Introduction and Objective: GLP-1 is secreted from enteroendocrine L cells and exerts multifaceted effects as well as its insulinotropic action. Recently, GLP-1 receptor agonists have been widely applied and have attracted attention as a treatment that not only improves hyperglycemia, but also prevents cardiovascular events. However, it remains unclear when, where, and how L cells are generated from their progenitors and differentiate.Methods: We previously established “GLP1-Timer” mice, in which a novel reporter construct eGFP-IRES-mRFP was inserted into the proglucagon gene. In the GLP1-Timer mouse, green fluorescence is detected first because the expression level of mRFP, controlled by the IRES, is lower than that of eGFP. This allows us to distinguish newly-generated L (early L) cells from more differentiated L (late L) cells using microscopy and flow cytometry. The stomach, duodenum, jejunum, ileum, and colon of adult GLP1-Timer mice were dissected to prepare frozen sections. Spatial information on early and late L cells was then analyzed by confocal microscopy, and the number of both types of L cells was quantified using flow cytometry.Results: Both green-fluorescent early L cells and green/red double-fluorescent late L cells were observed from the stomach to the colon by microscopy. Notably, about half of the early L cells were located higher (closer to the apical region) than the late L cells, suggesting that L-cell genesis occurs independently of their distance from the crypt. Flow cytometric quantification of early and late L cells demonstrated that the number of early L cells was highest in the ileum and colon (0.20% of intestinal epithelial cells) and significantly lower in the duodenum.Conclusion: The GLP1-Timer reporter system provided novel spatiotemporal insights into L-cell genesis and differentiation. L-cell genesis is likely to occur in a stochastic manner, independent of their distance from the crypt.
R. Fujishima: None. T. Taguchi: None. A. Suzuki: None. N. Shimizu: None. K. Kimura: None. S. Ito: None. T. Masaki: None. T. Miyatsuka: Speaker’s Bureau; Eli Lilly and Company, Novo Nordisk, Sumitomo Dainippon Pharma Co., Ltd, Kowa Company, Ltd, Mitsubishi Tanabe Pharma Corporation, Abbott Japan Co., Ltd, Novartis Pharmaceuticals Corporation, Ono Pharmaceutical Co., Ltd.
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