Zinc-induced Neuronal Death Research Articles

Carvacrol Inhibits Expression of Transient Receptor Potential Melastatin 7 Channels and Alleviates Zinc Neurotoxicity Induced by Traumatic Brain Injury.

Carvacrol is a monoterpenoid phenol produced by aromatic plants such as oregano. Although the exact mechanism by which carvacrol acts has not yet been established, it appears to inhibit transient receptor potential melastatin 7 (TRPM7), which modulates the homeostasis of metal ions such as zinc and calcium. Several studies have demonstrated that carvacrol has protective effects against zinc neurotoxicity after ischemia and epilepsy. However, to date, no studies have investigated the effect of carvacrol on traumatic brain injury (TBI)-induced zinc neurotoxicity. In the present study, we investigated the therapeutic potential of carvacrol for the prevention of zinc-induced neuronal death after TBI. Rats were subjected to a controlled cortical impact, and carvacrol was injected at a dose of 50 mg/kg. Histological analysis was performed at 12 h, 24 h, and 7 days after TBI. We found that carvacrol reduced TBI-induced TRPM7 over-expression and free zinc accumulation. As a result, subsequent oxidative stress, dendritic damage, and neuronal degeneration were decreased. Moreover, carvacrol not only reduced microglial activation and delayed neuronal death but also improved neurological outcomes after TBI. Taken together, these findings suggest that carvacrol administration may have therapeutic potential after TBI by preventing neuronal death through the inhibition of TRPM7 expression and alleviation of zinc neurotoxicity.

Zinc Accumulation Aggravates Cerebral Ischemia/Reperfusion Injury Through Inducing Endoplasmic Reticulum Stress.

Zinc is highly enriched in the central nervous system. Numerous evidences suggest that high concentration of zinc acts as a critical mediator of neuronal death in the ischemic brain, however, the possible mechanisms of neurotoxicity of zinc during cerebral ischemia/reperfusion (I/R) remain elusive. Endoplasmic reticulum (ER) is a storage location of intracellular zinc. ER stress related genes were up-regulated during zinc-induced neuronal death in vascular-type senile dementia. In the present study, we investigated whether intracellular accumulated zinc aggravates I/R injury through ER stress and ER stress-associated apoptosis. Male Sprague-Dawley rats were subjected to 90min middle cerebral artery occlusion (MCAO) and received either vehicle or zinc chelator TPEN 15mg/kg. The expression of ER stress related factors glucose-regulated protein 78 (GRP78) and phosphorylated eukaryotic initiation factor 2α (p-eIF2α), ER stress related apoptotic proteins CCAAT-enhancer-binding protein homologous protein (CHOP) and caspase-12, as well as anti-apoptotic factor B-cell lymphoma-2 (Bcl-2) were assessed 24h after reperfusion. Our results showed that the levels of GRP78 and p-eIF2α, as well as CHOP and caspase-12, were increased in ischemic brain, indicating that cerebral I/R triggers ER stress. Furthermore, GRP78, CHOP and caspase-12 were all colocalized with the zinc-specific dyes NG, suggesting that there is certain relationship between cytosolic labile zinc and ER stress following cerebral ischemia. Chelating zinc with TPEN reversed the expression of GRP78, p-eIF2α in ischemic rats. Moreover, CHOP and NeuN double staining positive cells, as well as caspase-12 and TUNEL double staining positive cells were also decreased after TPEN treatment, indicating that chelating zinc might inhibit ER stress and decreased ER stress associated neuronal apoptosis. In addition, TPEN treatment reversed the downregulated level of Bcl-2, which localized in the ER membrane and involved in the dysfunction of ER, confirming that the anti-apoptosis effects of chelating zinc following I/R are exerted via inhibition of the ER stress. Taken together, this study demonstrated that excessive zinc activates ER stress and zinc induced neuronal cell death is at least partially due to ER stress specific neuronal apoptosis in ischemic penumbra, which may provide an important mechanism of cerebral I/R injury.

A Novel Zinc Chelator, 1H10, Ameliorates Experimental Autoimmune Encephalomyelitis by Modulating Zinc Toxicity and AMPK Activation

Previous studies in our lab revealed that chemical zinc chelation or zinc transporter 3 (ZnT3) gene deletion suppresses the clinical features and neuropathological changes associated with experimental autoimmune encephalomyelitis (EAE). In addition, although protective functions are well documented for AMP-activated protein kinase (AMPK), paradoxically, disease-promoting effects have also been demonstrated for this enzyme. Recent studies have demonstrated that AMPK contributes to zinc-induced neurotoxicity and that 1H10, an inhibitor of AMPK, reduces zinc-induced neuronal death and protects against oxidative stress, excitotoxicity, and apoptosis. Here, we sought to evaluate the therapeutic efficacy of 1H10 against myelin oligodendrocyte glycoprotein 35-55-induced EAE. 1H10 (5 μg/kg) was intraperitoneally injected once per day for the entire experimental course. Histological evaluation was performed three weeks after the initial immunization. We found that 1H10 profoundly reduced the severity of the induced EAE and that there was a remarkable suppression of demyelination, microglial activation, and immune cell infiltration. 1H10 also remarkably inhibited EAE-associated blood-brain barrier (BBB) disruption, MMP-9 activation, and aberrant synaptic zinc patch formation. Furthermore, the present study showed that long-term treatment with 1H10 also reduced the clinical course of EAE. Therefore, the present study suggests that zinc chelation and AMPK inhibition with 1H10 may have great therapeutic potential for the treatment of multiple sclerosis.

