Contribution of a Non-β-Cell Source to β-Cell Mass during Pregnancy
Figure 3
Ngn3-positive cells in pancreatic islets during pregnancy.
A) Percentage of cells in the pancreatic islets of Ngn3:EGFP mice stained positive for EGFP (therefore expressing Ngn3, these cells will be denoted as Ngn3-EGFP+ in all images) but negative for endocrine hormones drops during pregnancy. The number of Ngn3-EGFP+ cells was expressed as a percentage of all the cells in the islets. Islets (outlined in white) were identified by staining for endocrine hormones. At least 10,000 cells were counted from each mouse. N = 3–4 separate mice were sampled at each gestational stage. “*”: p<0.05 in comparison to the non-pregnant (G0) mice by student’s t-test. B) A representative image of an islet at G0, G8 and G14. C) A representative image of TUNEL staining of an islet (outlined) at G6. Islets at postnatal day 2 (P2) and DNase treated pancreas served as controls. Black arrowheads indicate apoptotic (brown) cells. D) mRNA levels of Ngn3, as measured by qRT PCR in islets isolated from mice on days 0, 6, 9, and 15 of pregnancy expressed as mean ± SEM; N = 6 separate mice at each gestational stage. All expression levels are normalized to that of PGK-1 (phosphoglycerate kinase 1). “*”: p<0.05 and “**”: p<0.01 in comparison to non-pregnant (G0) mice by one-way ANOVA. E) Whole cell lysates were prepared from islets isolated from mice on days 0, 9, and 15 of pregnancy and immunoblotted for Ngn3. The optical density of the Ngn3 band was normalized to that of actin, which was used as the loading control. A representative blot is shown. Results represent mean ± SEM. N = 3–5 mice at each gestational stage from three separate experiments. “*”: p<0.05 in comparison to non-pregnant (G0) mice by one-way ANOVA.