CCN3 impairs glucose-stimulated insulin secretion.

<p>A) We evaluated the effects of <i>Ccn3</i> overexpression on insulin secretion by the hGH co-transfection system. In brief, we measured the hGH levels in the media of INS832/13 cells co-transduced with hGH and CCN3 or the empty plasmid and incubated at 2.8 mM (G2.8) or 16 mM glucose (G16). KCl (35 mM) was used as control. B) We studied the effects of CCN3 protein on insulin secretion. Insulin released in the culture medium was measured following incubation of INS832/13 cells in 2.8 mM or 16 mM glucose/KRBH medium or in the presence of 35 mM KCl. Secreted insulin levels were normalized to protein content. C) Measurements of intracellular calcium in living INS832/13 cells treated as described in (B) using the Fura-2 dye. We measured the ratio of fluorescence signals produced by excitation of 340 nm and 380 nm and detected at 510 nm to determine intracellular calcium concentrations. D) Glucose oxidation was measured as ¹⁴CO₂ production from [U-¹⁴C]-glucose in cells treated as described in (A) and the results were normalized to protein content. E) Cellular insulin content was measured by Elisa after lysis of the cells in acidic ethanol. Results represent mean ± SEM of three separate experiments carried out in triplicate. *, p<0.05.