Trans-anethole protects cortical neuronal cells against oxygen–glucose deprivation/reoxygenation

Trans-anethole has been studied on pharmacological properties such as anti-inflammation, anti-oxidative stress, antifungal and anticancer. However, to date, the anti-ischemic effects of trans-anethole have not been assessed. Therefore, we investigated the neuroprotection of trans-anethole against oxygen–glucose deprivation/reoxygenation (OGD/R)-induced cortical neuronal cell injury, an in vitro model of ischemia. The abilities of trans-anethole to block excitotoxicity, oxidative stress and mitochondrial dysfunction were evaluated in OGD/R-induced neurons. Trans-anethole significantly ameliorated OGD/R-induced neuronal cell injury by attenuating the intracellular calcium overload via the activation of NMDA receptors. Trans-anethole also inhibited OGD/R-induced reactive oxygen species overproduction, which may be derived from the scavenging activity in peroxyl radicals, assessed in an oxygen radical absorbance capacity assay. Furthermore, trans-anethole was shown to attenuate the depolarization of mitochondrial transmembrane. These results indicated that the neuroprotective effect of trans-anethole on OGD/R-induced neuronal injury might be due to its ability to inhibit excitotoxicity, oxidative stress and mitochondrial dysfunction. Considering these multiple pathways causing ischemic neuronal damage, the multi-functional effect of trans-anethole suggested that it may be effective in treating ischemic stroke.     

The involvement of TRPV4 on the hypoxia-induced oxidative neurotoxicity and apoptosis in a neuronal cell line: Protective role of melatonin

The hypoxia (HYPX)-mediated excessive generation of mitochondrial free reactive oxygen species (mROS) and the overload Ca²⁺ influx via the inhibition of TRPV4 are controlled by the treatment of antioxidants. However, the molecular mechanisms underlying melatonin (MLT)'s neuroprotection remains elusive. We investigated the role of MLT via modulation of TRPV4 on oxidative neurodegeneration and death in SH-SY5Y neuronal cells. The SH-SY5Y cells were divided into five groups as follows: control, MLT (1 mM for 2 h), HYPX (200 μM CoCl₂ for 24 h), HYPX + MLT, and HYPX + TRPV4 blockers (ruthenium red-1 μM for 30 min). The HYPX caused to the increase of TRPV4 current density and overload Ca²⁺ influx with an increase of mitochondrial membrane potential and mROS generation. The changes were not observed in the absence of TRPV4. When HYPX exposure and TRPV4 agonist (GSK1016790A)-induced TRPV4 activity were inhibited by the treatment of ruthenium red or MLT, the increase of mROS, lipid peroxidation, apoptosis, Zn²⁺ concentrations, TRPV4, caspase -3, caspase -9, Bax, and Bcl-2 expressions were restored via upregulation of reduced glutathione, glutathione peroxidase, and total antioxidant status. The levels of apoptosis and cell death in the cells were enriched with increases of caspase -3 and -9 activations, although they were decreased by MLT treatment. In conclusion, the treatment of MLT modulates HYPX-mediated mROS, apoptosis, and TRPV4-mediated overload Ca²⁺ influx and may provide an avenue for protecting HYPX-mediated neurological diseases associated with the increase of mROS, Ca²⁺, and Zn²⁺ concentration.     

Regulation of adenosine triphosphate-sensitive potassium channels suppresses the toxic effects of amyloid-beta peptide(25-35)

