title: Amikacin Inhibits miR-497 Maturation and Exerts Post-ischemic Neuroprotection.
abstract: MicroRNAs (miRNAs) are a group of small non-coding RNAs that regulate numerous signaling pathways involved in cerebral ischemia reperfusion injury. Recent finding demonstrated that miR-497 promotes ischemic neuronal death by negatively regulating anti-apoptotic proteins and therefore serves as a promising therapeutic target for cerebral ischemic injury. In this study, we present a systematic computational approach that includes 3D modeling, docking-based virtual screening, and molecular dynamics simulation to identify small-molecule inhibitors of pre-miR-497 maturation. The top hit, aminoglycosidic antibiotic, amikacin, formed a stable complex with pre-miR-497. Later, the protective efficacy of amikacin was evaluated against oxygen-glucose deprivation (OGD) and reoxygenation-induced neuronal cell death in SH-SY5Y cells and mouse organotypic hippocampal slice cultures. To confirm the inhibitory potential of amikacin on miR-497 maturation, quantitative real-time PCR was performed to check the expression of bcl-2, one of the primary anti-apoptotic targets of miR-497. Additionally, the expression level of mature miR-497 was quantified using TaqMan® MiRNA Assay Kit. Amikacin treatment effectively reduced OGD-induced cell death compared to control groups both in vitro and organotypic hippocampal slice cultures. Further, amikacin effectively increased the expression of bcl-2 in SH-SY5Y cells subjected to OGD. Interestingly, SH-SY5Y cells treated with amikacin displayed decreased expression of miR-497, probably due to inhibition of pre-miRic form. Our study provides strong evidence that amikacin inhibits miR-497 maturation and promotes ischemic neuronal survival by upregulating anti-apoptotic protein, bcl-2. Future studies directed at evaluating the neuroprotective efficacy and mechanism of amikacin animal models may lead to new therapeutic opportunities for preventing neuronal death after stroke.
authors.affiliation: School of Biotechnology, DBT - Centre for Bioinformatics, National Institute of Technology Calicut, Calicut, 673601, India.
authors.forename: S
authors.initials: S
authors.lastname: Sinoy
authors.affiliation: School of Biotechnology, DBT - Centre for Bioinformatics, National Institute of Technology Calicut, Calicut, 673601, India.|Department of Biotechnology, Manipal Institute of Technology, Manipal University, Manipal, 576104, India.
authors.forename: S M
authors.initials: SM
authors.lastname: Fayaz
authors.affiliation: School of Biotechnology, DBT - Centre for Bioinformatics, National Institute of Technology Calicut, Calicut, 673601, India.
authors.forename: K D
authors.initials: KD
authors.lastname: Charles
authors.affiliation: School of Biotechnology, DBT - Centre for Bioinformatics, National Institute of Technology Calicut, Calicut, 673601, India.
authors.forename: V K
authors.initials: VK
authors.lastname: Suvanish
authors.affiliation: Anatomical Institute, Department of Biomedicine Basel, University of Basel, Pestalozzistrasse 20, 4056, Basel, Switzerland.
authors.forename: Josef P
authors.initials: JP
authors.lastname: Kapfhammer
authors.affiliation: School of Biotechnology, DBT - Centre for Bioinformatics, National Institute of Technology Calicut, Calicut, 673601, India. rajanikant@nitc.ac.in.
authors.forename: G K
authors.initials: GK
authors.lastname: Rajanikant
mesh_terms: D000583:Amikacin / Q000494:pharmacology*; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002545:Brain Ischemia / Q000235:genetics* / Q000473:pathology*; D045744:Cell Line, Tumor; D005947:Glucose / Q000172:deficiency; D006624:Hippocampus / Q000473:pathology; D006801:Humans; D051379:Mice; D035683:MicroRNAs / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D056004:Molecular Dynamics Simulation; D000066829:Neuroprotection / Q000187:drug effects*; D018696:Neuroprotective Agents / Q000494:pharmacology; D010100:Oxygen; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects
chemical_list: C559640:MIRN497 microRNA, human; D035683:MicroRNAs; D018696:Neuroprotective Agents; D019253:Proto-Oncogene Proteins c-bcl-2; D000583:Amikacin; D005947:Glucose; D010100:Oxygen
keywords: Virtual screening; miRNA MD simulation; miRNA inhibition; miRNA modeling