Disruption of microRNA-214 during general anaesthesia prevents brain injury and maintains mitochondrial fusion by promoting Mfn2 interaction with Pkm2

Abstract

Duration of surgical general anaesthesia is associated with severe brain injury and neurological deficits. The specific mechanisms underlying post-general anaesthesia brain injury, however, still remain to be elucidated. Herein, we explore the role of microRNA-214 (miR-214) in the occurrence of brain injury after general anaesthesia and its underlying mechanism. Hippocampal tissues and neurons were isolated from rats exposed to 2% sevoflurane. TUNEL stains reflect hippocampal neuron apoptosis. Cultured hippocampal neurons stained with JC-1 and MitoTracker dyes were imaged by fluorescence microscope to visualize changes of mitochondrial membrane potential and mitochondrial fusion. Mitochondrial function was evaluated. Mitofusin 2 (Mfn2) binding to miR-214 or pyruvate kinase M2 (Pkm2) was confirmed by co-immunoprecipitation, immunofluorescence, dual luciferase reporter gene and RNA immunoprecipitation assays. After exposure to 2% sevoflurane, up-regulated miR-214 expression and impaired interaction between Mfn2 and Pkm2 were found in rat hippocampal tissues. Rats exposed to 2% sevoflurane also experienced neuronal injury, mitochondrial defects and deficits in the brain-derived neurotrophic factor (Bdnf) signalling. miR-214 was shown to target Mfn2 by impairing its binding with Pkm2. Inhibiting miR-214 expression using its specific inhibitor improved mitochondrial membrane potential, enhanced mitochondrial fusion, maintained mitochondrial function, restored interaction between Mfn2 and Pkm2, and activated the Bdnf signalling in cultured hippocampal neurons. Adenovirus infection of miR-214 inhibitor reduced neuron apoptosis and maintained mitochondrial function in the hippocampus of rats exposed to 2% sevoflurane. Taken together, the study demonstrates inhibition of miR-214 is cerebral protective against brain injury following general anaesthesia.

Keywords: Mitofusin 2; brain injury; general anaesthesia; microRNA-214; mitochondrial fusion; pyruvate kinase M2.