Expression Of Nogo-A Research Articles

Down-Regulation of Nogo-A and PirB in Ischemic Cortex with Remote Ischemic Preconditioning in Mice

Objectives: The present study aims to investigate the effect of Limb Remote Ischemic Preconditioning (LRIP) on the expression of Nogo-A and PirB in the cortex of mice with focal cerebral ischemia, and related pathways involving in axonal regeneration and neurological function recovery after cerebral ischemia. Methods: Adult male C57/BL6 mice were divided into sham-operated (sham), transient Middle Cerebral Artery Occlusion (MCAO), LRIP and anti- PirBAb treatment group. Samples were collected 48h after cerebral ischemia. The histopathologic changes were assessed by 1,3,5-Triphenyl-2H-Tetrazolium Chloride (TTC), and Hematoxylin and Eosin (HE) staining and TUNEL method. The expression of Nogo-A and PirB were determined by immunofluorescence, RT-PCR and Western blot respectively. Results: TTC staining showed that LRIP treatment reduced the infarct size of mice and anti-PirBAb treatment further decline the infarct size, which was accompanied with the decline of neurological deficit score and reduction of neuronal damage. LRIP treatment also reduced the TUNEL positive cells induced by MCAO and anti-PirBAb treatment further strengthened the effect of LRIP. Except sham group, the expressions of Nogo-A and PirB in other three groups all increased with varying degrees, among which MCAO group was the highest, LRIP group was the second and the anti-PirBAb group was the lowest. The expressions of growth associated protein 43 (GAP43) showed opposite tendency. Conclusions: LRIP plays beneficial influence on cerebral ischemia. LRIP and PirB inhibition combination has a better protective effect on nervous system after cerebral ischemia in mice.

Anti-apoptotic efficacy of Qingnao Yizhi formula in hypoxia/reoxygenation primary cortical neurons through the promotion of synaptic plasticity by modulating the NogoA-Nogo receptor/Rho-Rho kinase signaling pathway.

To evaluate the anti-apoptotic efficacy of Qingnao Yizhi formula (，QNYZ) in cultured cerebral cortical neuronal cells (CNCs) and the regulation of the NogoA-Nogo receptor (NgR)/Rho-Rho kinase (ROCK) signaling pathway. Primary cultured CNCs were randomly divided into the following groups: normal control group (N-C), hypoxia-reoxygenation group (H/R), high-dose QNYZ group (Q-H), low-dose QNYZ group (Q-L) butylphthalide (NBP) group, and Y-27632 (a selective ROCK transduction pathway inhibiter) group. Except those in the N-C group, CNCs were placed in hypoxic conditions for 24 h and then in reoxygenation conditions for 24 h. Cell media was changed every 48 h, and various assays were performed on the 7th day. Cell viability was evaluated by measuring mitochondrial dehydrogenase activity, using a CCK-8 assay, in triplicate. Synapsin (SYN) protein concentrations were evaluated by enzyme-linked immunosorbent assay. NogoA and RhoA protein expression were evaluated through Western blotting. The gene expression of NogoA, NgR, RhoA, and ROCK was evaluated by reverse transcription-polymerase chain reaction. Cell apoptosis was measured using a terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling assay. Compared with the N-C group, the cell viability of the H/R group decreased significantly (P < 0.05). The cell viability values for the Q-H and Q-L groups increased compared with that for the H/R group, and the difference was significant for the Q-H group (P < 0.05). The NogoA and RhoA protein levels and the NogoA, NgR, RhoA, and ROCK mRNA expression levels increased in the H/R group, compared with the N-C group, and decreased significantly in the Q-H and Q-L groups (P < 0.05) and in the Y-27632 group (P < 0.05) compared with the H/R group. The SYN levels in the Q-H, Q-L, and NBP groups significantly increased compared with that in the H/R group (P < 0.05). Compared with the H/R group, the numbers of apoptotic cells in the Q-H, Q-L, and NBP groups significantly decreased (P < 0.05). The presented study demonstrated that QNYZ exerted anti-apoptotic effects on H/R-induced CNCs, possibly through the modulation of the NogoA-NgR/Rho-ROCK signaling pathway and the promotion of synaptic plasticity in H/R CNCs.

Neurite Outgrowth Inhibitor (NogoA) Is Upregulated in White Matter Lesions of Complex Cortical Malformations.

