Spinal Cord Ischemia-reperfusion Injury Research Articles

Strategies for the development of in vitro models of spinal cord ischemia-reperfusion injury: Oxygen-glucose deprivation and reoxygenation

BackgroundIn vitro models tailored for spinal cord ischemia-reperfusion injury are pivotal for investigation of the mechanisms underlying spinal cord injuries. We conducted a two-phased study to identify the optimal conditions for establishing an in vitro model of spinal cord ischemia–reperfusion injury using primary rat spinal motor neurons. New methodIn the first phase, cell cultures were subjected to oxygen deprivation (OD) only, glucose deprivation (GD) only, or simultaneous deprivation of oxygen and glucose [oxygen-glucose deprivation (OGD)] for different durations (1, 2, and 6 h). In the second phase, different durations of re-oxygenation (1, 12, and 24 h) were applied after 1 h of OGD to determine the optimal duration simulating reperfusion injury. Results and comparison with existing method(s)GD for 6 h significantly reduced cell viability (91 % of control, P<0.001) and increase cytotoxicity (111 % of control, P<0.001). OGD for 1 h and 2 h, resulted in a significant decrease in cell viability (80 % of control P<0.001, respectively), and increase in cytotoxicity (130 % of control, P<0.001, respectively). Re-oxygenation for 1, 12, and 24 h worsened ischemic injury following 1 h of OGD (all P<0.05). ConclusionsOur results may provide a valuable guide to devise in vitro models of spinal cord ischemia–reperfusion injury using primary spinal motor neurons.

Time varying characteristic in somatosensory evoked potentials as a biomarker of spinal cord ischemic-reperfusion injury in rat.

Spinal cord ischemic-reperfusion injury (SCIRI) could occurs during surgical procedures without detection, presenting a complex course and an unfavorable prognosis. This may lead to postoperative sensory or motor dysfunction in areas innervated by the spinal cord, and in some cases, permanent paralysis. Timely detection of SCIRI and immediate waring can help surgeons implement remedial intervention to prevent irreversible spinal cord injury. Therefore, it is crucial to develop a precise and effective method for early detection of SCIRI. This study utilized rat models to simulate intraoperative SCIRI and employed somatosensory evoked potentials (SEP) for continuous monitoring during surgery. In this study, SEP signal changes were examined in six groups with varying severities of SCIRI and one normal control group. SEP signal changes were examined during operations in different groups and correlated with postoperative behavioral and histopathological data. The result demonstrated specific changes in SEP signals during SCIRI, termed as time-varying characteristics, which are associated with the duration of ischemia and subsequent reperfusion. Time-varying characteristics in SEP could potentially serve as a new biomarker for the intraoperative detection of SCIRI. This finding is significant for clinical surgeons to identify and guide early intervention of SCIRI timely. Additionally, this measurement is easily translatable to clinical application.

Ischemia-reperfusion injury after spinal cord decompressive surgery-An invivo rat model.

