Tetramethylpyrazine Nitrone Promotes the Clearance of Alpha-Synuclein via Nrf2-Mediated Ubiquitin-Proteasome System Activation.

Aggregation of α-synuclein (α-syn) and α-syn cytotoxicity are hallmarks of sporadic and familial Parkinson's disease (PD). Nuclear factor (erythroid-derived 2)-like 2 (Nrf2)-dependent enhancement of the expression of the 20S proteasome core particles (20S CPs) and regulatory particles (RPs) increases proteasome activity, which can promote α-syn clearance in PD. Activation of peroxisome proliferator-activated receptor γ co-activator 1α (PGC-1α) may reduce oxidative stress by strongly inducing Nrf2 gene expression. In the present study, tetramethylpyrazine nitrone (TBN), a potent-free radical scavenger, promoted α-syn clearance by the ubiquitin-proteasome system (UPS) in cell models overexpressing the human A53T mutant α-syn. In the α-syn transgenic mice model, TBN improved motor impairment, decreased the products of oxidative damage, and down-regulated the α-syn level in the serum. TBN consistently up-regulated PGC-1α and Nrf2 expression in tested models of PD. Additionally, TBN similarly enhanced the proteasome 20S subunit beta 8 (Psmb8) expression, which is linked to chymotrypsin-like proteasome activity. Furthermore, TBN increased the mRNA levels of both the 11S RPs subunits Pa28αß and a proteasome chaperone, known as the proteasome maturation protein (Pomp). Interestingly, specific siRNA targeting of Nrf2 blocked TBN's effects on Psmb8, Pa28αß, Pomp expression, and α-syn clearance. In conclusion, TBN promotes the clearance of α-syn via Nrf2-mediated UPS activation, and it may serve as a potentially disease-modifying therapeutic agent for PD.

Tetramethylpyrazine Nitrone alleviates D-galactose-induced murine skeletal muscle aging and motor deficits by activating the AMPK signaling pathway.

Tetramethylpyrazine nitrone (TBN), a novel derivative of tetramethylpyrazine (TMP) designed and synthesized by our group, possesses multi-functional mechanisms of action and displays broad protective effects in vitro and in animal models of age-related brain disorders such as stroke, Alzheimer's disease (AD), Amyotrophic Lateral Sclerosis (ALS) and Parkinson's disease (PD). In the present report, we investigated the effects of TBN on aging, specifically on muscle aging and the associated decline of motor functions. Using a D-galactose-induced aging mouse model, we found that TBN could reverse the levels of several senescence and aging markers including p16, p21, ceramides, and telomere length and increase the wet-weight ratio of gastrocnemius muscle tissue, demonstrating its efficacy in ameliorating muscle aging. Additionally, the pharmacological effects of TBN on motor deficits (gait analysis, pole-climbing test and grip strength test), muscle fibrosis (hematoxylin & eosin (HE), Masson staining, and αSMA staining), inflammatory response (IL-1ß, IL-6, and TNF-α), and mitochondrial function (ATP, mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) were also confirmed in the D-galactose-induced aging models. Further experiments demonstrated that TBN alleviated muscle aging and improved the decline of age-related motor deficits through an AMPK-dependent mechanism. These findings highlight the significance of TBN as a potential anti-aging agent to combat the occurrence and development of aging and age-related diseases.

Tetramethylpyrazine nitrone exerts neuroprotection via activation of PGC-1α/Nrf2 pathway in Parkinson's disease models.