Complex cortical malformations (CCMs), such as hemimegalencephaly and polymicrogyria, are associated with drug-resistant epilepsy and developmental impairment. They share certain neuropathological characteristics including mammalian target of rapamycin (mTOR) activation and an atypical number of white matter neurons. To get a better understanding of the pathobiology of the lesion architecture, we investigated the role of neurite outgrowth inhibitor A (NogoA), a known regulator of neuronal migration. Epilepsy surgery specimens from 16 CCM patients were analyzed and compared with sections of focal cortical dysplasia IIB (FCD IIB, n = 22), tuberous sclerosis complex (TSC, n = 8) as well as healthy controls (n = 15). Immunohistochemistry was used to characterize NogoA, myelination, and mTOR signaling. Digital slides were evaluated automatically with ImageJ. NogoA staining showed a significantly higher expression within the white matter of CCM and FCD IIB, whereas cortical tubers presented levels similar to controls. Further analysis of possible associations of NogoA with other factors revealed a positive correlation with mTOR and seizure frequency. To identify the main expressing NogoA cell type, double staining revealed dysmorphic neuronal white matter cells. Increased NogoA expression is associated with profound inhibition of neuritic sprouting and therefore contributes to a decrease in neuronal network complexity in CCM patients.

Nogo-A regulates myogenesis via interacting with Filamin-C

Among the three isoforms encoded by Rtn4, Nogo-A has been intensely investigated as a central nervous system inhibitor. Although Nogo-A expression is increased in muscles of patients with amyotrophic lateral sclerosis, its role in muscle homeostasis and regeneration is not well elucidated. In this study, we discovered a significant increase in Nogo-A expression in various muscle-related pathological conditions. Nogo−/− mice displayed dystrophic muscle structure, dysregulated muscle regeneration following injury, and altered gene expression involving lipid storage and muscle cell differentiation. We hypothesized that increased Nogo-A levels might regulate muscle regeneration. Differentiating myoblasts exhibited Nogo-A upregulation and silencing Nogo-A abrogated myoblast differentiation. Nogo-A interacted with filamin-C, suggesting a role for Nogo-A in cytoskeletal arrangement during myogenesis. In conclusion, Nogo-A maintains muscle homeostasis and integrity, and pathologically altered Nogo-A expression mediates muscle regeneration, suggesting Nogo-A as a novel target for the treatment of myopathies in clinical settings.

Effects of Nogo-A and its receptor on the repair of sciatic nerve injury in rats.

Regeneration of injured peripheral nerves is an extremely complex process. Nogo-A (neurite outgrowth inhibitor-A) inhibits axonal regeneration by interacting with Nogo receptor in the myelin sheath of the central nervous system (CNS). The aim of this study was to investigate the effects of Nogo-A and its receptor on the repair of sciatic nerve injury in rats. Sprague-Dawley rats (n=96) were randomly divided into 4 groups: control group (control), sciatic nerve transection group (model), immediate repair group (immediate repair), and delayed repair group (delayed repair). The rats were euthanized 1 week and 6 weeks after operation. The injured end tissues of the spinal cord and sciatic nerve were obtained. The protein expressions of Nogo-A and Nogo-66 receptor (NgR) were detected by immunohistochemistry. The protein expressions of Nogo-A, NgR, and Ras homolog family member A (RhoA) were detected by western blot. At 1 week after operation, the pathological changes in the immediate repaired group were less, and the protein expressions of Nogo-A, NgR, and RhoA in the spinal cord and sciatic nerve tissues were decreased (P<0.05) compared with the model group. After 6 weeks, the pathological changes in the immediate repair group and the delayed repair group were alleviated and the protein expressions decreased (P<0.05). The situation of the immediate repair group was better than that of the delayed repair group. Our data suggest that the expression of Nogo-A and its receptor increased after sciatic nerve injury, indicating that Nogo-A and its receptor play an inhibitory role in the repair process of sciatic nerve injury in rats.

Molecular effects of curcumin on the experimental autoimmune encephalomyelitis.

Experimental autoimmune encephalomyelitis (EAE) is an animal model of multiple sclerosis (MS). Previous studies have shown that myelin degradation during MS and EAE resulted in reduced expression of some of the proteins, e.g., the MBP (myelin basic protein), and increased expression of genes such as iNOS (Inducible nitric oxide synthase) and NOGO-A in the affected patients. In the present study, EAE was induced by immunizing Wistar rats (n=12) with homogenized spinal cord of guinea pig and Freund's complete adjuvant. Curcumin is an active ingredient in turmeric with anti-inflammatory properties, which has been studied in this article. In this study, the effect of curcumin administration on the change of the expression of MBP, NOGO-A, and iNOS genes was evaluated using the RT-PCR (Reverse transcription-polymerase chain reaction) technique. The obtained results indicated it could be concluded that curcumin was able to improve EAE by increasing the amount of MBP gene expression and reducing the intensity of NOGO-A expression.

