Establishment of lineage tracing method of mice small intestinal stem cell and application in radiation injury / 解放军医学杂志

ABSTRACT

Objective To establish a lineage tracing method with Lgr5-EGFP-CreERT2/ROSA26-stop-EYFP mouse for observing the role of Lgr5+ intestinal stem cells in the renovation of small intestine tissue, and investigate the effects of radiation on the reconstruction of small intestinal stem cell tissue. Methods-Lgr5-EGFP-CreERT2/ROSA26-stop-EYFP mice were identified by genotype analysis. Tamoxifen was used to induce the expression of fluorescent protein in mice. The differentiation of intestinal stem cells induced by Tamoxifen was observed by laser confocal microscopy. Lgr5-EGFP-CreERT2/ROSA26-stop-EYFP mice were irradiated with60Co γ-rays, and the effects of irradiation on the reconstruction of small intestinal stem cell tissue were examined. Results-Double positive mice were obtained by identification of Lgr5 gene (174bp) in DNA extraction from the mice tail with PCR method. In mice treated with single injection of Tamoxifen (100mg/kg), it was observed by laser confocal scanning microscopy that the Lgr5+ intestinal stem cells started dividing at the 5th day after inducement, reached the top of the villi at the 7th day, fluorescence appeared in a few of whole intestinal villi at the 14th day, and spread over more intestinal villi with dense fluorescence at the 45th day. However, in mice exposed to 15Gy irradiation, the intestinal villi fell off seriously without fluorescence in crypts. Conclusion-The lineage tracing model of intestinal stem cell has been successfully established and then used to evaluate the irradiation injuries to intestinal stem cells.