https://phaidra.vetmeduni.ac.at/o:2675 The Intricacies of Inflammatory Bowel Disease: A Preliminary Study of Redox Biology in Intestinal Organoids Csukovich, Georg Georg Csukovich 0000-0001-5615-666X University of Veterinary Medicine Vienna Burgener, Iwan Anton Iwan Anton Burgener 0000-0002-8516-0040 University of Veterinary Medicine Vienna Pratscher, Barbara Barbara Pratscher 0000-0001-5291-1365 University of Veterinary Medicine Vienna Huainig, Janina Janina Huainig University of Veterinary Medicine Vienna Troester, Selina Selina Troester 0000-0001-6268-4442 University of Veterinary Medicine Vienna MDPI text eng We evaluated the redox status, precisely glutathione levels, which have a major impact in cellular detoxification and antioxidant defence in IBD-derived and healthy intestinal organoids. Therefore, we wanted to explore the differences in terms of their redox balance and mitochondrial fitness. To this end, we introduced a Grx1-roGFP2 construct into the organoids by lentiviral transduction before performing a stress assay by treating the organoids with hydrogen peroxide and examined the GSH/GSSG ratio using confocal imaging. Using ratio imaging, we could detect statistically significant differences between healthy and IBD-derived samples. To gain more insight, we also performed a GSH/GSSG assay, which directly measured glutathione levels. This analysis revealed that both organoid lines had higher levels of oxidized glutathione due to the stress treatment demonstrated by a lower GSH/GSSG ratio compared to the untreated control. Nevertheless, the results showed no significant difference between healthy and IBD-derived organoids. We further challenged organoids with hydrogen peroxide after incubation with MitoTracker® to see if mitochondrial fitness might be different in IBD-derived organoids. However, these results were also very comparable. In summary, our preliminary findings indicate that both organoid lines demonstrate a well-functioning system in terms of analysis but show no clear difference between healthy and IBD-derived samples. intestinal organoid; redox biology; IBD; glutathione; oxidative stress; ROS; redox imaging http://creativecommons.org/licenses/by/4.0/ https://phaidra.vetmeduni.ac.at/api/object/o:2675/download 10.3390/organoids2030012 https://phaidra.vetmeduni.ac.at/o:605 application/pdf Organoids 2(3) (2023) Organoids 2 3 437.36 kB 2023