Fig. 3: IRE1α deficiency in ECs impairs insulin secretion capacity of islets from HFD-fed male mice.

Male Ern1^EC-KO and Ern1^fl/fl mice were fed a NC diet or HFD for 16 weeks. a Representative immunofluorescent staining of insulin/glucagon from pancreas sections. Insulin (Ins, red), Glucagon (Gcg, green), DAPI (blue). Scale bar, 50 μm. b, c Quantification of (b) Ins⁺ β-cell and (c) Gcg⁺ α-cell area per islet (n = 6 per NC group and n = 9 per HFD group). d, e Quantification of the area percentage of (d) Ins⁺ β-cells and (e) Gcg⁺ α-cells per islet (n = 6 per NC group and n = 9 per HFD group). f–k Primary islets were isolated after a 4-hour fast. f Glucose stimulated insulin secretion (GSIS) analysis. Islets pooled from 3 mice of each group were divided into the indicated replicates (30-50 islets per replicate) before incubation at 2.8 mM or 16.8 mM glucose for 1 hour. Insulin levels were then measured and are shown after normalization to the islet protein content. Data are representative of 2 independent experiments. g Islet insulin content was measured following GSIS, shown after normalization to islet protein content (n = 6 per group). h Perifusion analysis of insulin secretion dynamics for pooled islets from HFD-fed mice (n = 3 per group). Islets were stimulated sequentially by 16.8 mM glucose and KCl, and insulin levels are shown as fold changes relative to the unstimulated starting basal value. p.I: Phase I; p.II: Phase II. Shown are representative results from 2 independent experiments. i Quantification of glucose-stimulated biphasic insulin secretion and KCl-induced insulin release in (h) after normalization to the islet protein content (n = 6 mice per group from the 2 independent experiments). j Glucose-stimulated Ca²⁺ influx in whole islets from HFD-fed mice (n = 80 islets from 6 mice per genotype). Shown are representative fluorescent calcium signals in islets maintained at 2.8 mM glucose or following stimulation with 16.8 mM glucose for 150 seconds. Scale bar, 100 μm. k Dynamic monitoring of Ca²⁺ intensity in response to high glucose stimulation in (j). Data are presented as mean ± SEM by two-way ANOVA (b, f) or unpaired two-tailed Student’s t-test (i, k).