In this study, we treated PC12 cells with 0-20 μM amyloid-β peptide (25-35) for 24 hours to induce cytotoxicity, and found that 5-20 μM amyloid-β peptide (25-35) decreased PC12 cell viability, but adenosine triphosphate-sensitive potassium channel activator diazoxide suppressed the decrease in PC12 cell viability induced by amyloid-β peptide (25-35). Diazoxide protected PC12 cells against amyloid-β peptide (25-35)-induced increases in mitochondrial membrane potential and intracellular reactive oxygen species levels. These protective effects were reversed by the selective mitochondrial adenosine triphosphate-sensitive potassium channel blocker 5-hydroxydecanoate. An inducible nitric oxide synthase inhibitor, Nω-nitro-L-arginine, also protected PC12 cells from amyloid-β peptide (25-35)-induced increases in both mitochondrial membrane potential and intracellular reactive oxygen species levels. However, the H2O2-degrading enzyme catalase could not reverse the amyloid-β peptide (25-35)-induced increase in intracellular reactive oxygen species. A 24-hour exposure to amyloid-β peptide (25-35) did not result in apoptosis or necrosis, suggesting that the increases in both mitochondrial membrane potential and reactive oxygen species levels preceded cell death. The data suggest that amyloid-β peptide (25-35) cytotoxicity is associated with adenosine triphosphate-sensitive potassium channels and nitric oxide. Regulation of adenosine triphosphate-sensitive potassium channels suppresses PC12 cell cytotoxicity induced by amyloid-β peptide (25-35).     

RETRACTED ARTICLE: Pro-protein convertase-2/carboxypeptidase-E mediated neuropeptide processing of RGC-5 cell after in vitro ischemia

Objective

To observe the change of the neuropeptide pro-protein processing system in the ischemic retina ganglion cell-5 (RGC-5) cells, pro-protein convertase-2 (PC2), carboxypeptidase-E (CPE) and preproneuropeptide Y (preproNPY) protein levels in the ischemic RGC-5 cells and conditioned medium were analyzed.

Methods

The RGC-5 cell was differentiated in 0.1 μmol/L staurosporine for 24 h and then stressed by different doses of oxygen and glucose deprivation (OGD). The acute or chronic OGD-induced cell death rates were obtained by using PI or TUNEL staining. The protein expression levels were determined by using the Western blot method and PC2 activity analysis.

Results

The ischemia caused substantial cell death in an OGD dose-dependent manner. In the cells, proPC2 and preproNPY protein levels gradually increased whereas proCPE gradually decreased. After OGD, PC2 activity was decreased. In the conditioned medium, proPC2 and PC2 proteins gradually decreased whereas proCPE, CPE, and preproNPY proteins gradually increased.

Conclusion

These results demonstrated that OGD inhibited the neuropeptide pro-protein processing system by reducing PC2 activity and the maturation of proPC2. The aggregation of the pro-proteins and the increase of the active CPE excision adversely exacerbated the cell injury. The pro-protein processing system might play a critical role in the ischemic stress of RGC-5 cells.     

Neuroprotective effect of ginkgolide K on glutamate-induced cytotoxicity in PC 12 cells via inhibition of ROS generation and Ca(2+) influx

Glutamate is considered to be responsible for the pathogenesis of cerebral ischemia disease. [Ca²⁺]_i influx and reactive oxygen species (ROS) production are considered to be involved in glutamate-induced apoptosis process. In this study, we investigated the neuroprotective effects of ginkgolide K in the glutamate-induced rat's adrenal pheochromocytoma cell line (PC 12 cells) and the possible mechanism. Glutamate cytotoxicity in PC 12 cells was accompanied by an increment of malondialdehyde (MDA) content and lactate dehydrogenase (LDH) release, as well as Ca²⁺ influx, bax/bcl-2 ratio, cytochrome c release, caspase-3 protein and ROS generation, and reduction of cell viability and mitochondrial membrane potential (MMP). Moreover, treatment with glutamate alone resulted in decrease activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX) activity. However, pretreatment with ginkgolide K significantly reduced MDA content, LDH release, as well as Ca²⁺ influx, cytochrome c release, bax/bcl-2 ratio, caspase-3 protein and ROS production, and attenuated the decrease of cells viability and MMP. In addition, ginkgolide K remarkedly up-regulated SOD and GSH-PX activities. All these findings indicated that ginkgolide K protected PC12 cells against glutamate-induced apoptosis by inhibiting Ca²⁺ influx and ROS production. Therefore, the present study supports the notion that ginkgolide K may be a promising neuroprotective agent for the treatment of cerebral ischemia disease.     

