Abstract
Hypoxic-ischemic encephalopathy (HIE) is a common neonatal disease that can lead to high neonatal mortality rates. Previous studies have indicated that microRNAs (miRs) may be involved in the pathogenesis of HIE; however, the specific mechanisms underlying their involvement require further investigation. The aim of the present study was to investigate the roles of miR-204 and its target gene killin p53 regulated DNA replication inhibitor (KLLN) in HIE using rat HIE models. Brain injury was induced by surgery and incubation of hypoxic incubator brain using 10-day-old pup rats. On day 3, rats were sacrificed, and the infarct size of the brain was determined using a tetrazolium chloride assay. Terminal deoxynucleotidyl transferase UTP nick-end labeling staining was performed to detect the cell death rate in the brain tissue. Following this, the brain tissues were collected, and reverse transcription-quantitative polymerase chain reaction, western blot analysis and immunohistochemistry assays were performed to examine the expression levels of miR-204 and KLLN. Furthermore, neurons were cultured and transfected with miR-204 inhibitors or mimics, and the effect of miR-204 on the proliferation and apoptosis of neurons was examined using MTT and flow cytometric assays. Finally, a dual-luciferase reporter assay was performed to confirm whether KLLN is a direct target of miR-204. The expression of miR-204 was significantly downregulated and the expression of KLLN was significantly increased in the brain tissue of HIE rats (P<0.001). In addition, the transfection with miR-204 inhibitors significantly decreased the proliferation rates and significantly increased the apoptosis rate of neurons; however, transfection with miR-204 mimics prompted the opposite results. The dual-luciferase reporter assay also confirmed that KLLN is a direct target of miR-204. Taken together, the results of the present study demonstrated that miR-204 was downregulated in HIE and that miR-204 may serve important roles in the pathogenesis of HIE through targeting KLLN.