Developmental profiling of cell type specific Igf2 mRNA expression in mouse pancreas.
(A) Assessment of Cre recombinase specificity using the Rosa26YFP-stopfl/fl reporter mouse line at postnatal day 2. Cells with a functional Cre recombinase express YFP protein (green). Nkx3.2-Cre is active in mesenchyme-derived cells (pericytes, smooth muscle cells, stromal cells surrounding pancreatic ducts and acini, fibroblasts and myofibroblasts), Ptf1a-Cre in pancreatic epithelium (acinar cells, pancreatic islets and ducts) and RIP-Cre in pancreatic beta-cells. Sections were co-stained with insulin (INS) marker of pancreatic beta-cells; cholecystokinin (CK), marker of epithelial cells lining pancreatic ducts; amylase (AMY) marker of acinar cells and CD31, marker of endothelial cells. Scale bars are as following: 500 μm in panels i corresponding to each Cre-line; 50 μm in panels ii corresponding to each Cre-line; 100 μm (panel iii) and 200 μm (panels iv and v) for Nkx3.2-Cre. (B) Timeline of Igf2 mRNA expression measured by qRT-PCR in YFP+ or YFP–FACS isolated cells from offspring of Nkx3.2-Cre or RIP-Cre females mated with Rosa26YFP-stopfl/fl males (E–embryonic day; P–postnatal day). Highest levels of Igf2 expression are observed in the mesenchyme fraction (Nkx3.2-Cre YFP+ cells) throughout development. Expression data is normalized to Ppia and is shown as average + SD (n = 3–5 per time point and cell fraction). (C) Igf2 mRNA analysis by in situ hybridization at E19, P2 and P5 showing high levels of expression in mesenchymal cells (purple colour). AS–antisense probe; S–sense probe (negative control); Ac–acinar cells; Isl–pancreatic islet; Du–pancreatic duct; arrows point to cells expressing high levels of Igf2 mRNA. Scale bars are as follows: E19 (300 μm); P2 (100 μm); P5 (30 μm for higher resolution panels ii and iii and 300 μm for lower resolution panels i and iv).