Evidence suggest that overexpression of hypoxia-inducible factor-1α (HIF-1α) is linked to multidrug resistance of epilepsy. Here we explored whether aberrant expression of HIF-1α is regulated by miRNAs. Genome-wide microRNA expression profiling was performed on temporal cortex resected from mesial temporal lobe epilepsy (mTLE) patients and age-matched controls. miRNAs that are putative regulator of HIF-1α were predicted via target scan and confirmed by real-time quantitative polymerase chain reaction (RT-qPCR). Mimics or miRNA morpholino inhibitors were transfected in astrocytes and luciferase reporter assay was applied to detect HIF-11α expression. Microarray profiling identified down-regulated miR-153 as a putative regulator of HIF-1α in temporal cortex resected from surgical mTLE patients. RT-qPCR confirmed down-regulation of miR-153 in plasma of mTLE patients in an independent validation cohort. Knockdown of miR-153 significantly enhanced expression of HIF-1α while forced expression of miR-153 dramatically inhibited HIF-1α expression in pharmacoresistant astrocyte model. Luciferase assay established that miR-153 might inhibit HIF-1α expression via directly targeting two binding sites in the 3'UTR region of HIF-1α transcript. These data suggest that down-regulation of miR-153 may contribute to enhanced expression of HIF-1α in mTLE and serve as a novel biomarker and treatment target for epilepsy.