Postnatal expression and possible function of RANK and RANKL in the murine inner ear.

The bone encasing the inner ear, known as the otic capsule, is unique because it remodels little postnatally compared to other bones in the body. Previous studies established that osteoprotegerin (OPG) in the inner ear inhibits otic capsule remodeling. OPG acts as a decoy receptor of receptor activator of nuclear factor κB ligand (RANKL) to disrupt the interaction between RANKL and RANK, the primary regulators of bone metabolism. Here we studied the expression and function of RANK and RANKL in the murine cochlea. Using a combination of in situ hybridization, real-time quantitative RT-PCR, and western blot, we demonstrate that Rankl and Rank genes and their protein products are expressed in the intracochlear soft tissues and the otic capsule in a developmentally regulated manner. Using a culture of neonatal murine cochlear neurons, we show that the interaction between RANK and RANKL inhibits neurite outgrowth in these neurons, and is associated with upregulation of NOGO-A expression. Taken together, our results suggest that, in addition to regulating otic capsule bone remodeling, RANK and RANKL expressed by intracochlear soft tissues may also regulate spiral ganglion neuron function by affecting neurite outgrowth.

Effects of repetitive magnetic stimulation on motor function and GAP43 and 5-HT expression in rats with spinal cord injury.

ObjectivesSpinal cord injury (SCI) is a disabling central nervous system disorder. This study aimed to explore the effects of repetitive trans-spinal magnetic stimulation (rTSMS) of different spinal cord segments on movement function and growth-associated protein-43 (GAP43) and 5-hydroxytryptamine (5-HT) expression in rats after acute SCI and to preliminarily discuss the optimal rTSMS treatment site to provide a theoretical foundation and experimental evidence for clinical application of rTSMS in SCI.MethodsA rat T10 laminectomy SCI model produced by transient application of an aneurysm clip was used in the study. The rats were divided into group A (sham surgery), group B (acute SCI without stimulation), group C (T6 segment stimulation), group D (T10 segment stimulation), and group E (L2 segment stimulation).ResultsIn vivo magnetic stimulation protected motor function, alleviated myelin sheath damage, decreased NgR and Nogo-A expression levels, increased GAP43 and 5-HT expression levels, and inhibited terminal deoxynucleotidyl transferase dUTP nick end labeling-positive cells and apoptosis-related protein expression in rats at 8 weeks after the surgery.ConclusionsThis study suggests that rTSMS can promote GAP43 and 5-HT expression and axonal regeneration in the spinal cord, which is beneficial to motor function recovery after acute SCI.

Palmitoylethanolamide-Mediated Inhibition of Nogo-a Signaling Attenuates Pain in Sickle Mice