N-(quinolin-2-ylmethyl)butane-1,4-diamine,a polyamine analogue,attenuated injury in in vitro and in vivo models of cerebral ischemia

It has been widely recognized that glutamate (Glu)-induced cytotoxicity, intracellular calcium overload and excessive free radical production are the key players in the development and progression of ischemic brain injury. Since MK-801, an antagonist of N-methyl-d-aspartate (NMDA) receptor, showed many adverse reactions that hampered its clinical applications, development of safe and effective agent for the treatment of cerebral ischemia is eagerly required. This study was to investigate the effects of N¹-(quinolin-2-ylmethyl)butane-1,4-diamine (QMA), a polyamine analogue, on the in vitro and in vivo models of cerebral ischemic damage. The results revealed that pretreatment with QMA could attenuate Glu, putrescine (Put) and oxygen-glucose deprivation (OGD)-induced cell death, lipid peroxidation as well as the elevation of reactive oxygen species (ROS) and intracellular [Ca²⁺]_i in pheochromocytoma (PC12) cells and in rat primary cortical neurons. The results also demonstrated that QMA could inhibit NMDA-mediated intracellular [Ca²⁺]_i accumulation in rat primary cortical neurons and reduce brain infarct volume in middle cerebral artery occlusion (MCAO) rats. The present report suggested that polyamines played a crucial role in the pathological processes of cerebral ischemic damage and that QMA or other novel polyamine analogues could be promising therapeutic candidates for stroke by virtue of their anti-hypoxia and antioxidation property.     

Neuroprotective effects of the beta-catenin stabilization in an oxygen- and glucose-deprived human neural progenitor cell culture system

β-Catenin stabilization achieved either via GSK-3β specific inhibition or involving canonical Wnt signalling pathway, contributes to neuroprotection in an oxygen-glucose deprivation (4 h OGD) in vitro hypoxia model performed on human cortical neural progenitor cells previously differentiated into neurons and glia. Neuroprotection mechanisms include both acquiring tolerance to injury throughout preconditioning (72 h prior to OGD) or being pro-survival during 24 h reoxygenation after the insult. Four hours of OGD induced apoptotic cell death elevation to 73 ± 1% vs. 12% measured in control and the LDH level, indicative of necrotic cell injury, elevation by 67 ± 7% (set to 100%). A significant reduction in apoptosis occurred at 24 h reoxygenation with indirubin supplement which was 49 ± 6% at 2.5 μM BIO while LDH level was only 47 ± 5% of OGD. Kenpaullone was efficient in reducing both cell deaths at 5 μM (apoptosis 38 ± 1% and necrosis 33 ± 3% less than in OGD). Wnt agonist reduced apoptosis to 45 ± 3% at 0.01 μM, while LDH value was decreased to a level of 53 ± 5% of control. Our findings suggest that GSK-3beta inhibitors/β-catenin stabilizers may ultimately be useful drugs in neuroprotection and neuroregeneration therapies in vivo.     

Cinepazide maleate protects PC12 cells against oxygen–glucose deprivation-induced injury

Our previous study showed that cinepazide maleate (CM) was as effective and safe as mildronate in the treatment of acute ischemic stroke in a randomized, double-blind, active-controlled phase II multicenter trial, but underlying mechanism(s) is not well understood. As an extending study, here we demonstrated that CM could protect neuronal cells by affecting mitochondrial functions. PC12 cells were exposed to 2.5 h oxygen–glucose deprivation (OGD) followed by a 24 h reoxygenation, and then treated with different concentrations (1, 10, 100 μM) of CM. Among various concentrations, 10 μM CM exhibited most significant protection on PC12 cells against OGD injury. CM was found to suppress OGD-induced oxidative stress, as supported by its capability of reducing intracellular reactive oxygen species and malondialdehyde production and enhancing superoxide dismutase activity. Importantly, our results showed that CM could preserve mitochondrial functions, as revealed by its capability of stabilizing mitochondrial membrane potential, improving OGD-induced suppression of mitochondrial respiratory complex activities and enhancing ATP production. In summary, our present study provides the first evidence that CM can protect neuronal cells against OGD injury by preserving mitochondrial functions.     

