ABSTRACT
Seipin plays a crucial role in lipid metabolism and is involved in neurological disorders. However, the function and mechanism of action of seipin in acute ischemic stroke have not yet been elucidated. Here, we aimed to investigate the effect of seipin on neuroinflammation induced by oxygen-glucose deprivation/reoxygenation (OGD/R) and further explore the molecular mechanism by functional experiments. Our results revealed a significant decrease in seipin mRNA levels, accompanied by enhanced expression of TNF-α in patients with AIS, and a significant negative correlation between seipin and TNF-α was observed. Additionally, there was a negative correlation between seipin levels and the National Institutes of Health Stroke Scale (NIHSS) score. Furthermore, seipin levels were also decreased in middle cerebral artery occlusion/reperfusion (MCAO/R) mice and OGD/R-treated BV2 cells. RNA sequencing analysis showed that seipin knockdown altered the Toll-like receptor 3 (TLR3) signaling pathway. It was further confirmed in vitro that seipin knockdown caused significantly increased secretion of inflammatory factors including TNF-α, interleukin (IL)-1ß, and interferon (IFN)-ß. Meanwhile, seipin knockdown activated the Tlr3 signal pathway while this effect could be reversed by Tlr3 inhibitor in OGD/R treated BV2 cells. Furthermore, neuroinflammation induced by OGD/R was significantly reduced by seipin overexpression. Overall, our study demonstrate that seipin deficiency aggravates neuroinflammation by activating the TLR3/TRAF3/NF-κB signaling pathway after OGD/R stimuli, and suggest that seipin may be a potential therapeutic target for AIS.
Subject(s)
Glucose , NF-kappa B , Neuroinflammatory Diseases , Oxygen , Signal Transduction , TNF Receptor-Associated Factor 3 , Toll-Like Receptor 3 , Aged , Animals , Female , Humans , Male , Mice , Middle Aged , Cell Line , Disease Models, Animal , Glucose/metabolism , GTP-Binding Protein gamma Subunits/metabolism , GTP-Binding Protein gamma Subunits/genetics , Infarction, Middle Cerebral Artery/metabolism , Infarction, Middle Cerebral Artery/immunology , Ischemic Stroke/metabolism , Ischemic Stroke/immunology , Mice, Inbred C57BL , Neuroinflammatory Diseases/immunology , Neuroinflammatory Diseases/metabolism , NF-kappa B/metabolism , Oxygen/metabolism , Reperfusion Injury/metabolism , Reperfusion Injury/immunology , TNF Receptor-Associated Factor 3/metabolism , TNF Receptor-Associated Factor 3/genetics , Toll-Like Receptor 3/metabolism , Toll-Like Receptor 3/geneticsABSTRACT
BACKGROUND: Ischemic cerebrovascular disease (ICVD) is one of three fatal diseases in humans, along with heart disease and malignant tumors. Cerebral ischemia/reperfusion injury (CI/RI) is the primary cause of ICVD. Recently, seipin was reported to be crucial for lipid droplet formation, hepatic steatosis, and axonal atrophy. However, the function and mechanism of seipin in CI/RI has not been explicitly stated. METHODS: Oxygen-glucose deprivation/reoxygenation (OGD/R) hippocampal neuron cell line (HT-22) and middle cerebral artery occlusion (MCAO) in rats were established. The levels of apoptosis- and autophagy-related proteins and seipin were confirmed by western blot. Cell proliferation was assessed with CCK-8, and ischemic infarction and pathological structure were monitored by TTC and H&E staining, and tissue apoptosis was assessed through TUNEL assay. RESULTS: The proliferative activity was decreased, and apoptosis and autophagy pathways could also be induced in the OGD/R HT-22 cells. Seipin overexpression accelerated viability and inhibited apoptosis and autophagy in the OGD/R HT-22 cells. Moreover, the data revealed that seipin overexpression could also attenuate cerebral infarction, apoptosis. Autophagy pathways could be repressed by seipin in the MCAO rats. CONCLUSION: Seipin has a protective role against CI/RI in rats, and its mechanism might be relevant to the suppression of apoptosis and autophagy, suggesting that seipin might be a latent target for CI/RI.
Subject(s)
Brain Ischemia , Reperfusion Injury , Animals , Humans , Rats , Apoptosis , Autophagy , Brain Ischemia/complications , Cell Line , Glucose/metabolism , Infarction, Middle Cerebral Artery/metabolism , Oxygen/metabolism , Reperfusion Injury/prevention & controlABSTRACT
Ischemia/reperfusion (I/R) injury induces activation of the endoplasmic reticulum stress (ERS) pathway, accompanied by an increase in apoptosis. Multiple microRNAs (miRNAs/miRs) are dysregulated during I/R and contribute to I/R-induced injury. miRNAs act as suppressors of gene expression and negatively regulate gene expression by targeting the protein-coding sequence (CDS) of specific target mRNAs. Seipin is an endoplasmic reticulum protein that has recently been associated with ERS. We previously reported that seipin is the target gene of miR1873p. Therefore, we explored the involvement of miR-187-3p in I/R-induced ERS via the regulation of seipin. A rat MCAO/R model was established by 1â¯h of occlusion and 24â¯h reperfusion. Neurological deficits and infarction area were examined. PC12 cells were exposed to oxygenglucose deprivation/reoxygenation (OGD/R) to model I/R. Expression levels of miR-187-3p and proteins related to ERS and apoptosis were measured using RT-PCR, western blotting, immunofluorescence, and immunohistochemistry, respectively. TUNEL staining was used to assay apoptosis. MCAO/R-induced morphological changes were analyzed with Nissl staining and Hematoxylin-eosin staining. I/R-induced ERS was closely associated with an increase in miR-1873p and a decrease in seipin expression. miR-187-3p agomir further activated the ERS pathway and promoted apoptosis but decreased seipin expression levels; these effects were reversed by miR-187-3p antagomir. Moreover, seipin knockdown aggravated ERS in PC12 cells after OGD/R, and this change was rescued by seipin overexpression. miR-187-3p antagomir did not suppress ERS and apoptosis in seipin knockdown PC12 cells after OGD/R. Our findings demonstrate that the inhibition of miR1873p attenuated I/Rinduced cerebral injury by regulating seipin-mediated ERS.
Subject(s)
Brain Ischemia/metabolism , Endoplasmic Reticulum Stress/physiology , MicroRNAs/metabolism , Reperfusion Injury/metabolism , Animals , Apoptosis/physiology , GTP-Binding Protein gamma Subunits/metabolism , PC12 Cells , RatsABSTRACT
Stroke is a group of major diseases that cause death or disability in adults, with high incidence and lack of available therapeutic strategies. Although traditional Chinese medicine (TCM) has continuously achieved good effects in the therapy of stroke while there is still not convincing due to the limitation of blood-brain permeability, as well as the individual differences in usage and dosage. With the improvement of nanotechnology, TCM nanopreparation has gradually become a research hotspot in various fields due to its advantages in permeating the blood-brain barrier, targeting delivery, enhancing sustained-release drug delivery, changing the distribution in the body, and improving bioavailability. Zeolitic imidazolate framework-8 (ZIF-8) is an ideal nano-drug delivery system for adsorption, catalysis, and drug loading, which is a biocompatible metal-organic framework framed by 2-methylimidazole and zinc ions. At present, ZIF-8 was wildly used in the treatment of ischemic stroke. However, challenges remain persists for its clinical application, such as preparation technology, detection technology in vivo, targeting specificity, safety and stability, and so forth. Therefore, more efforts need to overcome the above problems to develop the application of TCM nanopreparations in the therapy of ischemia/reperfusion in the future.