Sickle cell disease (SCD) is characterized by multiple comorbidities including pain. SCD patients often use cannabinoids to alleviate pain, but their psychoactive effects and social stigma impose major challenges. Strategies to elevate endogenous cannabinoids (eCBs) are devoid of such challenges, but pharmacologic approaches showed adverse-effects in clinical trials. Therefore, we examined the potential of non-pharmacologic integrative approaches to elevate eCBs. Enriched high-energy diet has been shown to increase levels of eCBs (Argueta et al., Front Physiol 2019) and when combined with companionship reduced hyperalgesia in sickle mice (Tran et al., Blood 2016). We hypothesized that enriched diet and companionship would enhance eCBs without adverse effects and reduce hyperalgesia by inhibiting peripheral and central pro-nociceptive mechanisms. We fed male homozygous-BERK (sickle) mice, regular Rodent Diet (RD; 2018, Harlan) or customized high calorie enriched Sickle Mouse Diet (SD; 59M3, TestDiet), housed with or without a female companion (C+ or C-, respectively) for 3-weeks. RD/SD contain 18.6/26.4% protein, 6.2/11.1% fat, 24/27.5% carbohydrates and 18/26% kcal/g, respectively; and SD contains higher minerals, vitamins and ω-3 fatty acids compared to RD. Control HbAA-BERK and sickle mice were divided into 4 groups: [i] R/C-, RD, no companion, [ii] S/C-, SD without companion, [iii] R/C+, RD with companion, and [iv] S/C+, SD with companion. After 3-weeks of treatment, spinal cord eCBs were analyzed using targeted lipid quantitation with liquid chromatography mass spectrometry (LCMS). We observed a 20% decrease in palmitoylethanolamide (PEA), in sickle mice compared to control mice, in R/C- group (p&amp;lt;0.05). Further, we observed increased spinal PEA in S/C+ compared to R/C- sickle mice (~40%, p&amp;lt;0.05), which was concomitant with reduced mechanical, heat, and cold hyperalgesia in S/C+ sickle mice group (~80%, p&amp;lt;0.001; ~60%, p&amp;lt;0.01; &amp; ~30%, p&amp;lt;0.001, respectively). Therefore, sickle diet and companionship enhances endogenous spinal PEA which has an inhibitory effect on hyperalgesia in sickle mice. Treatment of control and sickle mice in R/C- group with PEA (i.p. 20 mg/kg/day) led to acute (1 hour) reduction of mechanical- (~40%, p&amp;lt;0.01) and cold-hyperalgesia (~40%, p&amp;lt;0.001) in sickle mice compared to pre-treatment, which was sustained during 3 day treatment, but had no effect on control mice which do not have hyperalgesia. PEA inhibits substance P (SP)-induced mast cell activity, and sickle mice show increased spinal SP, neuronal sensitization, peripheral nerve injury and mast cell activation (Tran et al., Blood 2017). Pain in SCD is both neuropathic and inflammatory. We examined if PEA inhibited the mechanisms that underlie spinal nerve repair by neurite outgrowth inhibitor, NOGO-A/reticulon-4, which regulates nerve regeneration via Rho Kinase (ROCK) signaling. NOGO-A contributes to inflammatory pain and hyperalgesia following spinal cord injury via NOGO receptor 1 in spinal cord. We observed that spinal NOGO-A expression and ROCK activity are upregulated (20% &amp; 100%, respectively) in sickle mice compared to control mice (all R/C-), which were inhibited upon 3-day treatment with PEA. We validated ROCK activity downstream of NOGO-A using SH-SY5Y neuroblastoma cells, simulating a sickle microenvironment with hemin (40 µM) and TNFα (1 ng/ml)(H+T). ROCK activity increased in H+T-treated SH-SY5Y cells compared to vehicle (~30%, p=0.05). In parallel, we analyzed the effect of PEA on extracellular traps (ET) in cutaneous mast cells from sickle mice induced by H+T in vitro. PEA treatment inhibited ET formation and extravasation of nuclear contents in H+T induced mast cells. Thus, PEA has the potential to attenuate neuropathic and inflammatory pain by inhibiting neuronal NOGO-A/ROCK pathway and mast cell activation in a sickle milieu. PEA has analgesic and anti-inflammatory effects on chronic pain in several clinical conditions. Therefore, our data suggest that diet and pleasure have the potential to upregulate pro-analgesic PEA that inhibits NOGO-A signaling and mast cell activation, leading to attenuation of hyperalgesia in sickle mice. Disclosures Gupta: Grifols: Research Funding; Cyclerion: Research Funding; 1910 Genetics: Research Funding; Novartis: Honoraria; Tautona Group: Honoraria; CSL Behring: Honoraria.

Nogo-A/S1PR2 Signaling Pathway Inactivation Decreases Microvascular Damage and Enhances Microvascular Regeneration in PDMCI Mice

