Unbiased molecular definition of epithelial barrier formation and defects driving inflammatory bowel disease

The intestinal epithelial barrier is one of the body’s largest mucosal surfaces. The cells involved reflect the numerous functions the epithelium must perform. The barrier requires time critical co-ordination with other intestinal cell compartments in utero for normal development. In maturity, dysregulation of the barrier or cross-talk can lead to disease such as inflammatory bowel disease (IBD). Despite the importance of the intestinal epithelium in development and health, characterisation of the origins of dysregulation are lacking. Questions also remain about the full spectrum of epithelial cell diversity and its mutualistic relationship with other intestinal compartments. This project characterises the development of the epithelial and mesenchymal cross talk in utero, in health and in IBD at high resolution using single cell RNA-sequencing (scRNAseq). Studying intestinal epithelial diversity in health and disease, isolating epithelial cells from IBD and controls and mapping over 11,000 cells from health and UC inflammation, mapped cell diversity through the maturation of epithelium. This identified a hitherto unappreciated BEST4/OTOP2 cell that sensed pH and was dysregulated in inflammation and cancer. Furthermore, a second project mapped human intestinal development from 8-22 post conceptual weeks. This charted 101 cell types across developmental time and through spatial transcriptomics (ST) could map these to tissue revealing origins of diverse cellular compartments along with fibroblast and intestinal stem cells across space and time. These results provide a platform that maps the human intestinal epithelium and previously unappreciated resolution and from which the normal developmental cues can be established and allow identification of the drivers of dysregulation in IBD.</p

Spatiotemporal Analysis of Human Intestinal Development at Single Cell Resolution: Supplementary Data

Supplementary data for the study "Spatiotemporal Analysis of Human Intestinal Development at Single Cell Resolution" which utilises single cell RNA-sequencing (scRNA-seq) and spatial transcriptomics (ST) to characterise the origins of human intestinal development between 8 and 22 post conceptual weeks. This supplementary data contains analysed sequencing data that was too large for main publications and details data that was used for comparisons of: cell type marker genes; ST varying genes; Transcription factor (TF) module nodes; Human Phenotype Ontology (HPO) disease comparisons across time and cell clusters; cell cluster comparisons and Gene Ontology Analysis. Specific content: A) An excel file with tabs of supplementary data (overview in first tab) including: 1 Sample overview 2 Endothelial Markers 3 Epithelial Markers 4 Fibroblast Markers 5 Immune Markers 6 Muscle Markers 7 Myofibroblast & Mesothelium markers 8 Neural markers 9 Pericyte Markers 10 Secretory epithelium markers 11 Compartment Markers 12 ST Distance varying genes 13 TF Tree Node Markers 14 TF Location Time Compartment 15 HPO Genes Time Varying 16 HPO Genes Cluster Specific 17 S2 Colon vs TI 18 S2 Time Course 19 Stem Cells TI vs Colon 20 Stem cells vs Stem progenitor 21 Stromal 4 differences 22 CITE-Seq Antibody Sequences 23 Neural GO BP 24 Morphogen modules 25 GO Terms ST dist genes B)Images of ST segments (n=8 tissue segments with low resolution and high resolution images of each

Unbiased molecular definition of epithelial barrier formation and defects driving inflammatory bowel disease (DPhil Thesis D Fawkner-Corbett)

The appendix and supplementary data to support the thesis "Unbiased molecular definition of epithelial barrier formation and defects driving inflammatory bowel disease" submitted by D Fawkner-Corbett, Balliol College, University of Oxford

Spatiotemporal Analysis of Human Intestinal Development at Single Cell Resolution: Supplementary Data

