ABSTRACT
Wang L, Fu G, Han R, Fan P, Yang J, Gong K, Zhao Z, Zhang C, Sun K, Shao GMALAT1 and NEAT1 Are Neuroprotective during Hypoxic Preconditioning in the Mouse Hippocampus Possibly by Regulation of NR2B High Alt Med Biol. 00:000-000, 2024. Background: The regulation of noncoding ribonucleic acid (ncRNA) has been shown to be involved in cellular and molecular responses to hypoxic preconditioning (HPC), a situation created by the induction of sublethal hypoxia in the brain. The ncRNAs metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) and nuclear paraspeckle assembly transcript 1 (NEAT1) are abundantly expressed in the brain, where they regulate the expression of various genes in nerve cells. However, the exact roles of MALAT1 and NEAT1 in HPC are not fully understood. Methods: A mouse model of acute repeated hypoxia was used as a model of HPC, and MALAT1 and NEAT1 levels in the hippocampus were measured using real-time polymerase chain reaction (PCR). The mRNA and protein levels of N-methyl-d-aspartate receptor subunit 2 B (NR2B) in the mouse hippocampus were measured using real-time PCR and western blotting, respectively. HT22 cells knocked-down for MALAT1 and NEAT1 were used for in vitro testing. Expression of NR2B, which is involved in nerve cell injury under ischemic and hypoxic conditions, was also evaluated. The levels of spectrin and cleaved caspase-3 in MALAT1 and NEAT1 knockdown HT22 cells under oxygen glucose deprivation/reperfusion (OGD/R) were determined by western blotting. Results: HPC increased the expression of MALAT1 and NEAT1 and decreased the expression of NR2B mRNA in the mouse hippocampus (p < 0.05). Knockdown of MALAT1 and NEAT1 increased both NR2B mRNA and protein levels nearly twofold and caused damage under OGD/R conditions in HT22 cells (p < 0.05). Conclusion: MALAT1 and NEAT1 exert neuroprotective effects by influencing the expression of NR2B.
ABSTRACT
BACKGROUND: Our previous research has demonstrated that hypoxic preconditioning (HPC) can improve spatial learning and memory abilities in adult mice. Adult hippocampal neurogenesis has been associated with learning and memory. The Neurogenic locus notch homolog protein (Notch) was involved in adult hippocampal neurogenesis, as well as in learning and memory. It is currently unclear whether the Notch pathway regulates hippocampal neuroregeneration by modifying the DNA methylation status of the Notch gene following HPC. METHOD: The HPC animal model and cell model were established through repeated hypoxia exposure using mice and the mouse hippocampal neuronal cell line HT22. Step-down test was conducted on HPC mice. Real-time PCR and Western blot analysis were used to assess the mRNA and protein expression levels of Notch1 and hairy and enhancer of split1 (HES1). The presence of BrdU-positive cells and Notch1 expression in the hippocampal dental gyrus (DG) were examined with confocal microscopy. The methylation status of the Notch1 was analyzed using methylation-specific PCR (MS-PCR). HT22 cells were employed to elucidate the impact of HPC on Notch1 in vitro. RESULTS: HPC significantly improved the step-down test performance of mice with elevated levels of mRNA and protein expression of Notch1 and HES1 (P < 0.05). The intensities of the Notch1 signal in the control group, the H group and the HPC group were 2.62 ± 0.57 × 107, 2.87 ± 0.84 × 107, and 3.32 ± 0.14 × 107, respectively, and the number of BrdU (+) cells in the hippocampal DG were 1.83 ± 0.54, 3.71 ± 0.64, and 7.29 ± 0.68 respectively. Compared with that in C and H group, the intensity of the Notch1 signal and the number of BrdU (+) cells increased significantly in HPC group (P < 0.05). The methylation levels of the Notch1 promoter 0.82 ± 0.03, 0.65 ± 0.03, and 0.60 ± 0.02 in the C, H, and HPC groups, respectively. The methylation levels of Notch1 decreased significantly (P < 0.05). The effect of HPC on HT22 cells exhibited similarities to that observed in the hippocampus. CONCLUSION: HPC may confer neuroprotection by activating the Notch1 signaling pathway and regulating its methylation level, resulting in the regeneration of hippocampal neurons.