Sulforaphane protects primary cultures of cortical neurons against injury induced by oxygen-glucose deprivation/reoxygenation via antiapoptosis

Objective To determine whether sulforaphane (SFN) protects neurons against injury caused by oxygen-glucose deprivation/reoxygenation (OGD/R) and, if so, to investigate the possible mechanisms. Methods Primary cultures of neurons were prepared from the cerebral cortex of 1-day-old Sprague-Dawley rats. On days 5-6 in vitro, the neurons were exposed to OGD for 1 h, followed by reoxygenation for 24 h. Cells were treated with 0, 0.1, 0.2, 0.5, 1, 2.5, or 5 μmol/L SFN, with or without 10 μmol/L LY294002, a PI3K-specific inhibitor, during OGD/R (a total of 25 h). After 24-h reoxygenation, MTT was used to assess viability and injury was assessed by Hoechst 33258/propidium iodide (PI) staining; immunofluorescence staining and Western blot were performed to detect molecular events associated with apoptosis. Results The MTT assay showed that 1 μmol/L SFN significantly increased viability, and Hoechst 33258/PI staining showed that the numbers of injured neurons were reduced significantly in the SFN group. Furthermore, immunofluorescence staining and Western blot showed that SFN increased Bcl-2 and decreased cleaved caspase-3 levels. Moreover, LY294002 inhibited the phosphorylated-Akt expression evoked by SFN, decreased Bcl-2 expression and increased cleaved caspase-3 expression. Conclusion SFN protects neurons against injury from OGD/R and this effect may be partly associated with an anti-apoptosis pathway.     

Neuroprotective effects of carnosine and homocarnosine on pheochromocytoma PC12 cells exposed to ischemia

The development of neuroprotective drugs against ischemic insults is hampered by the lack of pharmacological in vitro models. We developed an ischemic model using PC12 cell cultures exposed to oxygen-glucose-deprivation (OGD) followed by reoxygenation (18 hr) under regular atmospheric oxygen level. The toxicity induced in this model, that is partially caused by generation of reactive oxygen species (ROS), was measured morphologically as well as by the release of lactate dehydrogenase (LDH) and the prostaglandin PGE(2) from the cells. Carnosine and homocarnosine, histidine dipeptides antioxidants, found in high concentration in the brain, have been suggested to provide neuroprotection. Using the OGD model we found that 5 mM carnosine and 1 mM homocarnosine provided maximal neuroprotection of about 50% against OGD insult. This neuroprotective effect was similar to that of a known antioxidant, 4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl (tempol), and was not observed in a serum-deprivation toxicity model of PC12 cells, indicating that carnosine and homocarnosine may act as antioxidant-neuroprotective agents in the brain. Our ischemic model may provide a useful tool for investigating the mechanisms involved in the neuroprotection afforded by histidine dipeptides.     

Nordihydroguaiaretic acid protects hippocampal neurons against amyloid β-peptide toxicity, and attenuates free radical and calcium accumulation

Recent findings indicate that amyloid β-peptide (Aβ) can be neurotoxic by a mechanism involving an increase in the concentration of intracellular free Ca²⁺ ([Ca²⁺]_i) and the generation of free radicals. In the present study, the lipoxygenase inhibitor/antioxidant nordihydroguaiaretic acid (NDGA) protected cultured rat hippocampal neurons against the toxicity of Aβ in a concentration-dependent manner. Measurements of cellular oxidation (using the oxidation-sensitive dye 2,7-dichlorofluorescin) and intracellular free Ca²⁺ levels (using the Ca²⁺ indicator dye fura-2), showed that NDGA suppressed Aβ-induced accumulation of reactive oxygen species (ROS) and Ca²⁺; Ca²⁺ responses to glutamate were also suppressed by NDGA. NDGA prevented neuronal injury and accumulation of ROS induced by iron, indicating a role for NDGA as an antioxidant in NDGA-mediated neuroprotection. Another lipoxygenase inhibitor (AA861) also protected against Aβ and iron toxicity whereas the the 5-lipoxygenase-activating protein inhibitor L655,238 and the cyclooxygenase inhibitor indomethacin were ineffective. These findings suggest that NDGA can interupt a neurodegenerative pathway relevant to the pathophysiology of Alzheimer's disease.     

