The Role of Carbonic Anhydrases in Glucose-Stimulated Insulin Secretion from Insulinoma MIN6 Cells

Abstract

Background: Carbonic anhydrase (CA) catalyzes the formation of bicarbonate ions and protons from carbon dioxide and water, and also the reverse reaction, thus regulating acid-base balance and processes requiring HCO₃^- in the cell. Previous data from us and others have suggested that CAs participate in the regulation of insulin secretion. The aim of this study was to examine sublines of insulin-secreting MIN6 cells in terms of CA isoform transcript levels, and to analyze their roles in insulin secretion. Methods: Differential expression of CA isoforms in MIN6 cells differing in glucose responsiveness were analyzed by reverse transcription-polymerase chain reaction (RT-PCR). Next, we established stable cell lines with a doxycycline-inducible system overexpressing several of the CA isoforms. We also generated MIN6 sublines with shRNA-mediated suppression and studied glucose-stimulated insulin secretion, as well as glucose metabolism, using radiolabeled glucose in these cells. Results: We confirmed expression of a mitochondrial CA isoform, CA VB, and a cytosolic isoform, CA XIII, was significantly greater (4.18-fold, p < 0.01 for CA VB; 48.3-fold, p < 0.01 for CA XIII), while expression of CA XI, a cytosolic isoform that lacks catalytic activity, was lower (0.50-fold, p < 0.01), in highly glucose-responsive sublines. CA VB overexpression increased (1.32-fold, p < 0.05), while its knockdown suppressed (0.77-fold, p < 0.05), glucose-stimulated insulin secretion. In contrast, CA XIII overexpression and CA XIII knockdown failed to modulate insulin secretion evoked by glucose. CA XI overexpression produced no changes in insulin secretion. Changes in insulin secretion due to CA VB overexpression or knockdown were not accompanied by changes in [5-³H]glucose and [U-¹⁴C]glucose metabolism. Conclusions: CA VB serves as a positive regulator of glucose-stimulated insulin secretion in MIN6 cells.