Although decompression surgery is the optimal treatment for patients with severe degenerative cervical myelopathy (DCM), some individuals experience no improvement or even a decline in neurological function after surgery, with spinal cord ischemia-reperfusion injury (SCII) identified as the primary cause. Spinal cord compression results in local ischemia and blood perfusion following decompression is fundamental to SCII. However, owing to inadequate perioperative blood flow monitoring, direct evidence regarding the occurrence of SCII after decompression is lacking. The objective of this study was to establish a suitable animal model for investigating the underlying mechanism of spinal cord ischemia-reperfusion injury following decompression surgery for degenerative cervical myelopathy (DCM) and to elucidate alterations in neurological function and local blood flow within the spinal cord before and after decompression. Twenty-four Sprague-Dawley rats were allocated to three groups: the DCM group (cervical compression group, with implanted compression material in the spinal canal, n = 8), the DCM-D group (cervical decompression group, with removal of compression material from the spinal canal 4 weeks after implantation, n = 8), and the SHAM group (sham operation, n = 8). Von Frey test, forepaw grip strength, and gait were assessed within 4 weeks post-implantation. Spinal cord compression was evaluated using magnetic resonance imaging. Local blood flow in the spinal cord was monitored during the perioperative decompression. The rats were sacrificed 1 week after decompression to observe morphological changes in the compressed or decompressed segments of the spinal cord. Additionally, NeuN expression and the oxidative damage marker 8-oxoG DNA were analyzed. Following spinal cord compression, abnormal mechanical pain worsened, and a decrease in forepaw grip strength was observed within 1-4 weeks. Upon decompression, the abnormal mechanical pain subsided, and forepaw grip strength was restored; however, neither reached the level of the sham operation group. Decompression leads to an increase in the local blood flow, indicating improved perfusion of the spinal cord. The number of NeuN-positive cells in the spinal cord of rats in the DCM-D group exceeded that in the DCM group but remained lower than that in the SHAM group. Notably, a higher level of 8-oxoG DNA expression was observed, suggesting oxidative stress following spinal cord decompression. This model is deemed suitable for analyzing the underlying mechanism of SCII following decompressive cervical laminectomy, as we posit that the obtained results are comparable to the clinical progression of degenerative cervical myelopathy (DCM) post-decompression and exhibit analogous neurological alterations. Notably, this model revealed ischemic reperfusion in the spinal cord after decompression, concomitant with oxidative damage, which plausibly underlies the neurological deterioration observed after decompression.

Propofol alleviates spinal cord ischemia-reperfusion injury by preserving PI3K/AKT/GIT1 axis.

Spinal cord ischemia-reperfusion injury (SCIRI) is a major contributor to neurological damage and mortality associated with spinal cord dysfunction. This study aims to explore the possible mechanism of Propofol and G-protein-coupled receptor-interacting protein 1 (GIT1) in regulating SCIRI in rat models. SCIRI rat models were established and injected with Propofol, over expression of GIT1 (OE-GIT1), or PI3K inhibitor (LY294002). The neurological function was assessed using Tarlov scoring system, and Hematoxylin & Eosin (H&E) staining was applied to observe morphology changes in spinal cord tissues. Cell apoptosis, blood-spinal cord barriers (BSCB) permeability, and inflammatory cytokines were determined by TdT-mediated dUTP Nick-End Labeling (TUNEL) staining, evans blue (EB) staining, and enzyme-linked immuno sorbent assay (ELISA), respectively. Reverse transcription-quantitative polymerase chain reaction and western blot were used to detect the expression levels of GIT1, endothelial nitric oxide synthase (eNOS), PI3K/AKT signal pathway and apoptosis-related proteins. SCIRI rats had decreased expressions of GIT1 and PI3K/AKT-related proteins, whose expressions can be elevated in response to Propofol treatment. LY294002 can also decrease GIT1 expression levels in SCIRI rats. Propofol can attenuate neurological dysfunction induced by SCIRI, decrease spinal cord tissue injury and BSCB permeability in addition to suppressing cell apoptosis and inflammatory cytokines, whereas further treatment by LY294002 can partially reverse the protective effect of Propofol on SCIRI. Propofol can activate PI3K/AKT signal pathway to increase GIT1 expression level, thus attenuating SCIRI in rat models.

Retracted: Astragalin Protects against Spinal Cord Ischemia Reperfusion Injury through Attenuating Oxidative Stress-Induced Necroptosis.

[This retracts the article DOI: 10.1155/2021/7254708.].

Administration with curcumin alleviates spinal cord ischemia-reperfusion injury by regulating anti-oxidative stress and microglia activation-mediated neuroinflammation via Nrf2/NF-κB axis.