INTRODUCTION: Parkinson's disease (PD) is common neurodegenerative disease where oxidative stress and mitochondrial dysfunction play important roles in its progression. Tetramethylpyrazine nitrone (TBN), a potent free radical scavenger, has shown protective effects in various neurological conditions. However, the neuroprotective mechanisms of TBN in PD models remain unclear. OBJECTIVES: We aimed to investigate TBN's neuroprotective effects and mechanisms in PD models. METHODS: TBN's neuroprotection was initially measured in MPP+/MPTP-induced PD models. Subsequently, a luciferase reporter assay was used to detect peroxisome proliferator-activated receptor γ co-activator 1α (PGC-1α) promoter activity. Effects of TBN on antioxidant damage and the PGC-1α/Nuclear factor erythroid-2-related factor 2 (Nrf2) pathway were thoroughly investigated. RESULTS: In MPP+-induced cell model, TBN (30-300 µM) increased cell survival by 9.95 % (P < 0.05), 16.63 % (P < 0.001), and 24.09 % (P < 0.001), respectively. TBN enhanced oxidative phosphorylation (P < 0.05) and restored PGC-1α transcriptional activity suppressed by MPP+ (84.30 % vs 59.03 %, P < 0.01). In MPTP-treated mice, TBN (30 mg/kg) ameliorated motor impairment, increased striatal dopamine levels (16.75 %, P < 0.001), dopaminergic neurons survival (27.12 %, P < 0.001), and tyrosine hydroxylase expression (28.07 %, P < 0.01). Selegiline, a positive control, increased dopamine levels (15.35 %, P < 0.001) and dopaminergic neurons survival (25.34 %, P < 0.001). Additionally, TBN reduced oxidative products and activated the PGC-1α/Nrf2 pathway. PGC-1α knockdown diminished TBN's neuroprotective effects, decreasing cell viability from 73.65 % to 56.87 % (P < 0.001). CONCLUSION: TBN has demonstrated consistent effectiveness in MPP+-induced midbrain neurons and MPTP-induced mice. Notably, the therapeutic effect of TBN in mitigating motor deficits and neurodegeneration is superior to selegiline. The neuroprotective mechanisms of TBN are associated with activation of the PGC-1α/Nrf2 pathway, thereby reducing oxidative stress and maintaining mitochondrial function. These findings suggest that TBN may be a promising therapeutic candidate for PD, warranting further development and investigation.

Celastrol attenuates hepatitis C virus translation and inflammatory response in mice by suppressing heat shock protein 90ß.

Hepatitis C virus (HCV) infection is one of the major factors to trigger a sustained hepatic inflammatory response and hence hepatocellular carcinoma (HCC), but direct-acting-antiviral (DAAs) was not efficient to suppress HCC development. Heat shock protein 90 kDa (HSP90) is highly abundant in different types of cancers, and especially controls protein translation, endoplasmic reticulum stress, and viral replication. In this study we investigated the correlation between the expression levels of HSP90 isoforms and inflammatory response marker NLRP3 in different types of HCC patients as well as the effect of a natural product celastrol in suppression of HCV translation and associated inflammatory response in vivo. We identified that the expression level of HSP90ß isoform was correlated with that of NLRP3 in the liver tissues of HCV positive HCC patients (R2 = 0.3867, P < 0.0101), but not in hepatitis B virus-associated HCC or cirrhosis patients. We demonstrated that celastrol (3, 10, 30 µM) dose-dependently suppressed the ATPase activity of both HSP90α and HSP90ß, while its anti-HCV activity was dependent on the Ala47 residue in the ATPase pocket of HSP90ß. Celastrol (200 nM) halted HCV internal ribosomal entry site (IRES)-mediated translation at the initial step by disrupting the association between HSP90ß and 4EBP1. The inhibitory activity of celastrol on HCV RNA-dependent RNA polymerase (RdRp)-triggered inflammatory response also depended on the Ala47 residue of HSP90ß. Intravenous injection of adenovirus expressing HCV NS5B (pAde-NS5B) in mice induced severe hepatic inflammatory response characterized by significantly increased infiltration of immune cells and hepatic expression level of Nlrp3, which was dose-dependently ameliorated by pretreatment with celastrol (0.2, 0.5 mg/kg, i.p.). This study reveals a fundamental role of HSP90ß in governing HCV IRES-mediated translation as well as hepatic inflammation, and celastrol as a novel inhibitor of HCV translation and associated inflammation by specifically targeting HSP90ß, which could be developed as a lead for the treatment of HSP90ß positive HCV-associated HCC.

Tetramethylpyrazine nitrone activates hypoxia-inducible factor and regulates iron homeostasis to improve renal anemia.

