Potential Roles of miR-374a-5p in Mediating Neuroprotective Effects and Related Molecular Mechanism
Feng Jiang1, Min Yang1, Chuyan Wu2
1Neonatal Department, Obstetrics and Gynecology Hospital of Fudan University, No.419 Fangxie Road, Huangpu District, Shanghai, 200011, China.
Insights
MicroRNA-374a-5p (miR-374a-5p) is decreased in hypoxic-ischemic encephalopathy (HIE). Overexpressing miR-374a-5p inhibits PTEN/PI3K pathway, reducing apoptosis in HIE.
Area of Science:
- Molecular Biology
- Neuroscience
- Biochemistry
Background:
- Hypoxic-ischemic encephalopathy (HIE) is a serious neonatal condition.
- Reduced miR-374a-5p levels are observed in HIE patients.
- The molecular mechanisms underlying miR-374a-5p's role in HIE are unclear.
Purpose of the Study:
- To investigate the molecular mechanism of miR-374a-5p in HIE development.
- To explore miR-374a-5p as a potential therapeutic target for HIE.
Main Methods:
- Established an in vitro HIE model using oxygen/glucose deprivation (OGD) in PC12 cells.
- Quantified miR-374a-5p, PTEN, and apoptotic markers using RT-qPCR and Western blot.
- Assessed apoptosis via flow cytometry and validated miR-374a-5p targets using dual-luciferase reporter assay.
Main Results:
- miR-374a-5p was significantly downregulated in HIE models and patient cells.
- miR-374a-5p overexpression reduced OGD-induced apoptosis, while inhibition increased it.
- miR-374a-5p directly targeted PTEN, inhibiting its expression and the downstream PI3K pathway.
Conclusions:
- miR-374a-5p plays a protective role against apoptosis in HIE.
- The miR-374a-5p/PTEN/PI3K axis is a key pathway in HIE pathogenesis.
- Restoring miR-374a-5p levels may offer a therapeutic strategy for HIE.
Abstract:
Previous studies found that miR-374a-5p was decreased in infants suffering from the hypoxic-ischemic encephalopathy (HIE) compared with healthy control infants. However, the molecular mechanism of miR-374a-5p in the development of HIE remained unknown. This study is aimed to investigate the potential molecular pathway for shedding light on the treatment of HIE. An in vitro ischemia model in PC12 cells was established by oxygen/glucose deprivation (OGD). Reverse-transcription quantitative polymerase chain reaction and western blot were used to determine the levels of related genes or proteins in the OGD model or cells obtained from infants with HIE. Flow cytometry was conducted to quantify the apoptosis level of PC12 cells after OGD treatment. The TargetScan prediction algorithm was used to identify the potentially functional targets of miR-374a-5p. A dual-luciferase reporter assay was adopted to elucidate the sequences of miR-374a-5p binding to the 3'-UTR of potential target-PTEN. miR-374a-5p was downregulated in cells derived from human newborns with HIE, rat model with HIE, and PC12 cells after the OGD treatment. Inhibition of miR-374a-5p increased the expression of apoptotic markers and the apoptosis percentage of PC12 cells induced by OGD treatment while overexpression rescued the apoptosis. Meanwhile, PTEN expression was increased and suppressed after miR-374a-5p silence or overexpression, respectively. Upregulation of PTEN reversed the inhibitory effect of apoptotic markers elevation and PC12 cells apoptosis by the overexpression of miR-374a-5p after OGD treatment. PI3K pathway was required for the apoptosis effect caused by PTEN overexpression. We found that overexpression of miR-374a-5p reduced cell apoptosis through inhibiting PTEN/PI3K pathway in PC12 cells treated by OGD.
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