Spinal cord ischemia-reperfusion injury (SCII) ranks as the common complication after aortic surgery, usually leading to devastating post-operative paraplegia. Microglia over-activation and neuronal cell loss are key pathological features of SCII. Curcumin is involved in several I/R injuries. However, its underlying mechanism in SCII remains elusive. Here, curcumin attenuated oxygen and glucose deprivation/reoxygenation (OGD/R)-induced oxidative injury in PC12 neuronal cells by increasing cell viability, inhibiting cell apoptosis, lactate dehydrogenase, malondialdehyde levels, but elevating anti-oxidative superoxide dismutase and glutathione peroxidase levels. Furthermore, curcumin restrained OGD/R-evoked microglia M1 activation by decreasing microglia M1 polarization marker IBA-1 and iNOS transcripts. Moreover, the increased inflammatory cytokine levels of TNF-α and IL-6 in microglia under OGD/R conditions were suppressed after curcumin treatment. Importantly, neuronal cells incubated with a conditioned medium from OGD/R-treated microglia exhibited lower cell viability and higher apoptotic ratio, which were overturned when microglia were treated with curcumin. Intriguingly, curcumin could inhibit the activation of the NF-κB pathway by Nrf2 enhancement in OGD/R-treated PC12 cells and microglia. Notably, targeting Nrf2 signaling reversed the protective efficacy of curcumin against OGD/R-evoked oxidative insult in neuronal, microglia M1 activation, inflammatory response, and microglial activation-evoked neuronal death. In vivo, curcumin improved histopathologic injury and neurologic motor function in SCII rats and attenuated oxidative stress, microglia activation and neuroinflammation in spinal cord tissues, and activation of the Nrf2/NF-κB pathway. Thus, curcumin may alleviate SCII by mitigating I/R-evoked oxidative injury in neuron and microglia activation-induced neuroinflammation and neuron death through Nrf2/NF-κB signaling, supporting a promising therapeutic agent for SCII.

A slow-releasing donor of hydrogen sulfide inhibits neuronal cell death via anti-PANoptosis in rats with spinal cord ischemia‒reperfusion injury

BackgroundSpinal cord ischemia‒reperfusion injury (SCIRI) can lead to paraplegia, which leads to permanent motor function loss. It is a disastrous complication of surgery and causes tremendous socioeconomic burden. However, effective treatments for SCIRI are still lacking. PANoptosis consists of three kinds of programmed cell death, pyroptosis, apoptosis, and necroptosis, and may contribute to ischemia‒reperfusion-induced neuron death. Previous studies have demonstrated that hydrogen sulfide (H2S) exerts a neuroprotective effect in many neurodegenerative diseases. However, whether H2S is anti-PANoptosis and neuroprotective in the progression of acute SCIRI remains unclear. Thus, in this study we aimed to explore the role of H2S in SCIRI and its underlying mechanisms.MethodsMeasurements of lower limb function, neuronal activity, microglia/macrophage function histopathological examinations, and biochemical levels were performed to examine the efficacy of H2S and to further demonstrate the mechanism and treatment of SCIRI.ResultsThe results showed that GYY4137 (a slow-releasing H2S donor) treatment attenuated the loss of Nissl bodies after SCIRI and improved the BBB score. Additionally, the number of TUNEL-positive and cleaved caspase-3-positive cells was decreased, and the upregulation of expression of cleaved caspase-8, cleaved caspase-3, Bax, and Bad and downregulation of Bcl-2 expression were reversed after GYY4137 administration. Meanwhile, both the expression and activation of p-MLKL, p-RIP1, and p-RIP3, along with the number of PI-positive and RIP3-positive neurons, were decreased in GYY4137-treated rats. Furthermore, GYY4137 administration reduced the expression of NLRP3, cleaved caspase-1 and cleaved GSDMD, decreased the colocalization NeuN/NLRP3 and Iba1/interleukin-1β-expressing cells, and inhibited proinflammatory factors and microglia/macrophage polarization.ConclusionsH2S ameliorated spinal cord neuron loss, prevented motor dysfunction after SCIRI, and exerted a neuroprotective effect via the inhibition of PANoptosis and overactivated microglia-mediated neuroinflammation in SCIRI.

Effect of captopril on paraplegia caused by spinal cord ischemia-reperfusion injury in rats.