Renal anemia is one of the most common complications of chronic kidney disease and diabetic kidney disease. Despite the progress made in recent years, there is still an urgent unmet clinical need for renal anemia treatment. In this research, we investigated the efficacy and mechanism of action of the novel tetramethylpyrazine nitrone (TBN). Animal models of anemia including the streptozotocin (STZ)-induced spontaneously hypertensive rats (SHR) and the cisplatin (CDDP)-induced C57BL/6J mice are established to study the TBN's effects on expression of hypoxia-inducible factor and erythropoietin. To explore the mechanism of TBN's therapeutic effect on renal anemia, cobalt chloride (CoCl2) is used in Hep3B/HepG2 cells to simulate a hypoxic environment. TBN is found to increase the expression of hypoxia-inducible factor HIF-1α and HIF-2α under hypoxic conditions and reverse the reduction of HIFs expression caused by saccharate ferric oxide (SFO). TBN also positively regulates the AMPK pathway. TBN stimulates nuclear transcription and translation of erythropoietin by enhancing the stability of HIF-1α expression. TBN has a significant regulatory effect on several major biomarkers of iron homeostasis, including ferritin, ferroportin (FPN), and divalent metal transporter-1 (DMT1). In conclusion, TBN regulates the AMPK/mTOR/4E-BP1/HIFs pathway, and activates the hypoxia-inducible factor and regulates iron homeostasis to improve renal anemia.

Memantine nitrate MN-08 suppresses NLRP3 inflammasome activation to protect against sepsis-induced acute lung injury in mice.

Sepsis is a life-threatening organ dysfunction with devastating consequences, prominent among which is lung damage. Memantine, an N-methyl-D-aspartic acid receptor (NMDAR) antagonist, is able to alleviate acute lung injury (ALI). Nitric oxide (NO) suppresses NLRP3 inflammasome activation against lipopolysaccharide (LPS)-induced septic shock. MN-08, a novel nitrate derivative of memantine, possesses both the ability to antagonize NMDAR and release NO. In the present study, we aimed to investigate the protective effects of MN-08 against LPS-induced systemic inflammation and septic lung injury in mice, and to explore the underlying mechanisms of MN-08 in LPS-induced mice and THP-1 macrophages. MN-08 significantly increased the survival rate of septic mice, alleviated LPS-induced sepsis in mice via improving systemic inflammatory response syndrome and immune dysfunction, and attenuated pulmonary injury and inflammatory infiltration. More importantly, the therapeutic benefit of MN-08 for sepsis was greater than that of memantine and dexamethasone. Mechanistically, MN-08 exerted anti-inflammatory activity through inhibiting nuclear translocation of NF-κB, activation of the MAPK signaling pathway and the signaling transduction of STAT3/NF-κB. In addition, MN-08 suppressed NLRP3 inflammasome activation. Taken together, our studies demonstrate that MN-08 may be a promising therapeutic agent for sepsis-induced acute lung injury.

Neuroprotection against 1-Methyl-4-phenylpyridinium-induced cytotoxicity by naturally occurring polydatin through activation of transcription factor MEF2D.

Polydatin is the major active ingredient of Polygonum cuspidatum Sieb. Et Zucc. A recent study indicated that polydatin could protect against substantia nigra dopaminergic degeneration in rodent models associated with Parkinson's disease. However, mechanisms that underlie the neuroprotection of polydatin have not been fully elucidated. In the current study, the neuroprotective effects and detailed mechanisms of action of polydatin were investigated in Parkinson's disease-related cellular models. Polydatin dose- and time-dependently prevented neurotoxicity caused by 1-methyl-4-phenylpyridinium ion (MPP+) in primary cerebellar granule neurons. Moreover, we found that polydatin enhanced the activity of the transcription factor myocyte enhancer factor 2D (MEF2D) at both basal and pathological conditions using luciferase reporter gene assay. Additionally, western blot analysis revealed that polydatin could downregulate glycogen synthase kinase 3ß (GSK3ß), which is a negative regulator of MEF2D. Molecular docking simulations finally suggested an interaction between polydatin and a hydrophobic pocket within GSK3ß. All these results suggest that polydatin prevents MPP+-induced neurotoxicity via enhancing MEF2D through the inhibition of GSK3ß and that treatment with polydatin is worthy of further anti-Parkinson's disease study in future.

