A DS screening approach identifies a region of chromosome 21 associated with hyperglycemia.

<p>(A) Fasting blood glucose in Ts65Dn (n = 14), Dp16 (n = 11), Ts1Rhr (n = 12) and Tc1 (n = 23) mice (filled symbols) and their respective controls (open symbols). (B) Glucose tolerance test in Ts65Dn (n = 5) and control (n = 6) mice and (C) area under the curve analysis. (D) Glucose tolerance test in Tc1 (n = 9) and control (n = 8) mice and (E) area under the curve analysis. (F) Illustration of the trisomic regions in these DS models used and the 5 genes associated with hyperglycemia in DS that are up-regulated in human T2D islets. *<i>p</i> < 0.05, **<i>p</i> < 0.01.</p

GSIS is reduced <i>in vivo</i> in RCAN1^ox β-cells.

<p>(A) Plasma insulin is similar in WT and RCAN1^ox mice (n = 3). (B) <i>in vivo</i> GSIS is lower in RCAN1^ox mice as defined by (C) reduced area under the curve (AUC) for insulin secretion over 1 hour (n = 3). (D) Insulin sensitivity in WT (n = 4) and RCAN1^ox (n = 5) mice is not different as defined by (E) AUC analysis. (F) Plasma glucagon levels are similar in WT (n = 5) and RCAN1^ox (n = 4) mice. Data represents the mean ± SEM, *p<0.05.</p

RCAN1 expression and methylation in human and mouse Type 2 diabetic islets.

<p>(A) RCAN1 gene expression in human non-diabetic (ND, n = 77, blue symbols) and type 2 diabetic islets (T2D, n = 12, red symbols). (B) RCAN1 expression vs donor HbA1c (n = 89). RCAN1 expression and methylation status at sites (C) cg05056497, (D) cg05156137 and (E) cg21301258 are strongly correlated. (F) RCAN1.1 and RCAN1.4 protein expression in human islets. (G) Quantification of RCAN1 islet protein expression in human islets (n = 3), mouse islets (n = 5) and mouse MIN6 β-cells (n = 3). (H) RCAN1.1 and RCAN1.4 protein expression in control (db/+) and db/db islets. (I) Quantification of (G) (n = 6 control, n = 5 db/db). **<i>p</i> < 0.01, ***<i>p</i> < 0.001.</p

β-cell RCAN1 expression is increased under hyperglycemic conditions via Ca²⁺ and oxidative stress.

<p><b>(A)</b> RCAN1.1 expression (mRNA) is significantly higher in human pancreatic islets cultured at 25 mM glucose (black bars) when compared to islets cultured at 5.5 mM glucose (white bars) (n = 5 experiments). <b>(B)</b> RCAN1.1 and RCAN1.4 expression (protein) is significantly higher in mouse pancreatic islets cultured at 16.7 mM glucose (black bars) when compared to islets cultured at 5.5 mM glucose (white bars). (n = 4 experiments). <b>(C)</b> RCAN1.1 and RCAN1.4 expression (protein) is significantly higher in MIN6 cells cultured at 25 mM glucose for six days (black bars) compared to cells cultured at 5 mM glucose (white bars) for the same time period, (n = 5 experiments). <b>(D)</b> Representative Western blot showing reduced RCAN1.1 and RCAN1.4 expression in MIN6 cells cultured at 25 mM glucose +Nifedipine for 6 days compared to cells cultured at 25 mM glucose for the same time period. <b>(E)</b> Quantification of Western blot images, (n = 5 experiments). <b>(F)</b> Representative Western blot showing reduced RCAN1.1 and RCAN1.4 expression in MIN6 cells cultured at 25 mM glucose +NAC for 6 days compared to cells cultured at 25 mM glucose for the same time period. <b>(G)</b> Quantification of Western blot images, (n = 5 experiments). Data represents the mean ± SEM, *p<0.05, **p<0.01.</p

Glucose-dependent membrane depolarisation is reduced in RCAN1^ox β-cells.

<p>Voltage clamp recordings in (A) WT and (B) RCAN1^ox β-cells. Scale bar in A is 20ms and 20pA in A and B. (C) Current-voltage relationship in WT (n = 59) and RCAN1^ox (n = 39) β-cells. Current clamp recordings from (D) WT and (E) RCAN1^ox β-cells in response to 20mM glucose (arrow). (F) Membrane potential change in response to 20mM glucose in WT (n = 6) and RCAN1^ox (n = 7) β-cells (**<i>p</i> < 0.01). Current clamp recordings from (G) WT and (H) RCAN1^ox β-cells in response to tolbutamide (100μM, arrow). (I) Membrane potential change in response to tolbutamide in WT (n = 6) and RCAN1^ox (n = 6) β-cells.</p

Mitochondrial respiratory output is reduced in RCAN1^ox islets.

<p>(A) Oxygen consumption rate (OCR) in WT (n = 5) and RCAN1^ox (n = 6) islets under various conditions. (B) OCR in 3 mM or 20 mM glucose. (C) Islet ATP levels (n = 7 WT and n = 4 RCAN1^ox). (D) Methyl-succinate-induced (10 mM) insulin secretion (n = 4/group). (E) Effect of carboxyatractyloside (CATS, 200 μm) on insulin secretion in response to 20 mM glucose (n = 4–7). Immunocytochemical labelling of (F) RCAN1, (G) mitochondria (Mitotracker Red) and (H) both in MIN6 cells. *<i>p</i> < 0.05, **<i>p</i> < 0.01.</p

Parent-of-origin effect of <i>GRB10</i> rs933360 on insulin secretion and glucose levels.

<p>(A) No significant effect for CIR was observed from the paternally transmitted A-allele. (B) Carriers of the maternally transmitted A-allele showed lower CIR compared to the G-allele. (C) Carriers of the paternally transmitted A-allele had elevated fasting plasma glucose levels, whereas (D) the maternally transmitted A-allele was associated with lower fasting plasma glucose levels. Fin-Swe = Trios from Finland and Sweden, Amish = Amish Family Diabetes Study, Kuopio = Kuopio Offspring Study.</p