The incidence of mild cognitive impairment in Parkinson’s disease (PDMCI) is as high as 18–55%. However, the pathological mechanism of PDMCI is not yet clear. Our previous research showed that microvascular pathology and chronic cerebral hypoperfusion participated in the occurrence and development of PDMCI. Nogo-A has been suggested to be a negative regulator of microvascular regeneration in the central nervous system. Moreover, few insights have illuminated the mechanisms of Nogo-A and microvascular pathology in PDMCI. Therefore, we hypothesized that Nogo-A might be involved in the negative regulation of PDMCI angiogenesis. In this study, C57BL/6J mice were injected with Nogo-A-specific short hairpin RNA (shRNA-Nogo-A) in the lateral ventricle and intraperitoneally injected with a combination of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and probenecid. Subjects were classified into the following five groups for the Morris water maze test: control (CON), CON + shRNA-GFP, CON + shRNA-Nogo-A, PDMCI, and PDMCI + shRNA-Nogo-A. Furthermore, blood–brain barrier (BBB) permeability, fluorescein isothiocyanate (FITC)-conjugated dextran, transmission electron microscopy (TEM), immunofluorescence and Western blot analyses were performed. The results showed that MPTP could cause spatial memory and behavioral impairment, significant microvascular impairment and increased Nogo-A expression. When Nogo-A expression was downregulated, the cognitive and microvascular impairments were alleviated, and the expression of sphingosine-1-phosphate receptor 2 (S1PR2) and the RhoA/ROCK signaling pathway were inhibited. These findings suggested that Nogo-A could bind to S1PR2, activate related signaling pathways, and lead to the inhibition of vascular remodeling in PDMCI mice. This study indicated that Nogo-A downregulation could mediate microvascular remodeling and provide further insights into the pathogenesis of PDMCI.

Nogo-A aggravates oxidative damage in oligodendrocytes.

Nogo-A is considered one of the most important inhibitors of myelin-associated axonal regeneration in the central nervous system. It is mainly expressed by oligodendrocytes. Although previous studies have found regulatory roles for Nogo-A in neurite outgrowth inhibition, neuronal homeostasis, precursor migration, plasticity, and neurodegeneration, its functions in the process of oxidative injury are largely uncharacterized. In this study, oligodendrocytes were extracted from the cerebral cortex of newborn Sprague-Dawley rats. We used hydrogen peroxide (H2O2) to induce an in vitro oligodendrocyte oxidative damage model and found that endogenously expressed Nogo-A is significantly upregulated in oligodendrocytes. After recombinant virus Ad-ZsGreen-rat Nogo-A infection of oligodendrocytes, Nogo-A expression was increased, and the infected oligodendrocytes were more susceptible to acute oxidative insults and exhibited a markedly elevated rate of cell death. Furthermore, knockdown of Nogo-A expression in oligodendrocytes by Ad-ZsGreen-shRNA-Nogo-A almost completely protected against oxidative stress induced by exogenous H2O2. Intervention with a Nogo-66 antibody, a LINGO1 blocker, or Y27632, an inhibitor in the Nogo-66-NgR/p75/LINGO-1-RhoA-ROCK pathway, did not affect the death of oligodendrocytes. Ad-ZsGreen-shRNA-Nogo-A also increased the levels of phosphorylated extracellular signal-regulated kinase 1/2 and inhibited BCL2 expression in oligodendrocytes. In conclusion, Nogo-A aggravated reactive oxygen species damage in oligodendrocytes, and phosphorylated extracellular signal-regulated kinase 1/2 and BCL2 might be involved in this process. This study was approved by the Ethics Committee of Peking University People's Hospital, China (approval No. 2018PHC081) on December 18, 2018.

Forced forelimb use following stroke enhances oligodendrogenesis and functional recovery in the rat

Forced limb use, which forces the use of the impaired arm following stroke, improves functional recovery. The study was designed to investigate the mechanisms of recovery underlying forced impaired limbuse. Furthermore, forced unimpaired arm use was also performed in order to explore its effect on functional behavior. We hypothesized that forced forelimb use could improve functional recovery in rats that have had an experimentally induced ischemic stroke, through promoting the recruitment and differentiation of the oligodendrocyte progenitor cells (OPCs). Indeed the proliferation of Olig2 and NG2 positive cells, as well as the expression of myelin basic protein (MBP)were increased in the perilesional striatum, whereas quantitative changes of Olig2+ and NG2+ oligodendrocyte progenitor cells was not observed in the subventricular zone. Through comparing rats forced to rely on affected or unaffected forelimb, the results demonstrated that forced impaired limb use boosted functional recovery. At the same time forced unimpaired limb use deteriorated limb movement of injured side. In addition, the expression of NogoA is reduced, when the injured limb was used more, suggesting that it played a role in the repair of white matter.

