Aberrant expression of microRNAs (miRs) contributes to diabetic renal complications including renal hypertrophy and matrix protein accumulation. Reduced expression of PTEN by hyperglycemia contributes to these processes. We considered involvement of microRNA in the downregulation of PTEN. In the renal cortex of type 1 diabetic mice, we detected increased expression of miR-214 in association with decreased levels of PTEN and, enhanced Akt phosphorylation and fibronectin expression. Mesangial and proximal tubular epithelial cells exposed to high glucose showed augmented expression of miR-214. Mutagenesis studies using 3'UTR of PTEN in a reporter construct revealed PTEN as a direct target of miR-214, which controls its expression in both these cells. Overexpression of miR-214 decreased the levels of PTEN and increased Akt activity similar to high glucose and lead to phosphorylation of its substrates GSK3β, PRAS40 and tuberin. In contrast, quenching of miR-214 inhibited high glucose-induced Akt activation and its substrate phosphorylation; these changes were reversed by siRNAs against PTEN. Importantly, respective expression of miR-214 or anti-miR-214 increased or decreased the mTORC1 activity induced by high glucose. Furthermore, mTORC1 activity was controlled by miR-214-targeted PTEN via Akt activation. In addition, neutralization of high glucose-stimulated miR-214 expression significantly inhibited cell hypertrophy and expression of the matrix protein fibronectin. Finally, the anti-miR-214-induced inhibition of these processes was reversed by the expression of constitutively active Akt kinase and hyperactive mTORC1. These results uncover a significant role of miR-214 in the activation of mTORC1 that contributes to high glucose-induced mesangial and proximal tubular cell hypertrophy and fibronectin expression.