Figure 3

Bovine 2D enteroids consist of multi-epithelial cell lineages. A Reverse transcriptase-qPCR gene expression profiles of epithelial cell markers; Mucin2 (Muc2) for Goblet cells, Lysozyme C (Lyz C) for Paneth cells, Chromogranin A (CHGA) for Enteroendocrine cells and Leucine-rich Repeat containing G-protein Receptor 5, (Lgr5) for stem cells in 2D enteroid cultures grown on Matrigel coated 96-well plates in BEM/1% FBS media for 5 days, 3D enteroids cultured in IntestiCult media for 5 days and ileum tissue of 6-month old calves of enteroid origin. Data represents the median and error bars ± 95% confidence interval of four independent experiments. Representative confocal images of 5-day-old 2D enteroids demonstrating the presence of B villin (green) expression in enterocytes and C F-actin (green) filaments, D the bovine epithelial cell marker, cytokeratin 18 (green), and E UEA-1 (green) binding was observed in numerous cells. Nuclei are visualised by Hoechst staining (blue). The 2D enteroids were seeded at 2.5 × 10⁴ cells/well on Matrigel coated 8-well-chamber glass slides in BEM/1% FBS media for 5 days. Data is representative of five independent experiments of five individual calves.