained with periodic acid–Schiff, and goblet cells were counted by a pathologist (masked to treatment group), n = 6 or 7 biological replicates for each group. Bar indicates 50 μm at ×200 magnification. Statistics: Kaplan-Meier for survival analysis, two-way analysis of variance for clinical GVHD, nonparametric Mann-Whitney U test for histological GVHD scores, and crypts apoptosis. Yellow arrow marks necrotic cell; yellow asterisks marks neutrophils. *P < .05, **P < .01, ***P < .001, ****P < .0001.

VIP synthesis by donor pDCs limits GVHD in allo-HSCT recipients. (A-B) Percentage survival of B10.BR mice transplanted with 5 × 103 FACS-sorted HSCs and 1 × 106 MACS-enriched splenic T cells from C57BL/6J mice with the addition of 5 × 104 FACS-purified pDCs from either wild-type or VIP-KO mice. Survival and mean GVHD score of each group are shown. Biological replicates involved 18 independent experiments with n = 83 mice in wild-type pDCs, HSC, and 1MT group; n = 41 mice in the VIP-KO pDCs, HSC, and 1 × 106 group; n = 61 in the HSC and 1 × 106 T-cell group. (C-D) Survival and mean GVHD scores of mice that received 5 ×103 HSCs, 2 × 106 splenic T cells, and 5 × 104 FACS-purified wild-type or VIP-KO pDCs. N = 16 mice per group. (E) 7 days after B10.BR mice received 1 × 106 T cells, mice were euthanized, and jejunum, duodenum, and ileum were harvested, formalin-fixed, paraffin-embedded, 10-μm sections stained with hematoxylin and eosin, and histological GVHD scores were determined by a pathologist masked to treatment group. Bar indicates 50 μm at ×200 magnification. (F) Each dot represents a segment of small intestine. Four biological replicates and 12 technical replicates were analyzed for each group. Bar indicates 50 μm at ×200 magnification. (G-H) Histology score of colons in recipients of 1 × 106 T cells of recipients. Bar indicates 50 μm at ×200 magnification. (I-J) Seven days after the B10.BR mice received 2 × 106 T cells, colons were collected and stained with hematoxylin and eosin. Apoptotic crypts are counted by a pathologist. N = 8 in each group. Bar indicates 100 μm at ×40 magnification. (K-L) Seven days after allo-HSCT, formalin-fixed, paraffin-embedded sections of small intestine from recipients of 2 × 106 T cells were stained with periodic acid–Schiff, and goblet cells were counted by a pathologist (masked to treatment group), n = 6 or 7 biological replicates for each group. Bar indicates 50 μm at ×200 magnification. Statistics: Kaplan-Meier for survival analysis, two-way analysis of variance for clinical GVHD, nonparametric Mann-Whitney U test for histological GVHD scores, and crypts apoptosis. Yellow arrow marks necrotic cell; yellow asterisks marks neutrophils. *P < .05, **P < .01, ***P < .001, ****P < .0001.

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