Nephroprotective Effects of Tetramethylpyrazine Nitrone TBN in Diabetic Kidney Disease.

Diabetic kidney disease (DKD) is the leading cause of end-stage renal failure, but therapeutic options for nephroprotection are limited. Oxidative stress plays a key role in the pathogenesis of DKD. Our previous studies demonstrated that tetramethylpyrazine nitrone (TBN), a novel nitrone derivative of tetramethylpyrazine with potent free radical-scavenging activity, exerted multifunctional neuroprotection in neurological diseases. However, the effect of TBN on DKD and its underlying mechanisms of action are not yet clear. Herein, we performed streptozotocin-induced rat models of DKD and found that TBN administrated orally twice daily for 6 weeks significantly lowered urinary albumin, N-acetyl-ß-D-glycosaminidase, cystatin C, malonaldehyde, and 8-hydroxy-2'-deoxyguanosine levels. TBN also ameliorated renal histopathological changes. More importantly, in a nonhuman primate model of spontaneous stage III DKD, TBN increased the estimated glomerular filtration rate, decreased serum 3-nitrotyrosine, malonaldehyde and 8-hydroxy-2'-deoxyguanosine levels, and improved metabolic abnormalities. In HK-2 cells, TBN increased glycolytic and mitochondrial functions. The protective mechanism of TBN might involve the activation of AMPK/PGC-1α-mediated downstream signaling pathways, thereby improving mitochondrial function and reducing oxidative stress in the kidneys of DKD rodent models. These results support the clinical development of TBN for the treatment of DKD.

Therapeutic efficacy of novel memantine nitrate MN-08 in animal models of Alzheimer's disease.

Alzheimer's disease (AD) is a leading cause of dementia in elderly individuals and therapeutic options for AD are very limited. Over-activation of N-methyl-D-aspartate (NMDA) receptors, amyloid ß (Aß) aggregation, a decrease in cerebral blood flow (CBF), and downstream pathological events play important roles in the disease progression of AD. In the present study, MN-08, a novel memantine nitrate, was found to inhibit Aß accumulation, prevent neuronal and dendritic spine loss, and consequently attenuate cognitive deficits in 2-month-old APP/PS1 transgenic mice (for a 6-month preventative course) and in the 8-month-old triple-transgenic (3×Tg-AD) mice (for a 4-month therapeutic course). In vitro, MN-08 could bind to and antagonize NMDA receptors, inhibit the calcium influx, and reverse the dysregulations of ERK and PI3K/Akt/GSK3ß pathway, subsequently preventing glutamate-induced neuronal loss. In addition, MN-08 had favorable pharmacokinetics, blood-brain barrier penetration, and safety profiles in rats and beagle dogs. These findings suggest that the novel memantine nitrate MN-08 may be a useful therapeutic agent for AD.

Dual-Functional MN-08 Attenuated Pulmonary Arterial Hypertension Through Vasodilation and Inhibition of Pulmonary Arterial Remodeling.

[Figure: see text].

The Tetramethylpyrazine Analogue T-006 Alleviates Cognitive Deficits by Inhibition of Tau Expression and Phosphorylation in Transgenic Mice Modeling Alzheimer's Disease.

T-006, a small-molecule compound derived from tetramethylpyrazine (TMP), has potential for the treatment of neurological diseases. In order to investigate the effect of T-006 prophylactic treatment on an Alzheimer's disease (AD) model and identify the target of T-006, we intragastrically administered T-006 (3 mg/kg) to Alzheimer's disease (AD) transgenic mice (APP/PS1-2xTg and APP/PS1/Tau-3xTg) for 6 and 8 months, respectively. T-006 improved cognitive ability after long-term administration in two AD mouse models and targeted mitochondrial-related protein alpha-F1-ATP synthase (ATP5A). T-006 significantly reduced the expression of phosphorylated-tau, total tau, and APP while increasing the expression of synapse-associated proteins in 3xTg mice. In addition, T-006 modulated the JNK and mTOR-ULK1 pathways to reduce both p-tau and total tau levels. Our data suggested that T-006 mitigated cognitive decline primarily by reducing the p-tau and total tau levels in 3xTg mice, supporting further investigation into its development as a candidate drug for AD treatment.