This study investigated the effect of captopril (Cap) on spinal cord ischemia-reperfusion injury (SCII) in rats. Twenty-four adults male Wistar rats were randomly divided into four groups of six animals each: spinal cord ischemia-reperfusion (SCI-R) with Cap (SCI-R + Cap), SCI-R, sham-operated with Cap (SHAM + Cap), and SHAM. The 24 hr and 90 min before ischemia induction, Cap was administered intragastrically (100 mg kg-1) to the SHAM + Cap and SCI-R + Cap groups. Abdominal aortic clamping was performed in the SCI-R and SCI-R + Cap groups for 40 min. Hindlimb motor function was evaluated using the Tarlov Scale at 4, 6, 12, 24, 48, and 60 hr after SCII. The malondialdehyde (MDA), the ferric-reducing ability of plasma (FRAP) and prooxidant-antioxidant balance (PAB) values were also measured. Throughout the study period, the SCI-R group had significantly lower motor function scores compared to the other groups. The MDA and PAB levels were higher and the FRAP value was lower in the SCI-R group compared to in the SHAM group. The SCI-R + Cap had higher motor function scores compared to the SCI-R group at all time points. There were no significant differences in MDA concentration, FRAP and PAB values between the SCI-R + Cap and SCI-R groups. Captopril may act as a protective agent against SCII in rats based on hind limb motor function assessment.

Amantadine's Neuroprotective Effects in Rabbit Spinal Cord Ischemia/Reperfusion Model.

To examine the effects of amantadine, a drug with neuroprotective and anti-inflammatory activities on oxidative stress, tissue necrosis, apoptosis, and neurological recovery in an experimental rabbit spinal cord ischemia-reperfusion injury (SCIRI) model. A total of 32 rabbits were randomized into five groups: control, ischemia, vehicle, methylprednisolone (MP), and amantadine (AMT) (n=8/each). At 24th-hour neurological examination was performed, spinal cord tissues were collected, and biochemical and histopathological examinations were performed. When ischemia and vehicle groups were compared with control group, significant increase was seen in serum and tissue caspase-3, malondialdehyde (MDA), and myeloperoxidase (MPO) levels (p < 0.001); significant decrease was seen in serum and tissue catalase (CAT) levels (p < 0.001); and significant increase was seen in serum xanthine oxidase (XO) levels (p < 0.001). When the ischemia group and the MP and AMT groups were compared, low serum and tissue caspase-3 levels (p < 0.001), high serum and tissue CAT levels (p < 0.001), significantly low serum XO levels (p < 0.001), low serum and tissue MDA levels (p < 0.05) and tissue MPO levels (p < 0.001) were found. Both AMT and MP groups showed decreased histopathological score and higher number of normal neurons (p < 0.001) compared to ischemia group. Both AMT and MP showed better modified Tarlov scores compared to the ischemia group (p < 0.001). Our study found that AMT had antioxidant, anti-inflammatory, anti-apoptotic, and neuroprotective effects on SCIRI. We used biochemical, microscopic, and ultrastructural approaches to demonstrate these effects. AMT might be a candidate medication for SCIRI prophylaxis and treatment.

Protective Effect of Gum Arabic on Spinal Cord Ischemia-reperfusion Injury in Rats

Protective Effect of Gum Arabic on Spinal Cord Ischemia-reperfusion Injury in Rats

Ferrostatin-1 improves neurological impairment induced by ischemia/reperfusion injury in the spinal cord through ERK1/2/SP1/GPX4