Supplementary data for the study "Spatiotemporal Analysis of Human Intestinal Development at Single Cell Resolution" which utilises single cell RNA-sequencing (scRNA-seq) and spatial transcriptomics (ST) to characterise the origins of human intestinal development between 8 and 22 post conceptual weeks. This supplementary data contains analysed sequencing data that was too large for main publications and details data that was used for comparisons of: cell type marker genes; ST varying genes; Transcription factor (TF) module nodes; Human Phenotype Ontology (HPO) disease comparisons across time and cell clusters; cell cluster comparisons and Gene Ontology Analysis. Specific content: A) An excel file with tabs of supplementary data (overview in first tab) including: 1 Sample overview 2 Endothelial Markers 3 Epithelial Markers 4 Fibroblast Markers 5 Immune Markers 6 Muscle Markers 7 Myofibroblast & Mesothelium markers 8 Neural markers 9 Pericyte Markers 10 Secretory epithelium markers 11 Compartment Markers 12 ST Distance varying genes 13 TF Tree Node Markers 14 TF Location Time Compartment 15 HPO Genes Time Varying 16 HPO Genes Cluster Specific 17 S2 Colon vs TI 18 S2 Time Course 19 Stem Cells TI vs Colon 20 Stem cells vs Stem progenitor 21 Stromal 4 differences 22 CITE-Seq Antibody Sequences 23 Neural GO BP 24 Morphogen modules 25 GO Terms ST dist genes B)Images of ST segments (n=8 tissue segments with low resolution and high resolution images of each

Spatiotemporal Analysis of Human Intestinal Development at Single Cell Resolution: Supplementary Data

Supplementary data for the study "Spatiotemporal Analysis of Human Intestinal Development at Single Cell Resolution" which utilises single cell RNA-sequencing (scRNA-seq) and spatial transcriptomics (ST) to characterise the origins of human intestinal development between 8 and 22 post conceptual weeks. This supplementary data contains analysed sequencing data that was too large for main publications and details data that was used for comparisons of: cell type marker genes; ST varying genes; Transcription factor (TF) module nodes; Human Phenotype Ontology (HPO) disease comparisons across time and cell clusters; cell cluster comparisons and Gene Ontology Analysis. Specific content: A) An excel file with tabs of supplementary data (overview in first tab) including: 1 Sample overview 2 Endothelial Markers 3 Epithelial Markers 4 Fibroblast Markers 5 Immune Markers 6 Muscle Markers 7 Myofibroblast & Mesothelium markers 8 Neural markers 9 Pericyte Markers 10 Secretory epithelium markers 11 Compartment Markers 12 ST Distance varying genes 13 TF Tree Node Markers 14 TF Location Time Compartment 15 HPO Genes Time Varying 16 HPO Genes Cluster Specific 17 S2 Colon vs TI 18 S2 Time Course 19 Stem Cells TI vs Colon 20 Stem cells vs Stem progenitor 21 Stromal 4 differences 22 CITE-Seq Antibody Sequences 23 Neural GO BP 24 Morphogen modules 25 GO Terms ST dist genes B)Images of ST segments (n=8 tissue segments with low resolution and high resolution images of each

Multicentre observational study of adherence to Sepsis Six guidelines in emergency general surgery

BACKGROUND:Evidence-based interventions may reduce mortality in surgical patients. This study documented the prevalence of sepsis, adherence to guidelines in its management, and timing of source control in general surgical patients presenting as an emergency. METHODS:Patients aged 16 years or more presenting with emergency general surgery problems were identified over a 7-day period and then screened for sepsis compliance (using the Sepsis Six standards, devised for severe sepsis) and the timing of source control (whether radiological or surgical). Exploratory analyses examined associations between the mode (emergency department or general practitioner) and time of admission, adherence to the sepsis guidelines, and outcomes (complications or death within 30 days). RESULTS:Of a total of 5067 patients from 97 hospitals across the UK, 911 (18·0 per cent) fulfilled the criteria for sepsis, 165 (3·3 per cent) for severe sepsis and 24 (0·5 per cent) for septic shock. Timely delivery of all Sepsis Six guidelines for patients with severe sepsis was achieved in four patients. For patients with severe sepsis, 17·6-94·5 per cent of individual guidelines within the Sepsis Six were delivered. Oxygen was the criterion most likely to be missed, followed by blood cultures in all sepsis severity categories. Surgery for source control occurred a median of 19·8 (i.q.r. 10·0-35·4) h after diagnosis. Omission of Sepsis Six parameters did not appear to be associated with an increase in morbidity or mortality. CONCLUSION:Although sepsis was common in general surgical patients presenting as an emergency, adherence to severe sepsis guidelines was incomplete in the majority. Despite this, no evidence of harm was apparent

Systematic review and meta-analysis comparing outcomes following orchidopexy for cryptorchidism before or after 1 year of age

