Abstract
Autophagy has been shown to maintain neural system homeostasis during stroke. However, the molecular mechanisms underlying neuronal autophagy in ischemic stroke remain poorly understood. This study aims to investigate the regulatory mechanisms of the pathway consisting of MEG3 (maternally expressed gene 3), microRNA-378 (miR-378), and GRB2 (growth factor receptor-bound protein 2) in neuronal autophagy and neurological functional impairment in ischemic stroke. A mouse model of the middle cerebral artery occluded-induced ischemic stroke and an in vitro model of oxygen-glucose deprivation-induced neuronal injury were developed. To understand the role of the MEG3/miR-378/GRB2 axis in the neuronal regulation, the expression of proteins associated with autophagy in neurons was measured by Western blotting analysis, and neuron death was evaluated using a lactate dehydrogenase leakage rate test. First, it was found that the GRB2 gene, up-regulated in middle cerebral artery occluded-operated mice and oxygen-glucose deprivation-exposed neurons, was a target gene of miR-378. Next, miR-378 inhibited neuronal loss and neurological functional impairment in mice, as well as neuronal autophagy and neuronal death by silencing of GRB2. Confirmatory in vitro experiments showed that MEG3 could specifically bind to miR-378 and subsequently up-regulate the expression of GRB2, which in turn suppressed the activation of Akt/mTOR pathway. Taken together, these findings suggested that miR-378 might protect against neuronal autophagy and neurological functional impairment and proposed that a MEG3/miR-378/GRB2 regulatory axis contributed to better understanding of the pathophysiology of ischemic stroke.
Highlights
Stroke, a principal cause of disability and the second leading cause of death in the world, affects an estimated 17 million people each year, of which most are ischemic stroke [1, 2]
These findings suggested that miR-378 might protect against neuronal autophagy and neurological functional impairment and proposed that a maternally expressed gene 3 (MEG3)/miR-378/GRB2 regulatory axis contributed to better understanding of the pathophysiology of ischemic stroke
We investigated the interplay between MEG3, miR-378, and GRB2 in conjunction with the Akt/mTOR pathway in neuronal autophagy and neurological functional impairment caused by ischemic stroke
Summary
A principal cause of disability and the second leading cause of death in the world, affects an estimated 17 million people each year, of which most are ischemic stroke [1, 2]. Aggravation or alleviation of OGD-induced neuronal ischemic injury may be associated with the Akt/mTOR pathway-mediated autophagy [15] Another lncRNA, maternally expressed gene 3 (MEG3), is down-regulated in both MCAO mice and OGD-cultured HT22 cells and has been found to attenuate hypoxia-induced apoptosis of HT22 cells [16]. We investigated the interplay between MEG3, miR-378, and GRB2 in conjunction with the Akt/mTOR pathway in neuronal autophagy and neurological functional impairment caused by ischemic stroke. By analyzing the microarray profile GSE9391, the top 50 differentially expressed genes with the largest fold change are shown in a heat map (Fig. 1A) Among those genes, we found that GRB2 was up-regulated in patients with stroke (Fig. 1A).
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