Antidepressant-like effect of trans-resveratrol in chronic stress model: Behavioral and neurochemical evidences

Trans-resveratrol is a phenolic compound enriched in polygonum cuspidatum and has diverse biological activities. There is only limited information about the antidepressant-like effect of trans-resveratrol. The present study investigated whether trans-resveratrol has antidepressant-like activity in rats exposed to chronic stress by using two behavioral tasks, shuttle box and sucrose preference tests. The monoamines (5-HT, noradrenaline and dopamine) and their metabolites as well as monoamine oxidase (MAO) enzyme activities in different brain regions were also measured. Compared to unstressed rats, those exposed to chronic stress paradigm showed performance deficits in the shuttle box, reduced sucrose preference, less weight gain and the increase in the ratio of adrenal gland to body weight, which were reversed by chronic treatment with trans-resveratrol (40 and 80 mg/kg, i.g.). The neurochemical assay showed that higher dose of trans-resveratrol (80 mg/kg) produced a marked increase of 5-HT levels in three brain regions, the frontal cortex, hippocampus and hypothalamus. Noradrenaline and dopamine levels were also increased both in the frontal cortex and striatum. Furthermore, chronic treatment with trans-resveratrol was found to inhibit monoamine oxidase-A (MAO-A) activity in all the four brain regions, particularly in the frontal cortex and hippocampus; while MAO-B activity was not affected. These findings indicate that the antidepressant-like effect of trans-resveratrol involves the regulation of the central serotonin and noradrenaline levels and the related MAO-A activities.     

Differential vulnerability of immature murine neurons to oxygen-glucose deprivation

In vivo studies support selective neuronal vulnerability to hypoxia-ischemia (HI) in the developing brain. Since differences in intrinsic properties of neurons might be responsible, pure cultures containing immature neurons (6-8 days in vitro) isolated from mouse cortex and hippocampus, regions chosen for their marked vulnerability to oxidative stress, were studied under in vitro ischemic conditions-oxygen-glucose deprivation (OGD). Twenty-four hours of reoxygenation after 2.5 h of OGD induced significantly greater cell death in hippocampal than in cortical neurons (67.8% vs. 33.4%, P = 0.0068). The expression of neuronal nitric oxide synthase (nNOS) protein, production of nitric oxide (NO), and reactive oxygen species (ROS), as well as glutathione peroxidase (GPx) activity and intracellular levels of reduced glutathione (GSH), were measured as indicators of oxidative stress. Hippocampal neurons had markedly higher nNOS expression than cortical neurons by 24 h of reoxygenation, which coincided with an increase in NO production, and significantly greater ROS accumulation. GPx activity declined significantly in hippocampal but not in cortical neurons at 4 and 24 h after OGD. The decrease in GSH level in hippocampal neurons correlated with the decline of GPx activity. Our data suggest that developing hippocampal neurons are more sensitive to OGD than cortical neurons. This finding supports our in vivo studies showing that mouse hippocampus is more vulnerable than cortex after neonatal HI. An imbalance between excess prooxidant production (increased nNOS expression, and NO and ROS production) and insufficient antioxidant defenses created by reduced GPx activity and GSH levels may, in part, explain the higher susceptibility to OGD of immature hippocampal neurons.     