Spinal cord ischemia/reperfusion injury (SCIRI) induced by artificial aortic occlusion for a while during aortic surgery is a serious complication, leading to paraplegia and even death. Ferroptosis in the nervous system has been confirmed to contribute to neuronal death induced by SCIRI. Therefore, we investigated the therapeutic benefits of ferrostatin-1 (Fer-1, a ferroptosis inhibitor) and explored the mechanism and target of Fer-1 in SCIRI. Our results demonstrate that intrathecal injection of Fer-1 had a strong anti-SCIRI effect, improved ferroptosis-related indices, increased neurological function scores and motor neuron counts, and reduced BSCB leakage and neuroinflammation levels in the anterior horn. We found that SCIRI significantly elevated the levels of several important proteins, including SP1, p-ERK1/2/ERK1/2, COX2, TFR1, SLC40A1, SLC7A11, cleaved Caspase 3, GFAP, and Iba1, while reducing FTH1 and GPX4 protein expression, with no effect on ACSL4 expression. Fer-1 effectively ameliorated the ferroptosis-related changes in these proteins induced by SCIRI. However, for p-ERK1/2 and SP1, Fer-1 not only failed to reduce their expression but also significantly enhanced it. Fer-1 was injected into sham operation rats, abnormal increases in p-ERK1/2/ERK1/2 and SP1 were observed, along with an increase in GPX4. Fluorescent double labeling revealed that SP1 and GPX4 were expressed in neurons and astrocytes. Inhibitors of the ERK pathway (SCH772984) and siRNA against SP1 (AV-sh-SP1) significantly decreased the increase in SP1 and GPX4 protein levels, fluorescent density of SP1 and GPX4 in neurons, and the number of SP1-positive and GPX4-positive neurons induced by Fer-1. SCH772984 but not AV-sh-SP1 significantly reversed the decrease in GFAP and Iba1 induced by Fer-1. In conclusion, our results indicate that Fer-1 inhibited ferroptosis in spinal cord anterior horn neurons, improving neurological impairment and BSCB damage after SCIRI through the ERK1/2/SP1/GPX4 signaling pathway in rats.

Protective effect of valproic acid on ischemia-reperfusion induced spinal cord injury in a rat model

Purpose This study aims to determine the anti-inflammatory, antioxidant, and anti-apoptotic effects of valproic acid (VPA) on rat spinal cord tissue in ischemia-reperfusion (IR) injury model created by abdominal aorta occlusion. Materials and Methods Sprague Dawley rat (male sex) weighing 190–260 g divided into four experimental groups: control only underwent laparotomy, sham group, pre-IR injury (200 mg/kg dose), and post-IR injury (300 mg/kg) VPA. We measured serum levels of TNF-α, IL-6, IL-1β, IL-18, Total Oxidant Status (TOS) and Total Antioxidant Status (TAS), and serum Oxidative Stress Index (OSI) ratio, and tissue expression of Bax and Bcl2, Caspase3, and Bax/Bcl2 ratio. Results Serum IL-18 was higher in the sham than the control group(P = 0.001), and there were declines in the pre-IR treatment (P = 0.002) and the post-IR treatment when compared to sham (P = 0.001). Despite these reductions, IL-18 expression levels in both the pre- and post-IR treatment groups were higher than in the control group (P = 0.001 & P = 0.003). The favorable effects of pre-IR VPA administration on immunohistochemical biomarkers were superior to post-IR VPA administration. Conclusions Comparative analyses between prophylactic VPA administration and post-IR interventions revealed congruence in their anti-inflammatory and anti-apoptotic ramifications. VPA can reduce spinal cord IR injury in an aortic occlusion model of rats.

Diallyl trisulfide improves spinal cord ischemia–reperfusion injury damage by activating AMPK to stabilize mitochondrial function

BackgroundSpinal cord ischemia–reperfusion injury (SCII) is a catastrophic event, which can cause paraplegia in severe cases. In the reperfusion stage, oxidative stress was up-regulated, which aggravated the injury and apoptosis of neurons. As the main active ingredient of garlic, diallyl trisulfide (DATS) displays strong antioxidant capacity. However, it is unknown whether DATS can protect the neurons of SCII.Materials and methodsIn this study, the descending aorta at the distal end of the left subclavian artery was ligated and perfused again after 14 min. Samples including blood and spinal cord (L2–L5) were taken 24 h later for morphological and biochemical examination.ResultsAfter SCII, the rats showed motor dysfunction, increase apoptosis, malondialdehyde content, mitochondrial biogenesis and dynamic balance disorder. After the application of DATS, the adenosine monophosphate activated protein kinase (AMPK) was activated, the mitochondrial damage was improved, the oxidative stress was weakened, and the neuronal damage was recovered to some extent. However, the addition of compound C significantly weakened the protective effect of DATS.ConclusionOxidative stress caused by mitochondrial damage was one of the important mechanisms of neuronal damage in SCII. DATS could activate AMPK, stabilize mitochondrial biogenesis and dynamic balance, and reduce neuronal damage caused by oxidative stress.