Promoting Axonal Extension and Repair After Spinal Cord Injury by Inhibiting the Expression of Rho Gene Based on RNA Interference

Spinal cord injury causes central nervous system damage. Rho inhibits axonal regeneration. This study is intended to analyze the effect of inhibition of Rho expression on axonal repair. The oligodendrocytes were isolated and divided into NC group and shRNA-RhoA group followed by analysis of the average length of axon growth and average microtubule fluorescence density by immunofluorescence, Nogo, MAG and RhoA expression were by Real time PCR. Wistar rats were separated into control group; SCI group and shRNA-RhoA group followed by analysis of the BBB scores and the Reuter score of sensory function, RhoA expression by Real time PCR and Western blot, Caspase3 activity as well as Nogo and MAG expression by Real time PCR. Compared with NC group, shRNA-RhoA group showed significantly increased average length of axon growth and average microtubule fluorescence density at the distal axon and reduced expression of RhoA, Nogo and MAG (P &lt; 0 05). In comparison to control group, SCI group presented significantly increased RhoA expression, decreased BBB score, increased Reuter score and Caspase3 activity as well as elevated Nogo and MAG expression (P &lt; 0 05). The shRNA-RhoA group significantly decreased RhoA expression, increased BBB score, decreased Reuter score and Caspase3 activity, and reduced Nogo and MAG expression compared with SCI group (P &lt; 0 05). Inhibiting RhoA expression can promote axon extension and regenerative repair. Targeting RhoA reduces axon growth inhibitory factor expression, inhibits apoptosis and effectively alleviates SCI.

Tanshinone IIA Promotes Axonal Regeneration in Rats with Focal Cerebral Ischemia Through the Inhibition of Nogo-A/NgR1/RhoA/ROCKII/MLC Signaling.

PurposeThe aim of this study was to evaluate the neuroprotective effect of tanshinone IIA (TSA) on focal cerebral ischemia in rats and to investigate whether it was associated with Nogo-A/NgR1/RhoA/Rho-associated protein kinase 2 (ROCKII)/myosin light chain (MLC) signaling.MethodsIn this study, focal cerebral ischemia animal model was used. Neurological deficit scores and infarction volume were investigated to evaluate the neuroprotection of TSA. Hematoxylin-eosin staining, Nissl staining, and immunofluorescence staining were conducted to detect ischemic changes in brain tissue and changes in neurofilament protein 200 (NF200) and growth-associated protein-43 (GAP-43) expression, respectively. Western blotting and qRT-PCR analyses were used to detect the expression levels of NF200, GAP-43 and Nogo-A/NgR1/RhoA/ROCKII/MLC pathway-related signaling molecules.ResultsTSA treatment can improve the survival rate of rats, reduce the neurological score and infarct volume, and reduce neuron damage. In addition, TSA also increased axon length and enhanced expression of NF200 and GAP-43. Importantly, TSA significantly attenuated the expression of Nogo-A, NgR1, RhoA, ROCKII, and p-MLC, and thus inhibiting the activation of this signaling pathway.ConclusionTSA promoted axonal regeneration by inhibiting the Nogo-A/NgR1/RhoA/ROCKII/MLC signaling pathway, thereby exerting neuroprotective effects in cerebral ischemia rats, which provided support for the clinical application of TSA in stroke treatment.

The Effect of Mild Hypothermia on Nogo-A and Neurological Function in the Brain after Cardiopulmonary Resuscitation in Rats

Objective We investigated the dynamic changes of Nogo-A protein in brain and the effects of mild therapeutic hypothermia (MTH) on its expression after cardiopulmonary resuscitation (CPR). Methods: Western-blotting and neurological scoring of 45 rats subjected to cardiac arrest and CPR with and without MTR were performed to investigate the changes in the expression of Nogo-A protein in the hippocampus and cortex over a period of time ranging from 6 h to 72 h after restoration of spontaneous circulation (ROSC). Results: Nogo-A expression levels were increased at 6 h after CPR in the hippocampus and cortex, peaked at 24 h in the cortex, and at 48 h in the hippocampus. The expression of Nogo-A in the MTR group was significantly lower at 12 h (p < 0.05) compared to those with no MTR after ROSC. Conclusions: MTR blunts the expression of Nogo-A protein in the hippocampus and cortex after cardiac arrest and resuscitation, and MTR may provide cerebral protection after ischemia.

