RESUMO
Studies have shown that downregulation of nuclear-enriched autosomal transcript 1 (Neat1) may adversely affect the recovery of nerve function and the increased loss of hippocampal neurons in mice. Whether Neat1 has protective or inhibitory effects on neuronal cell apoptosis after secondary brain injury remains unclear. Therefore, the effects of Neat1 on neuronal apoptosis were observed. C57BL/6 primary neurons were obtained from the cortices of newborn mice and cultured in vitro, and an oxygen and glucose deprivation cell model was established to simulate the secondary brain injury that occurs after traumatic brain injury in vitro. The level of Neat1 expression in neuronal cells was regulated by constructing a recombinant adenovirus to infect neurons, and the effects of Neat1 expression on neuronal apoptosis after oxygen and glucose deprivation were observed. The experiment was divided into four groups: the control group, without any treatment, received normal culture; the oxygen and glucose deprivation group were subjected to the oxygen and glucose deprivation model protocol; the Neat1 overexpression and Neat1 downregulation groups were treated with Neat1 expression intervention techniques and were subjected to the in oxygen and glucose deprivation protocol. The protein expression levels of neurons p53-induced death domain protein 1 (PIDD1, a pro-apoptotic protein), caspase-2 (an apoptotic priming protein), cytochrome C (a pro-apoptotic protein), and cleaved caspase-3 (an apoptotic executive protein) were measured in each group using the western blot assay. To observe changes in the intracellular distribution of cytochrome C, the expression levels of cytochrome C in the cytoplasm and mitochondria of neurons from each group were detected by western blot assay. Differences in the cell viability and apoptosis rate between groups were detected by cell-counting kit 8 assay and terminal deoxynucleotidyl transferase dUTP nick-end labeling assay, respectively. The results showed that the apoptosis rate, PIDD1, caspase-2, and cleaved caspase-3 expression levels significantly decreased, and cell viability significantly improved in the Neat1 overexpression group compared with the oxygen and glucose deprivation group; however, Neat1 downregulation reversed these changes. Compared with the Neat1 downregulation group, the cytosolic cytochrome C level in the Neat1 overexpression group significantly decreased, and the mitochondrial cytochrome C level significantly increased. These data indicate that Neat1 upregulation can reduce the release of cytochrome C from the mitochondria to the cytoplasm by inhibiting the PIDD1-caspase-2 pathway, reducing the activation of caspase-3, and preventing neuronal apoptosis after oxygen and glucose deprivation, which might reduce secondary brain injury after traumatic brain injury. All experiments were approved by the Animal Ethics Committee of the First Affiliated Hospital of Chongqing Medical University, China, on December 19, 2020 (approval No. 2020-895).
RESUMO
OBJECTIVE: To investigate the effect of vildagliptin on pentamethazol (PTZ)-induced epilepsy in rats and explore the molecular mechanism. METHODS: Samples of temporal cortex from 23 patients with temporal lobe epilepsy were collected as epilepsy group and samples of temporal cortex from 14 patients with brain trauma were used as control group. Ninety male SD rats were randomly divided into control group (group A), PTZ-induced epilepsy group (group B), saline 2 mL/kg group (group C), vildagliptin 2.5 mg/kg group (group D), vildagliptin 5mg/kg group (group D) and vildagliptin 10 mg/kg group (group F). Use chronic model of epilepsy induced by PTZ (35 mg/kg) intraperitoneal injection for 3 consecutive weeks, and changes of behavior were observed. The expression of GLP-1R was detected by Western blotting and immunohistochemical (IHC) staining, and the expression of GLP-1 was detected by enzyme-linked immunosorbent assay (ELISA). The location of GLP-1R was detected by immunofluorescent staining. RESULTS: Immunofluorescent staining showed that the GLP-1R located in the neurons, and GLP-1R expression was obviously decreased both in patients with TLE and in rats with epilepsy. The latency time was prolonged and epilepsy attack time was decreased after vildagliptin treatment (P<0.05). GLP-1R expression was increased after vildagliptin treatment (P<0.05). ELISA showed the change of GLP-1 expression was the same as GLP-1R. CONCLUSION: Vildagliptin can suppress temporal lobe epilepsy in rats by up-regulating GLP-1 and GLP-1R expressions.
