title: Quantitative analysis of mucosal oxygenation using ex vivo imaging of healthy and inflamed mammalian colon tissue.
abstract: Colonic inflammation is associated with decreased tissue oxygenation, significantly affecting gut homeostasis. However, the crosstalk between O2consumption and supply in the inflamed tissue are not fully understood. Using a murine model of colitis, we analysed O2in freshly prepared samples of healthy and inflamed colon tissue. We developed protocols for efficient ex vivo staining of mouse distal colon mucosa with a cell-penetrating O2sensitive probe Pt-Glc and high-resolution imaging of O2concentration in live tissue by confocal phosphorescence lifetime-imaging microscopy (PLIM). Microscopy analysis revealed that Pt-Glc stained mostly the top 50-60 μm layer of the mucosa, with high phosphorescence intensity in epithelial cells. Measured O2values in normal mouse tissue ranged between 5 and 35 μM (4-28 Torr), tending to decrease in the deeper tissue areas. Four-day treatment with dextran sulphate sodium (DSS) triggered colon inflammation, as evidenced by an increase in local IL6 and mKC mRNA levels, but did not affect the gross architecture of colonic epithelium. We further observed an increase in oxygenation, partial activation of hypoxia inducible factor (HIF) 1 signalling, and negative trends in pyruvate dehydrogenase activity and O2consumption rate in the colitis mucosa, suggesting a decrease in mitochondrial respiration, which is known to be regulated via HIF-1 signalling and pyruvate oxidation rate. These results along with efficient staining with Pt-Glc of rat and human colonic mucosa reveal high potential of PLIM platform as a powerful tool for the high-resolution analysis of the intestinal tissue oxygenation in patients with inflammatory bowel disease and other pathologies, affecting tissue respiration.
authors: School of Biochemistry and Cell Biology, University College Cork, Cavanagh Pharmacy Building, College Road, Cork, Ireland. a.zhdanov@ucc.ie.
authors: Alexander V
authors: AV
authors: Zhdanov
authors: School of Biochemistry and Cell Biology, University College Cork, Cavanagh Pharmacy Building, College Road, Cork, Ireland.
authors: Irina A
authors: IA
authors: Okkelman
authors: APC Microbiome Institute, University College Cork, Cork, Ireland.
authors: Anna V
authors: AV
authors: Golubeva
authors: School of Biochemistry and Cell Biology, University College Cork, Cavanagh Pharmacy Building, College Road, Cork, Ireland.
authors: Barbara
authors: B
authors: Doerr
authors: APC Microbiome Institute, University College Cork, Cork, Ireland.|Department of Pharmacology and Therapeutics, University College Cork, Cork, Ireland.
authors: Niall P
authors: NP
authors: Hyland
authors: APC Microbiome Institute, University College Cork, Cork, Ireland.
authors: Silvia
authors: S
authors: Melgar
authors: APC Microbiome Institute, University College Cork, Cork, Ireland.
authors: Fergus
authors: F
authors: Shanahan
authors: APC Microbiome Institute, University College Cork, Cork, Ireland.|Department of Anatomy and Neuroscience, University College Cork, Cork, Ireland.
authors: John F
authors: JF
authors: Cryan
authors: School of Biochemistry and Cell Biology, University College Cork, Cavanagh Pharmacy Building, College Road, Cork, Ireland.
authors: Dmitri B
authors: DB
authors: Papkovsky
mesh_terms: D000818:Animals; D018938:Caco-2 Cells; D003092:Colitis / Q000276:immunology / Q000473:pathology*; D003106:Colon / Q000276:immunology / Q000473:pathology*; D006801:Humans; D007413:Intestinal Mucosa / Q000276:immunology / Q000473:pathology*; D008163:Luminescent Measurements; D008297:Male; D008810:Mice, Inbred C57BL; D018613:Microscopy, Confocal; D061848:Optical Imaging; D010100:Oxygen / Q000032:analysis* / Q000276:immunology; D017207:Rats, Sprague-Dawley; D013194:Staining and Labeling
chemical_list: D010100:Oxygen
keywords: Cell-penetrating phosphorescent probe; Colonic mucosa; FLIM/PLIM; Hypoxia; Intestinal inflammation; Oxygen imaging