Tetramethylpyrazine nitrone improves motor dysfunction and pathological manifestations by activating the PGC-1α/Nrf2/HO-1 pathway in ALS mice.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by progressive loss of upper and lower motor neurons that results in skeletal muscle atrophy, weakness and paralysis. Oxidative stress plays a key role in the pathogenesis of ALS, including familial forms of the disease arising from mutation of the gene coding for superoxide dismutase (SOD1). We have used the SOD1G93A ALS mouse model to investigate the efficacy of 2-[[(1,1-dimethylethyl)oxidoimino]-methyl]-3,5,6-trimethylpyrazine (TBN), a novel tetramethylpyrazine derivative armed with a powerful free-radical scavenging nitrone moiety. TBN was administered to mice by intraperitoneal or intragastric injection after the onset of motor deficits. TBN slowed the progression of motor neuron disease as evidenced by improved motor performance, reduced spinal motor neuron loss and the associated glial response, and decreased skeletal muscle fiber denervation and fibrosis. TBN treatment activated mitochondrial antioxidant activity through the PGC-1α/Nrf2/HO-1 pathway and decreased the expression of human SOD1. These findings suggest that TBN holds promise as a therapeutic agent for ALS.

Neuroprotective and neurogenic effects of novel tetramethylpyrazine derivative T-006 in Parkinson's disease models through activating the MEF2-PGC1α and BDNF/CREB pathways.

T-006, a new derivative of tetramethylpyrazine, has been recently found to protect against 6-hydroxydopamine (6-OHDA)-induced neuronal damage and clear α-synuclein (α-syn) by enhancing proteasome activity in an α-syn transgenic Parkinson's disease (PD) model. The effect of T-006 on the 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-induced PD model, however, has not been tested and T-006's neuroprotective mechanisms have not been fully elucidated. In this study, we further investigated the neuroprotective and neurogenic effects of T-006 and explored its underlying mechanism of action in both cellular and animal PD models. T-006 was able to improve locomotor behavior, increase survival of nigra dopaminergic neurons and boost striatal dopamine levels in both MPTP- and 6-OHDA-induced animals. T-006 treatment restored the altered expressions of myocyte enhancer factor 2D (MEF2D), peroxisome proliferator-activated receptor γ (PPARγ) co-activator 1α (PGC1α) and NF-E2-related factor 1/2 (Nrf1/2) via modulation of Akt/GSK3ß signaling. T-006 stimulated MEF2, PGC1α and Nrf2 transcriptional activities, inducing Nrf2 nuclear localization. Interestingly, T-006 promoted endogenous adult neurogenesis toward a dopaminergic phenotype by activating brain-derived neurotrophic factor (BDNF) and cAMP responsive element-binding protein (CREB) in 6-OHDA rats. Our work demonstrated that T-006 is a potent neuroprotective and neuroregenerative agent that may have therapeutic potential in the treatment of PD.

Multifunctional memantine nitrate significantly protects against glutamate-induced excitotoxicity via inhibiting calcium influx and attenuating PI3K/Akt/GSK3beta pathway.

Overactivation of N-methyl-D-aspartate (NMDA) receptors has been associated with neurodegenerative disorders such as Alzheimer's disease (AD), cerebral vascular disorders and amyotrophic lateral sclerosis (ALS). We have previously designed and synthesized a series of memantine nitrate and some of them have shown vessel dilatory effects and neuroprotective effects; however, the detailed mechanisms have not been elucidated. In this study, we further demonstrated that memantine nitrate-06 (MN-06), one of the novel compounds derived from memantine, possessed significant neuroprotective effects against glutamate-induced excitotoxicity in rat primary cerebellar granule neurons (CGNs). Pretreatment of MN-06 reversed the activation of GSK3b and the suppression of phosphorylated Akt induced by glutamate. In addition, the neuroprotective effects of MN-06 could be abolished by LY294002, the specific phosphatidylinositol 3-kinase (PI3-K) inhibitor. Ca2+ imaging shown that pretreatment of MN-06 prevented Ca2+ influx induced by glutamate. Moreover, MN-06 might inhibit the NMDA-mediated current by antagonizing NDMA receptors, which was further confirmed by molecular docking simulation. Taken together, MN-06 protected against glutamate-induced excitotoxicity by blocking calcium influx and attenuating PI3-K/Akt/GSK-3b pathway, indicating that MN-06 might be a potential drug for treating neurodegenerative disorders.

