Colonic MYO5B Expression Retains Fecal Formation but not Weight Loss in OLFM4+ Stem Cell-Specific MYO5B Deletion in Mice

Background and aim: Microvillus inclusion disease (MVID) is a congenital disorder characterized by severe diarrhea and is caused by inactivating mutations in motor protein myosin Vb (MYO5B). Mice lacking MYO5B in intestinal and colonic epithelia ( Vil-CreERT2;Myo5bfl/fl) display an MVID phenotype characterized by significant weight loss, lack of solid feces, and brush border defects. However, previous models have not discerned if this diarrheal phenotype is due to MYO5B loss in the small intestine specifically or both in the small intestine and colon. Using the small intestinal stem cell specific Cre driver under Olfm4, we phenotyped the mice lacking MYO5B only in the small intestine, while normal MYO5B expression is maintained in the colon. Methods: Olfm4-GFP-CreERT2;R26R-LSL-tdTom and Myo5bfl/fl mice were crossbred to generate Olfm4-GFP-CreERT2;R26R-LSL-tdTom;Myo5bfl/fl (Olfm4DMYO5B) mice. Adult Olfm4DMYO5B mice and control littermates received tamoxifen injection at day 0. GI tissues were collected 5 and 7 days post tamoxifen. Results: Following Cre induction, Olfm4DMYO5B mice exhibited progressive weight loss starting at day 5 and on average lost 20.6% on day 7. At day 7, the small intestine contained clear luminal contents, similar to that of the previous MVID model mice. However, the colon of day 7 Olfm4DMYO5B mice contained small, solid feces. An elongation of PCNA+ crypts was identified on day 5 throughout the small intestine. Villus blunting and microvillus inclusions were prominent in the small intestinal enterocytes on day 7, but not on day 5. The day 7 Olfm4DMYO5B small intestine demonstrated a decrease in tuft cell numbers, suggesting differentiation deficits in the MYO5B-lacking stem cells. However, the colon of Olfm4DMYO5B mice showed intact morphology. Colonic crypt depth and tuft cell numbers at both days 5 and 7 were similar to those of control mice. Immunostaining confirmed the maintenance of MYO5B expression in the colon and progressive loss of MYO5B in the small intestine of Olfm4DMYO5B mice. At day 5, MYO5B was specifically lost in small intestinal crypts, and the loss of MYO5B expanded to the villus tips on day 7. These MYO5B-lacking cells were tdTom+, including enterocytes, goblet, and rare tuft cells. Following the loss of MYO5B, small intestinal enterocytes lacked proper apical localization of numerous transporters and enzymes, including Na+-glucose cotransporter 1 and dipeptidyl peptidase 4. Conclusion: Recently established Olfm4DMYO5B mice demonstrated that the specific MYO5B deletion in the small intestinal stem cells causes hyperproliferation and differentiation deficits. Colonic MYO5B function is suffcient for the formation of solid feces, however, these mice experience prominent weight loss, likely due to the MVID-like malabsorption in the small intestine. NIH R01DK128190, RC2DK118640. This is the full abstract presented at the American Physiology Summit 2024 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.