Fig. 1: Immune-mediated GI damage impacts the transcriptome of intestinal stem cells, upregulating STAT1 in GVHD.

a–d, f–h Single cell RNA sequencing (scRNA-seq) of freshly isolated ileal crypt cells from healthy B6 mice (Naive), B6-into-B6 syngeneic BMT recipients without GVHD (Syn), or B10.BR-into-B6 MHC-mismatched allogeneic BMT recipients with GVHD (Allo) 5 days after transplant. a Study design: Prior to scRNA-seq, BMT recipients were transplanted with both marrow and purified donor T cells following pre-transplant conditioning (TBI; 5.5 Gy x 2). b UMAP plots indicating the experimental conditions for sequenced cells (left) and distinct clusters with their cell type annotation (right). Star symbol indicates ISC clusters. c UMAP plots highlighting gene expression signatures corresponding to each cell type. d Phenotypic distances among epithelial populations. Heatmap shows average phenotypic distances between experimental conditions; histogram shows the distribution of phenotypic distances between non-GVHD and GVHD conditions (syn-BMT and allo-BMT) in units of log-normalized gene expression. e Phenotypic distances among epithelial populations from SI crypt scRNA-seq after LP-into-B6 MHC-matched (minor-antigen-mismatched) allo-BMT in units of log-normalized gene expression. Heatmap shows each population’s average phenotypic distance between homeostatic (Naive), non-GVHD T-cell-depleted transplant (BM), and GVHD-inducing BM + T cell transplant (BM + T); histogram shows the distribution of phenotypic distances between non-GVHD and GVHD transplant conditions (BM vs. BM + T). f GSEA comparing ISCs from syn-BMT and MHC-mismatched allo-BMT; NES: normalized enrichment score. g Volcano plot showing transcription factor genes differentially expressed (computed using MAST) in ISCs following syn-BMT and allo-BMT. Positive log fold change indicates increased expression after allo-BMT. h Expression of Stat1 in crypt epithelial populations highlighting upregulation in ISCs after allo-BMT; gene expression imputed using MAGIC (n = 3685 Naive, 3371 Syngeneic, 4670 Allogeneic cells); E enterocytes, EEC enteroendocrine cells, G goblet cells, I ISCs, P Paneth cells, T tuft cells. The boxplots represent three quartiles, and the whiskers indicate 1.5 times the interquartile range.