Expression of Nogo-A in dorsal root ganglion in rats with cauda equina injury

ObjectiveTo investigate the expression of Nogo-A in dorsal root ganglion (DRG) in rats with cauda equina injury and the therapeutic effects of blocking Nogo-A and its receptor. Methods and materialsFifty-eight male Sprague-Dawley rats were divided randomly into either the sham operation group (n = 24) or the cauda equina compression (CEC) control group (n = 34). Behavioral, histological, and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) analyses were conducted to assess the establishment of the model. The dynamic expression change of Nogo-A was evaluated using real time-qPCR. Immunofluorescence was used to evaluate the expression of Nogo-A in the DRG and cauda equina. Furthermore, 20 male Sprague-Dawley rats were equally divided into 4 groups, including the sham group, the CEC group, the NEP1-40 (the NgR antagonist peptide) treatment group, and the JTE-013 (the S1PR2 antagonist) treatment group. Behavioral assessments and western blotting were used to evaluate the therapeutic effect of cauda equina injury via blocking Nogo-A and its receptor. ResultsTactile allodynia and heat hyperalgesia in the CEC model developed as soon as 1 day after surgery and recovered to normal at 7 days, which was followed by the downregulation of Nogo-A in DRG neurons. However, the locomotor function impairment in the CEC model showed a different prognosis from the sensory function, which was consistent with the expression change of Nogo-A in the spinal cord. Immunofluorescence results also demonstrated that Nogo A-positive/NF200-negative neurons and axons increased in the DRG and cauda equina 7 days after surgery. Surprisingly, Schwann cells, which myelinate axons in the PNS, also expressed considerable amounts of Nogo-A. Then, after blocking the Nogo-A/NgR signaling pathway by NEP1-40, significant improvement of mechanical allodynia was identified in the first 2 days after the surgery. Western blotting suggested the NEP1-40 treatment group had lower expression of cleaved caspase-3 than the CEC and JTE-013 treatment group. ConclusionNeuronal Nogo-A in the DRG may be involved in regeneration and play a protective role in the CEC model. Whereas Nogo-A, released from the injured axons or expressed by Schwann cells, may act as an inhibiting factor in the process of CEC repairment. Thus, blocking the Nogo-A/NgR signaling pathway can alleviate mechanical allodynia by apoptosis inhibition.

Expression of Olig2, Nestin, NogoA and AQP4 have no impact on overall survival in IDH-wildtype glioblastoma.

Despite many years of research efforts and clinical trials the prognosis of patients diagnosed with glioblastoma remains very poor. The oligodendrocyte transcription factor 2 (Olig2) was identified as a marker for glioma stem cells, which are believed to be responsible for glioma recurrence and therapy resistance. In this retrospective analysis we assessed the prognostic value of oligodendroglial and glioma stem cell markers in 113 IDH-wildtype glioblastomas. Immunohistochemical staining for Olig2, NogoA, AQP4 and Nestin was performed in combination with sequencing of IDH1 and IDH2 as well as promotor methylation analysis of the MGMT gene. Even though differences in overall survival according to Olig2 expression were observed, univariate and multivariate survival analysis did not reveal a firm significant prognostic impact of Olig2, NogoA, AQP4 or Nestin expression. Additionally, no differences in the expression of these markers depending on clinical status, age or gender were found. The established independent prognostic factors age<65, Karnofsky Performance Status> = 70 and methylated MGMT gene promoter were significant in the multivariate analysis. In conclusion expression of oligodendroglial and glioma stem cell markers do not have an independent prognostic effect in IDH-wildtype glioblastoma.

Neuroprotective Effects of Bone Marrow Mononuclear Cells Therapy in a Rat Model of Ischemia Stroke

Objective: Bone marrow mononuclear cells (BMMCs) are considered a potential approach to promote the recovery of stroke-induced neurological deficit. However, the exact mechanism of BMMCs in nerve function recovery is still unclear. Methods: Adult Sprague-Dawley (SD) rat models of cerebral ischemia-reperfusion injury was established by using thread method. BMMCs were transplanted into rat models. Neurological deficits were evaluated by Longa score scale. Immunohistochemistry assay were employed to examine the expression of GFAP and Nogo-A around the ischemic foci in the right frontal lobe. Caspase-3 activity was examined by Western Blot. Results: Rats in BMMCs group had lessened neurological deficits and cleaved Caspase-3 expression on day 21 after reper-fusion, as well as higher expression of GFAP [(37.62±2.45) vs. (27.62±1.69) and (38.00±1.85) vs. (27.25±1.83), P &lt; 0.05] and lower expression of Nogo-A [(28.88±2.64) vs. (32.50±1.60) and (23.87±2.36) vs. (32.00±1.85), P &lt; 0.05] on day 14 and 21 after reperfusion. Meanwhile, the expression of Nogo-A on day 21 was lower than that on day 14 after reperfusion [(23.87±2.36) vs. (28.88±2.64), P &lt; 0.05] in BMMCs group. Conclusion: These findings suggested that BMMCs treatment could improve the functional recovery of neurological deficits in rats with MCAO, which was probably related to enhanced expression of GFAP and reduced Nogo-A expression and Caspase-3 activity in the ischemic brain tissues.