Transplantation of MiR-28-5p-Modified BMSCs Promotes Functional Recovery After Spinal Cord Injury

Traumatic spinal cord injury (TSCI) is a prevalent central nervous system condition that imposes a significant burden on both families and society, affecting more than 2 million people worldwide. Recently, there has been increasing interest in bone marrow mesenchymal stem cell (BMSC) transplantation as a promising treatment for spinal cord injury (SCI) due to their accessibility and low immunogenicity. However, the mere transplantation of BMSCs has limited capacity to directly participate in the repair of host spinal cord nerve function. MiR-28-5p, identified as a key differentially expressed miRNA in spinal cord ischemia–reperfusion injury, exhibits differential expression and regulation in various neurological diseases. Nevertheless, its involvement in this process and its specific regulatory mechanisms in SCI remain unclear. Therefore, this study aimed to investigate the potential mechanisms through which miR-28-5p promotes the neuronal differentiation of BMSCs both in vivo and in vitro. Our results indicate that miR-28-5p may directly target Notch1, thereby facilitating the neuronal differentiation of BMSCs in vitro. Furthermore, the transplantation of lentivirus-mediated miR-28-5p-overexpressed BMSCs into SCI rats effectively improved footprint tests and Basso, Beattie, and Bresnahan (BBB) scores, ameliorated histological morphology (hematoxylin–eosin [HE] and Nissl staining), promoted axonal regeneration (MAP2 and growth-associated protein 43 [GAP43]), and facilitated axonal remyelination (myelin basic protein [MBP]). These findings may suggest that miR-28-5p-modified BMSCs could serve as a therapeutic target to enhance the behavioral and neurological recovery of SCI rats.

Melatonin Exerts an Anti-Panoptoic Role in Spinal Cord Ischemia-Reperfusion Injured Rats.

Paraplegia is a serious consequence of spinal cord ischemia-reperfusion (SCIR) injury, which leads to neuron death and permanent loss of motor function. However, there is no effective treatment for SCIR. Melatonin exerts a neuroprotective effect in neurodegenerative diseases. However, whether pyroptosis, apoptosis, and necroptosis (PANoptosis) is the primary cause of the massive neural death in SCIR is unknown, and if melatonin exhibits anti-PANoptotic effect in rescuing the disastrous damage is to be decided. This study indicates that melatonin confers neuroprotection in SCIR, attenuating the loss of Nissl body and improving Basso, Beattie & Bresnahan locomotor rating scale scores. Specifically, the apoptotic hallmarks in neurons are increased in SCIR injured spinal cord compared to the sham group. The upregulated trend is reversed by melatonin while the effect of melatonin is abolished by the administration of luzindole, a selective melatonin receptor antagonist. Moreover, similar patterns are found in the necroptotic markers in neurons, the pyroptotic indicators, and the interleukin-1β staining in microglia. In conclusion, PANoptosis may underlie the mass neural death and paraplegia in SCIR, and melatonin confers neuroprotection to the spinal cord via inhibiting PANoptosis.

Exosomes derived from human placental mesenchymal stem cells in combination with hyperbaric oxygen synergically alleviates spinal cord ischemia-reperfusion injury

Spinal cord ischemia-reperfusion injury (IR) is a terrible non-traumatic injury that occurs after abdominal aortic occlusion and causes serious damage to neurological function. Several treatment strategies have been suggested for IR, but they were not unable to effectively improve these conditions. Herein we investigated whether exosomes derived from human placental mesenchymal stem cells (hpMSCs-Exos) in combination with hyperbaric oxygen (HBO) could alleviate injury and promote recovery in IR rats. Eighty male Sprague-Dawley rats were randomly allocated into five equal groups. In addition to the control group that only underwent laparotomy, IR animals were planned into four groups as follows: IR group; IR-Exos group; IR-HBO group; and IR-Exos + HBO group. Neurological function evaluated before, 6 h, 12 h, 24 h, and 48 h after injury. After the last neurological evaluation, tissue samples were obtained for stereological, biochemical, and molecular assessments. Our results indicated that the neurological function scores (MDI), the numerical density of neurons, the levels of antioxidative factors (GSH, SOD, and CAT), and anti-inflammatory cytokine (IL-10) were considerably greater in treatment groups than in the IR group, and these changes were more obvious in the IR-Exos + HBO ones. This is while the numerical density of glial cells, the levels of an oxidative factor (MDA) and inflammatory cytokines (IL-1β, TNF-α, and IL-18), as well as the expression of an apoptotic protein (caspase-3) were meaningfully decreased in treatment groups, especially IR-Exos + HBO group, compared to the IR group. Generally, it was found that co-administration of hpMSCs-Exos and HBO has synergistic neuroprotective effects in the rats undergoing IR.

