The proliferation and migration features of Lgr5+ stem cells and transit amplifying cells in the intestine are dynamically changed during the postnatal development
Authors: Geng Hua , Subramanian Saravanan , Bu Heng-Fu , Tan Xiao-Di 
Published: May 2023

Intestines continue to develop after birth, and the early neonatal period is a critical phase of gastrointestinal (GI) maturation. The intestinal epithelium takes a central role in mucosal physiological functions, and it experiences renewal ability throughout life. Lgr5+ crypt base columnar stem cells (Lgr5+-CBC) and transit amplifying cells (TACs) are proliferative cells, and their physiological features can be affected by inflammation, microbiome, and environmental exposure. However, it is unclear whether and how the activity of Lgr5+-CBCs and TACs changes during postnatal development. In this study, we aimed to advance our knowledge in this aspect. Lgr5EGFP-IRES-creERT2 mice at different developmental stages including fetuses, postnatal (P) pups, and adults were subjected to BrdU/EdU dual-labeling to determine S-phase proliferative cells in the crypts and to monitor intestinal epithelial cell (IEC) migration along the crypt-villus axis for up to 72 hours. We used immunohistochemistry, click chemistry, and multi-panel RNAscope FISH to determine EdU+, BrdU+, Lgr5 +, Prom1 +, and Pcna + cells in the intestinal epithelium. We also examined the expression of multiple progenitor markers during postnatal development in the intestine using qRT-PCR. In parallel, GFP+ IECs were collected using fluorescence-activated cell sorting (FACS) and processed for qRT-PCR to determine Lgr5 expression. We found that crypt containing Lgr5+-CBC and well-organized TAC zone with EdU+ and/or Pcna + cells started to develop after birth (P0) and well-formed adult-like crypt structure at P14 in mouse pups. After this time point, the arrangement and number of Lgr5+ CBC per crypt became stable throughout adulthood. Meanwhile, these proliferative IECs were lack of this pattern during the fetal and early postnatal periods, although they were restricted to the base of the villi. Our qRT-PCR analysis showed altered expression of Lgr5 and other fetal progenitor markers (i.e. Olfm4, Axin2, Ascl2, Prom1, and Hopx) during postnatal development in the intestine. Interestingly, qRT-PCR analysis of FACS-sorted GFP+ cells (P0, P7, P14, and adult) revealed that Lgr5 expression was significantly higher in P0, suggesting CBC stem cells possess more active stemness after birth. In combined BrdU staining and EdU pulse-chase assay, we found that IEC migration was halted at P0 and P1, in which BrdU label-retaining cells resided within the crypt. Among them, a group of BrdU+/EdU+ cells showed active proliferative potential in 24 hours. In contrast, IEC migration was detected in P2, in which BrdU+ cells migrated along the crypt-villi axis, and only a few portions of BrdU label-retaining cells resided within the crypt and maintained EdU+ active proliferation. Collectively, our data indicated that the activity of Lgr5+-CBC and TACs are dynamically changed during postnatal GI development. The mechanisms underlying these alterations need further study.

This work is supported by grants from NIH and U.S. Department of Veterans Affairs.

This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.