Stromal-Like Cells Are Found in Peripheral Blood of Patients With Inflammatory Bowel Disease and Correlate With Immune Activation State.

INTRODUCTION: Recent studies have identified a critical role of stromal-immune cell interactions in immunity and immune tolerance. Transcriptomic profiling has implicated stromal cells in immune-mediated disorders including the 2 common forms of inflammatory bowel disease (IBD), Crohn's disease (CD), and ulcerative colitis (UC). Stromal-immune interactions may edify inflammatory state and the development of IBD-related complications such as fibrosis, yet the lack of protein markers has hampered studying stromal-immune perturbation. METHODS: In this study, we designed a 40-color spectral flow cytometry assay to characterize hematopoietic and nonhematopoietic cells in intestinal biopsies and matched blood samples from patients with CD or UC. RESULTS: We identified circulating stromal-like cells that are significantly more abundant in IBD blood samples than in healthy controls. Those cells expressed podoplanin (PDPN), a commonly used marker for fibroblasts, and they were associated with activated and memory T and B cells and altered natural killer cell, monocyte, and macrophage populations. PDPN + cells in the blood correlated with PDPN + cells in the colon. Principal component analysis distinctly separated healthy blood samples from IBD blood samples, with stromal-like cells and B-cell subtypes dominating the IBD signature; Pearson correlation detected an association between PDPN + stromal-like cells and B-cell populations in IBD blood and gut biopsies. DISCUSSION: These observations suggest that PDPN + cells in the blood may serve as a biomarker of IBD. Understanding the relationship between stromal cells and immune cells in the intestine and the blood may provide a window into disease pathogenesis and insight into therapeutic targets for IBD.

Lamina Propria Phagocyte Profiling Reveals Targetable Signaling Pathways in Refractory Inflammatory Bowel Disease.

BACKGROUND AND AIMS: Lamina propria phagocytes are key mediators of inflammatory bowel disease (IBD). We aimed to understand the transcriptomic and functional differences in these cells based on location, disease type, inflammation state, and medication use in patients with IBD. METHODS: Phagocytic immune cells in the lamina propria, as defined by the marker CD11b, were isolated from 54 unique patients (n = 111 gut mucosal biopsies). We performed flow cytometry for cell phenotyping (n = 30) and RNA sequencing with differential gene expression analysis (n = 58). We further cultured these cells in vitro and exposed them to janus kinase inhibitors to measure cytokine output (n = 27). Finally, we matched patient genomic data to our RNA sequencing data to perform candidate gene expression quantitative trait locus analysis (n = 34). RESULTS: We found distinct differences in gene expression between CD11b+ cells from the colon vs ileum, as well as in different inflammatory states and, to a lesser degree, IBD types (Crohn's disease or ulcerative colitis). These genes mapped to targetable immune pathways and metabolic and cancer pathways. We further explored the janus kinase-signal transducer and activator of transcription pathway, which was upregulated across many comparisons including in biopsies from anti-tumor necrosis factor refractory patients. We found that isolated CD11b+ cells treated with janus kinase inhibitors had decreased secretion of cytokines tumor necrosis factora and interleukin-8 (P ≤ .05). We also found 3 genetic variants acting as expression quantitative trait loci (P ≤ .1) within our CD11b+ data set. CONCLUSIONS: Lamina propria phagocytes from IBD mucosa provide pathogenetic clues on the nature of treatment refractoriness and inform new targets for therapy.

Genetic dissection of chlorate respiration in Pseudomonas stutzeri†PDA reveals syntrophic (per)chlorate reduction.

Genes important for growth of Pseudomonas stutzeri PDA on chlorate were identified using a randomly DNA bar-coded transposon mutant library. During chlorate reduction, mutations in genes encoding the chlorate reductase clrABC, predicted molybdopterin cofactor chaperon clrD, molybdopterin biosynthesis and two genes of unknown function (clrE, clrF) had fitness defects in pooled mutant assays (Bar-seq). Markerless in-frame deletions confirmed that clrA, clrB and clrC were essential for chlorate reduction, while clrD, clrE and clrF had less severe growth defects. Interestingly, the key detoxification gene cld was essential for chlorate reduction in isogenic pure culture experiments, but showed only minor fitness defects in Bar-seq experiments. We hypothesized this was enabled through chlorite dismutation by the community, as most strains in the Bar-seq library contained an intact cld. In support of this, Δcld grew with wild-type PDA or ΔclrA, and purified Cld also restored growth to the Δcld mutant. Expanding on this, wild-type PDA and a Δcld mutant of the perchlorate reducer Azospira suillum PS grew on perchlorate in co-culture, but not individually. These results demonstrate that co-occurrence of cld and a chloroxyanion reductase within a single organism is not necessary and raises the possibility of syntrophic (per)chlorate respiration in the environment.