Mechanisms of polydatin against spinal cord ischemia–reperfusion injury based on network pharmacology, molecular docking and molecular dynamics simulation

BackgroundPolydatin has shown considerable pharmacological activities in ischemia–reperfusion injuries of various organs. However, its effects and mechanisms in spinal cord ischemia–reperfusion injury have not been fully established. In this study, the mechanisms of polydatin against spinal cord ischemia–reperfusion injury were investigated via network pharmacology, molecular docking and molecular dynamics simulation. MethodsSpinal cord ischemia–reperfusion injury-related targets were obtained from the GeneCards database, while polydatin-related action targets were obtained from the CTD and SwissTarget databases. A protein–protein interaction network of potential targets was constructed using the String platform. After selecting the potential key targets, GO functional enrichment and KEGG pathway enrichment analyses were performed via the Metascape database, and a network map of “drug-target-pathway-disease” constructed. The relationships between polydatin and various key targets were assessed via molecular docking. Molecular dynamics simulation was conducted for optimal core protein–compound complexes obtained by molecular docking. ResultsTopological analysis of the PPI network revealed 14 core targets. GO functional enrichment analysis revealed that 435 biological processes, 12 cell components and 29 molecular functions were enriched while KEGG pathway enrichment analysis revealed 91 enriched signaling pathways. Molecular docking showed that polydatin had the highest binding affinity for MAPK3, suggesting that MAPK3 is a key target of polydatin against spinal cord ischemia–reperfusion injury. Molecular dynamics simulations revealed good binding abilities between polydatin and MAPK3. ConclusionsPolydatin exerts its effects on spinal cord ischemia–reperfusion injury through multiple targets and pathways. MAPK3 may be a key target of polydatin in spinal cord ischemia–reperfusion injury.

EGFR-Activated JAK2/STAT3 Pathway Confers Neuroprotection in Spinal Cord Ischemia-Reperfusion Injury: Evidence from High-Throughput Sequencing and Experimental Models.

This study aimed to investigate the molecular mechanisms underlying spinal cord ischemia-reperfusion (SCI/R) injury. Through RNA-Seq high-throughput sequencing and bioinformatics analysis, we found that EGFR was downregulated in the spinal cord of SCI/R mice and may function via mediating the JAK2/STAT3 signaling pathway. In vitro cell experiments indicated that overexpression of EGFR activated the JAK2/STAT3 signaling pathway and reduced neuronal apoptosis levels. In vivo animal experiments further confirmed this conclusion, suggesting that EGFR inhibits SCI/R-induced neuronal apoptosis by activating the JAK2/STAT3 signaling pathway, thereby improving SCI/R-induced spinal cord injury in mice. This study revealed the molecular mechanisms of SCI/R injury and provided new therapeutic strategies for treating neuronal apoptosis.

Xenon postconditioning attenuates neuronal injury after spinal cord ischemia/reperfusion injury by targeting endoplasmic reticulum stress-associated apoptosis.