Pharmacological Characterizations of anti-Dementia Memantine Nitrate via Neuroprotection and Vasodilation in Vitro and in Vivo.

We have previously designed and synthesized a series of novel memantine nitrates, and some of them have shown neuroprotective effects; however, the detailed mechanisms remain unknown. In this study, we demonstrated that MN-12, one of the memantine nitrates, concentration-dependently protected against glutamate-induced neurotoxicity in rat primary cultured cerebellar granule neurons (CGNs). Western blotting assays revealed that MN-12 might possess neuroprotective effects through the inhibition of ERK pathway and activation of PI3K/Akt pathway concurrently. Moreover, MN-12 concentration-dependently dilated precontracted rat middle cerebral artery through activation of NO-cGMP pathway ex vivo. In the 2-vessel occlusion (2VO) rat model, MN-12 alleviated the impairments of spatial memory and motor dysfunction possibly via neuroprotection and improvement of the cerebral blood flow. Furthermore, the results of preliminary pharmacokinetic studies showed that MN-12 might quickly distribute to the major organs including the brain, indicating that MN-12 could penetrate the blood-brain barrier. Taken together, MN-12 might provide multifunctional therapeutic benefits for dementia associated with Alzheimer's disease, vascular dementia, and ischemic stroke, via neuroprotection and vessel dilation to improve the cerebral blood flow.

Sirt3-dependent deacetylation of COX-1 counteracts oxidative stress-induced cell apoptosis.

The mitochondrial complexes are prone to sirtuin (Sirt)3-mediated deacetylation modification, which may determine cellular response to stimuli, such as oxidative stress. In this study, we show that the cytochrome c oxidase (COX)-1, a core catalytic subunit of mitochondrial complex IV, was acetylated and deactivated both in 2,2'-azobis(2-amidinopropane) dihydrochloride-treated NIH/3T3 cells and hydrogen peroxide-treated primary neuronal cells, correlating with apoptotic cell death induction by oxidative stress. Inhibition of Sirt3 by small interfering RNA or the inhibitor nicotinamide induced accumulation of acetylation of COX-1, reduced mitochondrial membrane potential, and increased cell apoptosis. In contrast, overexpression of Sirt3 enhanced deacetylation of COX-1 and inhibited oxidative stress-induced apoptotic cell death. Significantly, rats treated with ischemia/reperfusion injury, a typical oxidative stress-related disease, presented an inhibition of Sirt3-induced hyperacetylation of COX-1 in the brain tissues. Furthermore, K13, K264, K319, and K481 were identified as the acetylation sits of COX-1 in response to oxidative stress. In conclusion, COX-1 was discovered as a new deacetylation target of Sirt3, indicating that the Sirt3/COX-1 axis is a promising therapy target of stress-related diseases.-Tu, L.-F., Cao, L.-F., Zhang, Y.-H., Guo, Y.-L., Zhou, Y.-F., Lu, W.-Q., Zhang, T.-Z., Zhang, T., Zhang, G.-X., Kurihara, H., Li, Y.-F., He, R.-R. Sirt3-dependent deacetylation of COX-1 counteracts oxidative stress-induced cell apoptosis.

The dual-functional memantine nitrate MN-08 alleviates cerebral vasospasm and brain injury in experimental subarachnoid haemorrhage models.