Effects of bone marrow mesenchymal stem cells transplantation on the recovery of neurological functions and the expression of Nogo-A, NgR, Rhoa, and ROCK in rats with experimentally-induced convalescent cerebral ischemia.

BackgroundTo investigate the effects of intravenous transplantation of bone marrow mesenchymal stem cells (BMSCs) on neurological function in rats with experimentally-induced convalescent cerebral ischemia and the expression of Nogo-A, NgR, Rhoa, and ROCK expression.MethodsBMSCs were isolated and cultured in vitro using the whole bone marrow adherent method. Eighty-one adult male Sprague-Dawley rats were divided at random into three groups: the sham-operated group, the cerebral ischemia group, and the BMSC treatment group (n=27 rats per group). In the latter two groups, the middle cerebral artery occlusion (MCAO) model was performed by the modified Zea Longa method. After MCAO, rats in the sham-operated and cerebral ischemic groups were injected with 1 mL of phosphate buffered saline (PBS) via the tail vein. In the BMSC-treatment group, 1 mL of the BMSC suspension (containing 3×106 BMSCs) was injected through the rats’ femoral vein. At 12, 24, and 72 h after BMSC transplantation, modified neurological deficit scores (mNSS) were used to assess neurological function. TTC (2,3,5-triphenyl tetrazolium chloride) staining was used to measure the ischemic lesion volume, and the distribution of Nogo-A protein was observed by immunohistochemistry. The expressions of Nogo-A, NgR, Rhoa, and ROCK were detected by Western blot.ResultsAt 72 h after BMSC transplantation, the mNSS scores were significantly lower in the BMSC treatment group than those in the cerebral ischemia group (7.50±0.55 vs. 8.67±0.52, P<0.01), and the ischemic lesions volume was significantly reduced. The expressions of Nogo-A, NgR, RhoA, and ROCK were significantly decreased compared with the controls (P<0.05).ConclusionsThe transplantation of BMSCs can improve neurological function in rats after convalescent cerebral ischemia, and their therapeutic effect may be related to the downregulation of Nogo-A, NgR, RhoA, and ROCK expression.

Blockade of Nogo-A/Nogo-66 receptor 1 (NgR1) Inhibits Autophagic Activation and Prevents Secondary Neuronal Damage in the Thalamus after Focal Cerebral Infarction in Hypertensive Rats

Focal cerebral infarction leads to autophagic activation, which contributes to secondary neuronal damage in the ipsilateral thalamus. Although Nogo-A deactivation enhances neuronal plasticity, its role in autophagic activation in the thalamus after ischemic stroke remains unclear. This study aimed to investigate the potential roles of Nogo-A/Nogo-66 receptor 1 (NgR1) in autophagic activation in the ipsilateral thalamus after cerebral infarction. Focal neocortical infarction was established using the middle cerebral artery occlusion (MCAO) method. Secondary damage in the ipsilateral thalamus was assessed by Nissl staining and immunostaining. The expression of Nogo-A, NgR1, Rho-A and Rho-associated coiled-coil containing protein kinase 1 (ROCK1) as well as autophagic flux were evaluated by immunofluorescence and immunoblotting. The roles of Nogo-A–NgR1 signaling in autophagic activation were determined by intraventricular delivery of an NgR1 antagonist peptide, NEP1–40, at 24 h after MCAO. The results showed that Nogo-A and NgR1 overexpression temporally coincided with marked increases in the levels of Beclin1, LC3-II and sequestosome 1 (SQSTM1)/p62 in the ipsilateral thalamus at seven and fourteen days after MCAO. In contrast, NEP1–40 treatment significantly reduced the expression of Rho-A and ROCK1 which was accompanied by marked reductions of LC3-II conversion as well as the levels of Beclin1 and SQSTM1/p62. Furthermore, NEP1–40 treatment significantly reduced neuronal loss and gliosis in the ipsilateral thalamus, and accelerated somatosensory recovery at the observed time-points after MCAO. These results suggest that blockade of Nogo-A–NgR1 signaling inhibits autophagic activation, attenuates secondary neuronal damage in the ipsilateral thalamus, and promotes functional recovery after focal cerebral cortical infarction.