Fig. 3.
Impaired β-cell proliferation in miR-375/ob double-knockout mice. (A) Relative miR-375 expression in lepob/lepob (ob/ob) mice and wild-type (WT) controls measured by real-time PCR and normalized to U6 (N1) or miR-107 (N2) expression levels. (B) Representative 8-μm sections of pancreas from 10-week-old 375/ob (miR-375−/− leptin−/−) and ob/ob mice visualized by immunofluorescence after staining with insulin (green) and glucagon (red). (Bar, 50 μm.) (C) β-Cell mass in 10-week-old WT (gray bar), 375KO (black bar), ob/ob (dark gray bar), and 375/ob (open bar) mice is quantified and reported as mean ± SE. (D–F) Quantification of β-cell number (insulin-positive cells), total endocrine cell number (insulin, glucagon, and somatostatin-positive cells) and α-cell number (glucagon-positive cells) per total pancreatic area in wild-type (gray bars), 375KO (black bars), ob/ob (dark gray bars), and 375/ob (open bars) 10-week-old mice. (G) Ratio of α-cell number to islet cell number in wild-type (gray bar), 375KO (black bar), ob/ob (dark gray bar), and 375/ob (open bar) 10-week-old mice. (H) Quantification of percentage of Ki-67 insulin-positive nuclei within insulin-positive cells of 10-week-old 375/ob (black bar) and ob/ob (gray bar) mice. n > 30 for each genotype. (I) Random-fed blood glucose levels in 375/ob (open squares), ob/ob (filled circles), 375KO (filled squares), and wild-type littermate control (WT) (open circles) mice. (J) Plasma insulin levels in random-fed, 10-week-old 375/ob (black bar) and ob/ob (gray bar) mice. (K) Hepatic glucose production measured after sodium [2-14C]pyruvate was administered by i.p. injection into random-fed, 10-week-old 375/ob (black bar) and ob/ob (gray bar) mice. Data are presented as means ± SE. n = 4–6 animals per genotype unless otherwise noted. *, P < 0.05; **, P < 0.01; ***, P < 0.001.