Objective To explore the effect of microRNA-199a (miR-199a) on neuronal apoptosis in rats with cerebral infarction and its role in regulating silent information regulator 1 (SIRT1).
Methods The experimental model of cerebral infarction rats was constructed, and 60 rats were randomly divided into sham operation group, cerebral infarction group, miR-199a inhibition group, miR-199a negative control group, and miR-199a inhibition+SIRT1 inhibition group, with 12 rats in each group. Neurobehavioral tests were used to evaluate and judge the changes of nerve function in rat, the hematoxylin-eosin (HE) staining method was used to observe the pathological changes of neurons in the brain tissue of rats in each group, flow cytometry was used to detect the apoptosis of cerebral cortex neurons, fluorescence quantitative polymerase chain reaction (PCR) was used to detect the expression levels of miR-199a and SIRT1 mRNA in the brain tissues of each group, and Western blot method was used to detect the expression levels of SIRT1 and Cleaved caspase-3 proteins in the brain tissues of each group. Dual luciferase reporter system was used to verify the targeted regulation relationship between miR-199a and SIRT1.
Results Compared with the sham operation group, rats in the cerebral infarction group had degeneration and neuronal necrosis in the brain tissues, which were accompanied by the neuronal apoptosis rate, protein expression of Cleaved caspase-3, neurological score and miR-199a expression significantly increased, and the SIRT1 mRNA and protein expression of SIRT1 significantly reduced (P < 0.05); compared with the miR-199a negative control group, the degree of neuronal necrosis in the miR-199a inhibition group was weakened and the structure of brain tissue gradually recovered, which were accompanied by the neuronal apoptosis rate, protein expression of Cleaved caspase-3, neurological score and miR-199a expression significantly reduced, and the SIRT1 mRNA and protein expression of SIRT1 significantly increased (P < 0.05); compared with miR-199a inhibition group, the degrees of neuron necrosis and brain tissue degeneration in the miR-199a inhibition + SIRT1 inhibition group became severer, which were accompanied by the neuronal apoptosis rate, protein expression of Cleaved caspase-3 and neurological score significantly increased, and the SIRT1 mRNA and protein expression of SIRT1 significantly reduced(P < 0.05); there were no significant differences in the indicators mentioned above between the miR-199a negative control group and the cerebral infarction group (P>0.05); the results of double luciferase reporter gene experiment showed that miR-199a could specifically bind to wild-type SIRT1 3′-UTR, and there was a targeted regulation relationship between two indexes.
Conclusion Inhibiting the expression of miR-199a can targeted increase the expression level of SIRT1, relieve neuronal apoptosis in rats with cerebral infarction, and improve neurological function.
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