miR-29a Negatively Affects Glucose-Stimulated Insulin Secretion and MIN6 Cell Proliferation via Cdc42/-Catenin Signaling.

BACKGROUND

Diabetes is a progressive metabolic disease characterized by hyperglycemia. Functional impairment of islet cells can occur to varying degrees. This impairment can initially be compensated for by proliferation and metabolic changes of cells. Cell division control protein 42 (Cdc42) and the microRNA (miRNA) miR-29 have important roles in -cell proliferation and glucose-stimulated insulin secretion (GSIS), which we further explored using the mouse insulinoma cell line MIN6.

METHODS

Upregulation and downregulation of miR-29a and Cdc42 were accomplished using transient transfection. miR-29a and Cdc42 expression was detected by real-time PCR and western blotting. MIN6 proliferation was detected using a cell counting kit assay. GSIS under high-glucose (20.0 mM) or basal-glucose (5.0 mM) stimulation was detected by enzyme-linked immunosorbent assay. The miR-29a binding site in the Cdc42 mRNA 3'-untranslated region (UTR) was determined using bioinformatics and luciferase reporter assays.

RESULTS

miR-29a overexpression inhibited proliferation ( < 0.01) and GSIS under high-glucose stimulation ( < 0.01). Cdc42 overexpression promoted proliferation ( < 0.05) and GSIS under high-glucose stimulation ( < 0.05). miR-29a overexpression decreased Cdc42 expression ( < 0.01), whereas miR-29a downregulation increased Cdc42 expression ( < 0.01). The results showed that the Cdc42 mRNA 3'-UTR is a direct target of miR-29a . Additionally, Cdc42 reversed miR-29a-mediated inhibition of proliferation and GSIS ( < 0.01). Furthermore, miR-29a inhibited -catenin expression ( < 0.01), whereas Cdc42 promoted -catenin expression ( < 0.01).

CONCLUSION

By negatively regulating Cdc42 and the downstream molecule -catenin, miR-29a inhibits MIN6 proliferation and insulin secretion.