BACKGROUND AND PURPOSE: Cerebral vasospasm and neuronal apoptosis after subarachnoid haemorrhage (SAH) is the major cause of morbidity and mortality in SAH patients. So far, single-target agents have not prevented its occurrence. Memantine, a non-competitive NMDA re3ceptor antagonist, is known to alleviate brain injury and vasospasm in experimental models of SAH. Impairment of NO availability also contributes to vasospasm. Recently, we designed and synthesized a memantine nitrate MN-08, which has potent dual functions: neuroprotection and vasodilation. Here, we have tested the therapeutic effects of MN-08 in animal models of SAH. EXPERIMENTAL APPROACH: Binding to NMDA receptors (expressed in HEK293 cells), NO release and vasodilator effects of MN-08 were assessed in vitro. Therapeutic effects of MN-08 were investigated in vivo, using rat and rabbit SAH models. KEY RESULTS: MN-08 bound to the NMDA receptor, slowly releasing NO in vitro and in vivo. Consequently, MN-08 relaxed the pre-contracted middle cerebral artery ex vivo and increased blood flow velocity in small vessels of the mouse cerebral cortex. It did not, however, lower systemic blood pressure. In an endovascular perforation rat model of SAH, MN-08 improved the neurological scores and ameliorated cerebral vasospasm. Moreover, MN-08 also alleviated cerebral vasospasm in a cisterna magna single-injection model in rabbits. MN-08 attenuated neural cell apoptosis in both rat and rabbit models of SAH. Importantly, the therapeutic benefit of MN-08 was greater than that of memantine. CONCLUSION AND IMPLICATIONS: MN-08 has neuroprotective potential and can ameliorate vasospasm in experimental SAH models.

Tetramethylpyrazine Nitrone Reduces Oxidative Stress to Alleviate Cerebral Vasospasm in Experimental Subarachnoid Hemorrhage Models.

Cerebral vasospasm is one of the deleterious complications after subarachnoid hemorrhage (SAH), leading to delayed cerebral ischemia and permanent neurological deficits or even death. Free radicals and oxidative stress are considered as crucial causes contributing to cerebral vasospasm and brain damage after SAH. Tetramethylpyrazine nitrone (TBN), a derivative of the clinically used anti-stroke drug tetramethylpyrazine armed with a powerful free radical scavenging nitrone moiety, has been reported to prevent brain damage from ischemic stroke. The present study aimed to investigate the effects of TBN on vasospasm and brain damage after SAH. Two experimental SAH models were used, a rat model by endovascular perforation and a rabbit model by intracisternal injection of autologous blood. The effects of TBN on SAH were evaluated assessing basilar artery spasm, neuronal apoptosis, and neurological deficits. TBN treatment significantly attenuated vasospasm, improved neurological behavior functions and reduced the number of apoptotic neurons in both the SAH rats and rabbits. Mechanistically, TBN suppressed the increase in 3-nitrotyrosine and 8-hydroxy-2-deoxyguanosine immuno-positive cells in the cortex of SAH rat brain. Western blot analyses indicated that TBN effectively reversed the altered expression of Bcl-2, Bax and cytochrome C, and up-regulated nuclear factor erythroid-derived 2-like 2 (Nrf2) and hemeoxygenase-1 (HO-1) protein expressions. In the in vitro studies, TBN inhibited H2O2-induced bEnd.3 cell apoptosis and reduced ROS generation. Additionally, TBN alleviated the contraction of rat basilar artery rings induced by H2O2 ex vivo. In conclusion, TBN ameliorated SAH-induced cerebral vasospasm and neuronal damage. These effects of TBN may be attributed to its anti-oxidative stress effect and up-regulation of Nrf2/HO-1.

Novel neuroprotective tetramethylpyrazine analog T-006 promotes neurogenesis and neurological restoration in a rat model of stroke.

Neuronal death is among the deleterious pathological changes that occur after cerebral ischemia and can lead to transient or permanent neurological deficits. The tetramethylpyrazine analog T-006 has been shown to be a multifunctional neuroprotective agent; however, its neuroprotective effect and mechanism of action have not been studied in ischemic stroke model rats. This study investigated the neuroprotective effects of T-006 in rat stroke model using a battery of behavioral and molecular biological tests. Results indicated that T-006 treatment significantly improved neurological function and behavior. Double immunofluorescence staining showed that T-006 visibly improved the number of NeuN/BrdU, Nestin/BrdU, and DCX/BrdU cells and induced neuronal regeneration. Western blot analyses indicated that T-006 upregulated neurogenesis-related protein expression of postsynaptic density protein 95, brain-derived neurotrophic factor, synaptophysin, and myelin basic protein. Collectively, these data suggest that T-006 stimulated neurogenesis in rats with middle cerebral artery occlusion and restored neurological functions.