Fig. 4.

In-ovo feeding (IOF) shifts pre-hatch stem cell localizations toward villus bottoms. (A, B) Representative images of RNAscope in-situ hybridization (ISH) of stem cell marker leucine-rich repeat containing G-protein coupled receptor 5 (Lgr5) at embryonic day 19 (E 19) in the bottom (B), middle (M) and top (T) regions of each villus (separated by dotted lines) in control (non-injected) (A) and IOF-Gln embryos (B). Arrowheads indicate Lgr5 punctate expression within each region. Lgr5 probes were visualized with fast-red and tissues were counterstained with hematoxylin. Images were captured at X200 magnifications. Scale bars, 50 μm. (C) Percentage of Lgr5 punctate expression in each villus region, relative to total Lgr5 expression in the entire villus. Values are means ± SEM. Different uppercase letters mark significant differences between age/treatment group in each villus region by Tukey–Kramer HSD, P < 0.05.