Division of Bone Marrow Transplant and Immune Deficiency, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229; Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati OH 45229.
Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati OH 45229; Division of Gastroenterology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229; Sheba Medical Center, Hashomer, affiliated with the Aviv University, Israel 52620.
Haematologica. 2023 Jul 1;108(7):1803-1816. doi: 10.3324/haematol.2022.282035.
We performed transcriptomic analyses on freshly frozen (n=21) and paraffin-embedded (n=35) gastrointestinal (GI) biopsies from children with and without acute acute GI graft-versus-host disease (GvHD) to study differential gene expressions. We identified 164 significant genes, 141 upregulated and 23 downregulated, in acute GvHD from freshy frozen biopsies. CHI3L1 was the top differentially expressed gene in acute GvHD, involved in macrophage recruitment and bacterial adhesion. Mitochondrial genes were among the top downregulated genes. Immune deconvolution identified a macrophage cellular signature. Weighted gene co-expression network analysis showed enrichment of genes in the ERK1/2 cascade. Transcriptome data from 206 ulcerative colitis (UC) patients were included to uncover genes and pathways shared between GvHD and UC. Comparison with the UC transcriptome showed both shared and distinct pathways. Both UC and GvHD transcriptomes shared an innate antimicrobial signature and FCγ1RA/CD64 was upregulated in both acute GvHD (log-fold increase 1.7, P=0.001) and UC. Upregulation of the ERK1/2 cascade pathway was specific to GvHD. We performed additional experiments to confirm transcriptomics. Firstly, we examined phosphorylation of ERK (pERK) by immunohistochemistry on GI biopsies (acute GvHD n=10, no GvHD n=10). pERK staining was increased in acute GvHD biopsies compared to biopsies without acute GvHD (P=0.001). Secondly, plasma CD64, measured by enzyme-linked immunsorbant assay (n=85) was elevated in acute GI GvHD (P<0.001) compared with those without and was elevated in GVHD compared with inflammatory bowel disease (n=47) (P<0.001), confirming the upregulated expression seen in the transcriptome.
我们对新鲜冷冻（n=21）和石蜡包埋（n=35）的胃肠道（GI）活检组织进行了转录组分析，这些活检组织来自患有和不患有急性 GI 移植物抗宿主病（GvHD）的儿童，以研究差异基因表达。我们在新鲜冷冻活检组织的急性 GvHD 中鉴定出 164 个显著差异表达的基因，其中 141 个上调，23 个下调。CHI3L1 是急性 GvHD 中表达差异最大的基因，参与巨噬细胞募集和细菌黏附。下调基因中包括线粒体基因。免疫去卷积鉴定出巨噬细胞细胞特征。加权基因共表达网络分析显示 ERK1/2 级联相关基因富集。纳入 206 例溃疡性结肠炎（UC）患者的转录组数据，以揭示 GvHD 和 UC 之间共享的基因和途径。与 UC 转录组比较，发现既有共同途径，也有独特途径。UC 和 GvHD 转录组均具有固有抗菌特征，且急性 GvHD（log 倍增加 1.7，P=0.001）和 UC 中 FCγ1RA/CD64 均上调。ERK1/2 级联途径的上调仅见于 GvHD。我们进行了额外的实验来验证转录组结果。首先，我们通过免疫组化在 GI 活检组织（急性 GvHD n=10，无 GvHD n=10）上检测 ERK 的磷酸化（pERK）。与无急性 GvHD 的活检组织相比，急性 GvHD 活检组织中的 pERK 染色增加（P=0.001）。其次，通过酶联免疫吸附试验（n=85）测量的血浆 CD64 水平在急性 GI GvHD 中升高（P<0.001），与无 GvHD 相比升高，与炎症性肠病（n=47）相比升高（P<0.001），证实了转录组中观察到的上调表达。