The purpose of the study is to explore the underlying mechanisms of xenon (Xe) which protects against spinal cord ischemia/reperfusion injury (SCIRI). A SCIRI rat model was induced by abdominal artery occlusion for 85 min and reperfusion. Xe postconditioning (50% Xe) was administered 1 h after 1 h of reperfusion. At reperfusion time points (2, 4, 6, and 24 h), rats were treated with spinal cord scans by MRI to assess the time of peak spinal cord injury after SCIRI. Subsequently, endoplasmic reticulum (ER) stress inhibitor sodium 4-phenylbutyrate (4-PBA) was administered by daily intraperitoneal injection (50 mg/kg) for 5 days before SCIRI. At 4 h after reperfusion, motor function, immunofluorescence staining, hematoxylin and eosin (HE) staining, Nissl staining, TUNEL staining, real-time reverse transcription polymerase chain (RT-PCR) reaction, and western blot analyses were performed to investigate the protective effects of Xe against SCIRI. In the rat I/R model, spinal cord edema peaked at reperfusion 4 h. SCIRI activated ER stress, which was located in neurons. Xe postconditioning remarkably alleviated hind limb motor function, reduced neuronal apoptosis rate, increased the number of normal neurons, and inhibited the expression of ER stress-related protein in spinal cord. Furthermore, the administration of the ER stress inhibitor 4-PBA strongly decreased ER stress-induced apoptosis following SCIRI. Xe postconditioning inhibits ER stress activation, which contributes to alleviate SCIRI by suppressing neuronal apoptosis.

SYVN1 attenuates ferroptosis and alleviates spinal cord ischemia–reperfusion injury in rats by regulating the HMGB1/NRF2/HO-1 axis

BackgroundThe ferroptosis of neurons is an important pathological mechanism of spinal cord ischemia reperfusion injury (SCIRI). Previous studies showed that synoviolin 1 (SYVN1) is a good prognostic marker of neurodegenerative diseases, but its mechanism is still unclear. This study aims to explore the role of SYVN1 in the ferroptosis of neurons and to clarify its internal mechanism. MethodsRat primary spinal cord neurons were treated with oxygen-glucose deprivation (OGD) for 1, 4 or 8 h, and then cell viability, ROS and MDA levels, glutathione peroxidase (GSH-Px) activity, and the expression of ferroptosis-related proteins GPX4, FTH1 and PTGS2 were detected. OGD/R-induced neurons were transfected with pcDNA-SYVN1 or si-HMGB1, and then cell functions were detected. Transmission electron microscope (TEM) was used to detect cell ferroptosis. The interplay between SYVN1 and high mobility group box 1 (HMGB1) was confirmed with Co-immunoprecipitation (Co-IP) assay. The stability of HMGB1 was measured by ubiquitination assay. Also, cells were treated with pcDNA-SYVN1 or together with ubiquitination inhibitor MG132, as well as treated with pcDNA-SYVN1 and pcDNA-HMGB1 or together with NRF2 activator dimethyl fumarate (DMF), and then Western blotting was used to detect the expression of HMGB1, nuclear NRF2 and HO-1 proteins. In addition, SD rats were occluded left common carotid artery and aortic arch to establish a SCIRI rat model. And rats were injected intrathecal with adenovirus-mediated SYVN1 overexpression vector (Ad-SYVN1, 2 μL, virus titer 5 × 1013 transduction unit [TU]/mL) to overexpress SYVN1. The motion function of rats was quantified using the Basso Rat Scale (BMS) for Locomotion. The ferroptosis and the number of neurons in the spinal cord tissue of rats were detected. ResultsSYVN1 overexpression inhibited ferroptosis of SCIRI rats and OGD/R-treated primary spinal cord neurons, and down-regulated the expression of HMGB1. In terms of mechanism, the binding of SYVN1 and HMGB1 promoted the ubiquitination and degradation of HMGB1, and negatively regulated the expression of HMGB1. Moreover, under OGD/R conditions, MG132 treatment or HMGB1 overexpression eliminated the inhibitory effect of SYVN1 overexpression on the ferroptosis of neurons and the activation of the NRF2/HO-1 pathway, and DMF treatment abolished the inhibition of HMGB1 overexpression on the NRF2/HO-1 pathway. Finally, in vivo experiments showed that SYVN1 overexpression could alleviate the spinal cord ischemia–reperfusion injury in rats by down-regulating HMGB1 and promoting the activation of the NRF2/HO-1 pathway. ConclusionSYVN1 regulates ferroptosis through the HMGB1/NRF2/HO-1 axis to prevent spinal cord ischemia–reperfusion injury.