Zn2+‐induced ERK activation mediates PARP‐1‐dependent ischemic‐reoxygenation damage to oligodendrocytes

Much of the cell death following episodes of anoxia and ischemia in the mammalian central nervous system has been attributed to extracellular accumulation of glutamate and ATP, which causes a rise in [Ca²⁺]_i, loss of mitochondrial potential, and cell death. However, restoration of blood flow and reoxygenation are frequently associated with exacerbation of tissue injury (the oxygen paradox). Herein we describe a novel signaling pathway that is activated during ischemia‐like conditions (oxygen and glucose deprivation; OGD) and contributes to ischemia‐induced oligodendroglial cell death. OGD induced a retarded and sustained increase in extracellular signal‐regulated kinase 1/2 (ERK1/2) phosphorylation after restoring glucose and O₂ (reperfusion‐like conditions). Blocking the ERK1/2 pathway with the MEK inhibitor UO126 largely protected oligodendrocytes against ischemic insults. ERK1/2 activation was blocked by the high‐affinity Zn²⁺ chelator TPEN, but not by antagonists of AMPA/kainate or P2X7 receptors that were previously shown to be involved in ischemic oligodendroglial cell death. Using a high‐affinity Zn²⁺ probe, we showed that ischemia induced an intracellular Zn²⁺ rise in oligodendrocytes, and that incubation with TPEN prevented mitochondrial depolarization and ROS generation after ischemia. Accordingly, exposure to TPEN and the antioxidant Trolox reduced ischemia‐induced oligodendrocyte death. Moreover, UO126 blocked the ischemia‐induced increase in poly‐[ADP]‐ribosylation of proteins, and the poly[ADP]‐ribose polymerase 1 (PARP‐1) inhibitor DPQ significantly inhibited ischemia‐induced oligodendroglial cell death—demonstrating that PARP‐1 was required downstream in the Zn²⁺‐ERK oligodendrocyte cell death pathway. Chelation of cytosolic Zn²⁺, blocking ERK signaling, and antioxidants may be beneficial for treating CNS white matter ischemia‐reperfusion injury. Importantly, all the inhibitors of this pathway protected oligodendrocytes when applied after the ischemic insult. © 2012 Wiley Periodicals, Inc.     

N-methyl-d-aspartate and hypoxia induced Ca-changes in the CA1 region of the hippocampal slice

Hypoxic neuronal depolarization was accompanied by a large decrease in extracellular [Ca²⁺]. After reoxygenation, the time at which [Ca²⁺] normalized was correlated with the extent of recovery of N-methyl-d-aspartate (NMDA) and synaptic responses. There was no evidence that the NMDA receptor system was more disrupted following hypoxia than the receptors involved in synaptic transmission. The Na⁺/K⁺ pump appeared to be better able to recover from hypoxia than the NMDA responses or synaptic transmission.     

Different amyloidogenic peptides share a similar mechanism of neurotoxicity involving reactive oxygen species and calcium

The amyloid β-peptide (Aβ) that accumulates as insoluble plaques in the brains of Alzheimer's victims can be neurotoxic, by a mechanism that may involve generation of reactive oxygen species (ROS) and destabilization of cellular calcium homeostasis. We now provide evidence that the mechanism of neurotoxicity of two other amyloidogenic peptides (APs), human amylin and β2-microglobulin, also involves induction of ROS and elevation of [Ca²⁺]_i. Human amylin, β2-microglobulin and Aβ1–40 all caused significant death of neurons in rat hippocampal cell cultures during 24–48h exposure periods. Rat amylin, a non-AP, was not neurotoxic. Each AP caused an elevation of rest [Ca²⁺]_i during a 20 h exposure period, and promoted a sustained elevation of [Ca²⁺]_i following exposure to glutamate which was significantly greater than controls. Each AP induced accumulation of ROS in neurons which preceded elevation of [Ca²⁺]_i. Several antioxidants, including propyl gallate, vitamin E and the spin-trapping compound N-tert-butyl-α-phenylnitrone attenuated the elevation of [Ca²⁺]_i and neurotoxicity induced by the peptides. The data indicate that different APs share a common mechanism of neurotoxicity involving free radical accumulation and destabilization of [Ca²⁺]_i homeostasis.     