Background Current guidelines recommend orchidopexy for cryptorchidism by 12 months of age, yet this is not universally adhered to. The aim of this systematic review and meta-analysis was to compare outcomes between orchidopexies performed before and after 1 year of age. Methods MEDLINE and Embase were searched (September 2015) using terms relating to cryptorchidism, orchidopexy and the outcomes of interest. Studies were eligible for inclusion if they compared orchidopexy at less than 1 year of age (early) with orchidopexy at 1 year or more of age (delayed) and reported the primary outcome (testicular atrophy) or one of the secondary outcomes (fertility potential, postoperative complication, malignancy). Studies were excluded when more than 50 per cent of infants had intra-abdominal testes, or the population included infants with disorders of sexual differentiation. Additional studies were identified through reference list searching. Unpublished data were sought from the ORCHESTRA study investigators. Results Fifteen eligible studies were identified from 1387 titles. There was no difference in atrophy rate between early orchidopexy and delayed orchidopexy (risk ratio 0⋅64, 95 per cent c.i. 0⋅25 to 1⋅66; 912 testes). Testicular volume was greater (mean difference 0⋅06 (95 per cent c.i. 0⋅01 to 0⋅10) ml; 346 testes) and there were more spermatogonia per tubule (mean difference 0⋅47 (0⋅31 to 0⋅64); 382 testes) in infants undergoing early orchidopexy, with no difference in complication rate (risk ratio 0⋅68, 0⋅27 to 1⋅68; 426 testes). No study reported malignancy rate. Conclusions Atrophy and complication rates do not appear different between early and delayed orchidopexy, and fertility potential may be better with early orchidopexy. Imprecision of the available data limits the robustness of these conclusions.</p

Enteric nervous system stem cells associated with thickened extrinsic fibers in short segment aganglionic Hirschsprung's disease gut are absent in the total colonic and intestinal variants of disease

Despite current treatments patients with Hirschsprung's disease (HSCR) suffer significant long-term morbidity. Therefore, there is increasing interest in adjunctive therapies, such as using enteric nervous system stem cells (ENSSC), isolated from typical aganglionic bowel. The source of these cells is unclear however it is hypothesized that they are present in the thickened nerve trunks in aganglionic short and long segment HSCR gut. These cells should therefore be absent in total colonic and pan intestinal HSCR where these thickened fibers are absent.Cells were isolated from samples of short segment HSCR gut (n=18) and total colonic and total intestinal HSCR gut (n=2). Acetylcholinesterase histochemistry confirmed the presence/absence of thickened nerve trunks. P75 immunofluorescence highlighted ENSSC at isolation and after 10days in culture in both groups.ENSSC were not isolated or cultured from total colonic and total intestinal HSCR gut where thickened nerve trunks were absent. In contrast 10.0% (+/-1.9 SEM) of cells from short segment HSCR gut were ENSSC at isolation rising to 22.7% (+/-2.9 SEM) after 10days in culture.These results associate ENSCC with thickened nerve trunks and also suggest that the aganglionic bowel segment in total colonic and intestinal HSCR cannot be used as a source of ENSCC for adjunctive therapy

Enteric nervous system stem cells associated with thickened extrinsic fibers in short segment aganglionic Hirschsprung's disease gut are absent in the total colonic and intestinal variants of disease

Despite current treatments patients with Hirschsprung's disease (HSCR) suffer significant long-term morbidity. Therefore, there is increasing interest in adjunctive therapies, such as using enteric nervous system stem cells (ENSSC), isolated from typical aganglionic bowel. The source of these cells is unclear however it is hypothesized that they are present in the thickened nerve trunks in aganglionic short and long segment HSCR gut. These cells should therefore be absent in total colonic and pan intestinal HSCR where these thickened fibers are absent.Cells were isolated from samples of short segment HSCR gut (n=18) and total colonic and total intestinal HSCR gut (n=2). Acetylcholinesterase histochemistry confirmed the presence/absence of thickened nerve trunks. P75 immunofluorescence highlighted ENSSC at isolation and after 10days in culture in both groups.ENSSC were not isolated or cultured from total colonic and total intestinal HSCR gut where thickened nerve trunks were absent. In contrast 10.0% (+/-1.9 SEM) of cells from short segment HSCR gut were ENSSC at isolation rising to 22.7% (+/-2.9 SEM) after 10days in culture.These results associate ENSCC with thickened nerve trunks and also suggest that the aganglionic bowel segment in total colonic and intestinal HSCR cannot be used as a source of ENSCC for adjunctive therapy