Cdk5rap3 is essential for intestinal Paneth cell development and maintenance

摘要

a Scheme of mouse breeding to generate IEC-specific KO mice. b Quantitative RT-PCR analysis of Cdk5rap3 mRNA using the primers specific for floxed exons (n = 3 mice per genotype); Total RNA was isolated from intestinal crypts. c Immunoblotting of Cdk5rap3 in the lysate of intestinal epithelial cell lysates. d Immunohistochemistry of Cdk5rap3 in wild-type and Cdk5rap3 KO ileal sections. e Significant loss of Paneth cells in Cdkrap3∆/∆IEC intestine. The number of Paneth and goblet cells were counted in double-blinded fashion from more than 20 crypts, and 4 mice of each phenotypes were scored. ****p < 0.0001 (n = 4 mice per genotype). f Lysozyme staining of ileal sections of wild-type and Cdkrap3∆/∆IEC mice. Lysozyme-positive cells per crypt were scored. ***p < 0.001 (n = 4 mice per genotype). g Electron micrographs of the crypts of wild-type and Cdk5rap3∆/∆IEC mice. Mitotic cells with condensed chromatin were marked by solid arrows. h Quantitative RT-PCR analysis of cell type-specific gene expression. Total RNA was isolated from scraped cells of ileal section. *p < 0.05, **p < 0.01, ***p < 0.001 (n = 4 mice per genotype). i The number of enteroendocrine cells (Chromogranin A-positive) in wild-type and Cdkrap3∆/∆IEC intestine. The number of ChA+ cells per crypt-villus axis was counted and four mice of each phenotypes were scored. j Mitotic cells (phospho-Histone H3 Ser10 staining) in wild-type and Cdk5rap3∆/∆IEC ileal sections. The number of p-H3+ cells per crypt-villus axis was scored. **p < 0.01. (n = 4 mice per genotype).