rAAV/ABAD-DP-6His attenuates oxidative stress- induced injury of PC 12 cells

Our previous studies have revealed that amyloid β(Aβ)-binding alcohol dehydrogenase(ABAD) decoy peptide antagonizes Aβ42-induced neurotoxicity. However, whether it improves oxidative stress injury remains unclear. In this study, a recombinant adenovirus constitutively secreting and expressing Aβ-ABAD decoy peptide(rAAV/ABAD-DP-6His) was successfully constructed. Our results showed that rAAV/ABAD-DP-6His increased superoxide dismutase activity in hydrogen peroxide-induced oxidative stress-mediated injury of PC12 cells. Moreover, rAAV/ABADDP-6His decreased malondialdehyde content, intracellular Ca2+ concentration, and the level of reactive oxygen species. rAAV/ABAD-DP-6His maintained the stability of the mitochondrial membrane potential. In addition, the ATP level remained constant, and apoptosis was reduced. Overall, the results indicate that rAAV/ABAD-DP-6His generates the fusion peptide, Aβ-ABAD decoy peptide, which effectively protects PC12 cells from oxidative stress injury induced by hydrogen peroxide, thus exerting neuroprotective effects.     

Antioxidant effects of the orientin and vitexin in Trollius chinensis Bunge in D-galactose-aged mice

Total flavonoids are the main pharmaceutical components of Trollius chinensis Bunge, and orientin and vitexin are the monomer components of total flavonoids in Trollius chinensis Bunge. In this study, an aged mouse model was established through intraperitoneal injection of D-galactose for 8 weeks, followed by treatment with 40, 20, or 10 mg/kg orientin, vitexin, or a positive control (vitamin E) via intragastric administration for an additional 8 weeks. Orientin, vitexin, and vitamin E improved the general medical status of the aging mice and significantly increased their brain weights. They also produced an obvious rise in total antioxidant capacity, superoxide dismutase, catalase, and glutathione peroxidase levels in the serum, and the levels of superoxide dismutase, catalase and glutathione peroxidase, Na+-K+-ATP enzyme, and Ca2+-Mg2+-ATP enzyme in the liver, brain and kidneys. In addition, they significantly reduced malondialdehyde levels in the liver, brain and kidney and lipofuscin levels in the brain. They also significantly improved the neuronal ultrastructure. The 40 mg/kg dose of orientin and vitexin had the same antioxidant capacity as vitamin E. These experimental findings indicate that orientin and vitexin engender anti-aging effects through their antioxidant capacities.     

Amyloid-β disrupts unitary calcium entry through endothelial NMDA receptors in mouse cerebral arteries

Transient increases in intracellular Ca²⁺ activate endothelium-dependent vasodilatory pathways. This process is impaired in cerebral amyloid angiopathy, where amyloid-β_(1-40) accumulates around blood vessels. In neurons, amyloid-β impairs the Ca²⁺-permeable N-methyl-D-aspartate receptor (NMDAR), a mediator of endothelium-dependent dilation in arteries. We hypothesized that amyloid-β_(1-40) reduces NMDAR-elicited Ca²⁺ signals in mouse cerebral artery endothelial cells, blunting dilation. Cerebral arteries isolated from 4-5 months-old, male and female cdh5:Gcamp8 mice were used for imaging of unitary Ca²⁺ influx through NMDAR (NMDAR sparklets) and intracellular Ca²⁺ transients. The NMDAR agonist NMDA (10 µmol/L) increased frequency of NMDAR sparklets and intracellular Ca²⁺ transients in endothelial cells; these effects were prevented by NMDAR antagonists D-AP5 and MK-801. Next, we tested if amyloid-β_(1-40) impairs NMDAR-elicited Ca²⁺ transients. Cerebral arteries incubated with amyloid-β_(1-40) (5 µmol/L) exhibited reduced NMDAR sparklets and intracellular Ca²⁺ transients. Lastly, we observed that NMDA-induced dilation of pial arteries is reduced by acute intraluminal amyloid-β_(1-40), as well as in a mouse model of Alzheimer’s disease, the 5x-FAD, linked to downregulation of Grin1 mRNA compared to wild-type littermates. These data suggest that endothelial NMDAR mediate dilation via Ca²⁺-dependent pathways, a process disrupted by amyloid-β_(1-40) and impaired in 5x-FAD mice.