MiR-212 Attenuates MPP+-Induced Neuronal Damage by Targeting KLF4 in SH-SY5Y Cells
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Purpose: Parkinson’s disease (PD) is a common age-dependent neurodegenerative disease. MiR-212 has been demonstrated toexert protective effects in several neurological disorders. The present study aimed to investigate the role and underlying molecularmechanism of miR-212 in PD. Materials and Methods: 1-methyl-4-phenylpyridinium (MPP+)-induced SH-SY5Y cells were applied as a PD model in vitro. RTqPCRwas used to measure the expression of miR-212 and Kruppel-like factor 4 (KLF4) mRNA. Western blot analysis was performedto detect the protein levels of KLF4, Notch1 and Jagged1. Cell viability and apoptosis were determined by the Cell CountingKit-8 and flow cytometry, respectively. Quantitative analysis of caspase-3 activity, lactate dehydrogenase (LDH), reactiveoxygen species (ROS), superoxide dismutase (SOD), tumor necrosis factor-α (TNF-α), and interleukin-1 beta (IL-1β) was conductedwith corresponding ELISA kits. Dual-luciferase reporter assay was employed to evaluate the relationship between miR-212 and KLF4. Results: MiR-212 was downregulated in MPP+-induced SH-SY5Y cells. Also, miR-212 alleviated MPP+-induced SH-SY5Y cell damage,embodied by increased cell viability, decreased caspase-3 activity, LDH release, ROS production, TNF-α, and IL-1β expression,as well as elevated SOD levels. KLF4 was a direct target of miR-212, and miR-212 repressed KLF4 expression in a post-transcriptionalmanner. Moreover, miR-212-mediated protection effects were abated following KLF4 expression restoration in MPP+-induced SH-SY5Y cells, represented as lowered cell viability and enhanced apoptotic rate. Furthermore, Notch signaling wasinvolved in the regulation of miR-212/KLF4 axis in MPP+-induced SH-SY5Y cells. Conclusion: miR-212 might attenuate MPP+-induced neuronal damage by regulating KLF4/Notch signaling pathway in SH-SY5Ycells, a promising target for PD therapy.
Abstract
Purpose: Parkinson’s disease (PD) is a common age-dependent neurodegenerative disease. MiR-212 has been demonstrated toexert protective effects in several neurological disorders. The present study aimed to investigate the role and underlying molecularmechanism of miR-212 in PD. Materials and Methods: 1-methyl-4-phenylpyridinium (MPP+)-induced SH-SY5Y cells were applied as a PD model in vitro. RTqPCRwas used to measure the expression of miR-212 and Kruppel-like factor 4 (KLF4) mRNA. Western blot analysis was performedto detect the protein levels of KLF4, Notch1 and Jagged1. Cell viability and apoptosis were determined by the Cell CountingKit-8 and flow cytometry, respectively. Quantitative analysis of caspase-3 activity, lactate dehydrogenase (LDH), reactiveoxygen species (ROS), superoxide dismutase (SOD), tumor necrosis factor-α (TNF-α), and interleukin-1 beta (IL-1β) was conductedwith corresponding ELISA kits. Dual-luciferase reporter assay was employed to evaluate the relationship between miR-212 and KLF4. Results: MiR-212 was downregulated in MPP+-induced SH-SY5Y cells. Also, miR-212 alleviated MPP+-induced SH-SY5Y cell damage,embodied by increased cell viability, decreased caspase-3 activity, LDH release, ROS production, TNF-α, and IL-1β expression,as well as elevated SOD levels. KLF4 was a direct target of miR-212, and miR-212 repressed KLF4 expression in a post-transcriptionalmanner. Moreover, miR-212-mediated protection effects were abated following KLF4 expression restoration in MPP+-induced SH-SY5Y cells, represented as lowered cell viability and enhanced apoptotic rate. Furthermore, Notch signaling wasinvolved in the regulation of miR-212/KLF4 axis in MPP+-induced SH-SY5Y cells. Conclusion: miR-212 might attenuate MPP+-induced neuronal damage by regulating KLF4/Notch signaling pathway in SH-SY5Ycells, a promising target for PD therapy.
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