An Overfeeding-Induced Obesity Mouse Model Reveals Necessity for Sin3a in Postnatal Peak β-Cell Mass Acquisition

Abstract

The increase of functional β-cell mass is paramount to maintaining glucose homeostasis in the setting of systemic insulin resistance and/or augmented metabolic load. Understanding compensatory mechanisms that allow β-cell mass adaptation may allow for the discovery of therapeutically actionable control nodes. In this study, we report the rapid and robust β-cell hyperplasic effect in a mouse model of overfeeding-induced obesity (OIO) based on direct gastric caloric infusion. By performing RNA sequencing in islets isolated from OIO mice, we identified Sin3a as a novel transcriptional regulator of β-cell mass adaptation. β-Cell-specific Sin3a knockout animals showed profound diabetes due to defective acquisition of postnatal β-cell mass. These findings reveal a novel regulatory pathway in β-cell proliferation and validate OIO as a model for discovery of other mechanistic determinants of β-cell adaptation.

Figure 1

Overfeeding induces rapid β-cell mass acquisition. A: Representative insulin staining (red) in whole pancreata from saline-infused control and OIO mice and quantification of β-cell mass (n = 5–8 mice/group). Scale bars: 500 μm. B: Representative images from pancreatic sections of control and OIO mice stained with insulin (green), Ki67 (red), and DAPI (blue), with quantification of Ki67⁺ β-cells (n = 5–11 mice/group). Scale bars: 20 μm. C: Quantification of individual β-cell size in OIO and control mice. Violin plots of all data points are shown; average and median cell size are provided in the text. Data are group means. **P < 0.01, ***P < 0.001 by two-tailed t test.

Figure 2

OIO induces profound changes in the islet transcriptome. A: Change in body weight from baseline in OIO and saline-infused control mice. Data are mean ± SEM. *P < 0.05, ***P < 0.001 by two-tailed t test. B: Volcano plot showing differentially expressed genes (DEGs) (adjusted [adj.] P < 0.05) in islets from 9-day OIO mice compared with saline-infused controls. C: Biological processes corresponding to upregulated DEGs in OIO islets, after gene ontology enrichment analysis using Enrichr. D–G: GSEA plot for upregulated DEGs in sorted proliferating β-cells (11) (D), isolated islets from HFD-fed mice (4) (E), pregnant mice (5) (F), or S961-treated mice (10) (G) compared with a ranked list of DEGs in OIO islets, with normalized enrichment score (NES) and false discovery rate (FDR) q value as indicated. H: Transcription factor (TF) enrichment of upregulated DEGs in OIO islets, showing that 158 of the top 250 upregulated DEGs are associated with E2f4, Foxm1, or Sin3a. Network shown and expanded in Supplementary Fig. 2. DOWN, downregulated; UP, upregulated.

Figure 3

β-Sin3aKO mice develop diabetes and impaired β-cell mass expansion. A and B: Glucose (A) and insulin (B) levels in 8-week-old male β-Sin3aKO and control mice after 4 h of fasting. C: Glucose tolerance test in 8-week-old male β-Sin3aKO (▪) and control mice (○). D: Body weight of 8-week-old male β-Sin3aKO and control mice. E: Representative images from pancreatic sections of 8-week-old male β-Sin3aKO and control mice stained with insulin (green), glucagon (red), and DAPI (blue). Scale bars: 20 μm. F: Quantification of β-cell mass at different postnatal stages in β-Sin3aKO and control mice (n = 3–5 mice/group). Data are group mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001 by two-tailed t test.

Figure 4

Impaired cell cycle progression and β-cell death in β-Sin3aKO mice. A: Quantification of Ki67⁺ β-cells in pancreatic sections in P2 β-Sin3aKO and control mice (n = 5–6 mice/group). B–D: Representative images and quantification of BrdU⁺ (B), phospho-histone H3⁺ (pHH3) (C), and phospho-histone H2A.X⁺ (γH2AX) (D) β-cells in pancreatic sections in P2 β-Sin3aKO and control mice (n = 4–5 mice/group). E and F: Representative images of TUNEL and Ki67 staining in pancreatic sections from β-Sin3aKO, with orthogonal projection shown in F. G: Quantification of TUNEL⁺ and TUNEL⁺/Ki67⁺ β-cells in pancreatic sections from P2 β-Sin3aKO and control mice (n = 4 mice/group). Scale bars: 20 μm. Data are group means. **P < 0.01, ***P < 0.001 by two-tailed t test.