Nickel Enhances Zinc-Induced Neuronal Cell Death by Priming the Endoplasmic Reticulum Stress Response.

Trace metals such as zinc (Zn), copper (Cu), and nickel (Ni) play important roles in various physiological functions such as immunity, cell division, and protein synthesis in a wide variety of species. However, excessive amounts of these trace metals cause disorders in various tissues of the central nervous system, respiratory system, and other vital organs. Our previous analysis focusing on neurotoxicity resulting from interactions between Zn and Cu revealed that Cu2+ markedly enhances Zn2+-induced neuronal cell death by activating oxidative stress and the endoplasmic reticulum (ER) stress response. However, neurotoxicity arising from interactions between zinc and metals other than copper has not been examined. Thus, in the current study, we examined the effect of Ni2+ on Zn2+-induced neurotoxicity. Initially, we found that nontoxic concentrations (0–60 μM) of Ni2+ enhance Zn2+-induced neurotoxicity in an immortalized hypothalamic neuronal cell line (GT1-7) in a dose-dependent manner. Next, we analyzed the mechanism enhancing neuronal cell death, focusing on the ER stress response. Our results revealed that Ni2+ treatment significantly primed the Zn2+-induced ER stress response, especially expression of the CCAAT-enhancer-binding protein homologous protein (CHOP). Finally, we examined the effect of carnosine (an endogenous peptide) on Ni2+/Zn2+-induced neurotoxicity and found that carnosine attenuated Ni2+/Zn2+-induced neuronal cell death and ER stress occurring before cell death. Based on our results, Ni2+ treatment significantly enhances Zn2+-induced neuronal cell death by priming the ER stress response. Thus, compounds that decrease the ER stress response, such as carnosine, may be beneficial for neurological diseases.

Involvement of SAPK/JNK Signaling Pathway in Copper Enhanced Zinc-Induced Neuronal Cell Death

Zinc (Zn) plays an important role in many organisms in various physiological functions such as cell division, immune mechanisms and protein synthesis. However, excessive Zn release is induced in pathological situations and causes neuronal cell death. Previously, we reported that Cu ions (Cu2+) markedly exacerbates Zn2+-induced neuronal cell death by potentiating oxidative stress and the endoplasmic reticulum stress response. In contrast, the stress-activated protein kinase/c-Jun amino-terminal kinase (SAPK/JNK) signaling pathway is important in neuronal cell death. Thus, in this study, we focused on the SAPK/JNK signaling pathway and examined its involvement in Cu2+/Zn2+-induced neurotoxicity. Initially, we examined expression of factors involved in the SAPK/JNK signaling pathway. Accordingly, we found that phosphorylated (ie, active) forms of SAPK/JNK (p46 and p54) are increased by CuCl2 and ZnCl2 co-treatment in hypothalamic neuronal mouse cells (GT1-7 cells). Downstream factors of SAPK/JNK, phospho-c-Jun, and phospho-activating transcription factor 2 are also induced by CuCl2 and ZnCl2 co-treatment. Moreover, an inhibitor of the SAPK/JNK signaling pathway, SP600125, significantly suppressed neuronal cell death and activation of the SAPK/JNK signaling pathway induced by CuCl2 and ZnCl2 cotreatment. Finally, we examined involvement of oxidative stress in activation of the SAPK/JNK signaling pathway, and found that human serum albumin-thioredoxin fusion protein, an antioxidative protein, suppresses activation of the SAPK/JNK signaling pathway. On the basis of these results, our findings suggest that activation of ZnCl2-dependent SAPK/JNK signaling pathway is important in neuronal cell death, and CuCl2-induced oxidative stress triggers the activation of this pathway.

Identifying New AMP-Activated Protein Kinase Inhibitors That Protect against Ischemic Brain Injury.

We recently reported that AMP-activated protein kinase (AMPK) contributes to zinc-induced neuronal death by inducing Bim, a pro-apoptotic Bcl-2 homology domain 3-only protein, in a liver kinase B1 (LKB1)-dependent manner. Current data suggest AMPK plays key roles in excitotoxicity and ischemic brain injury, with zinc neurotoxicity representing at least one mechanism of ischemic neuronal death. Inhibition of AMPK could be a viable therapeutic strategy to prevent ischemic brain injury following stroke. This prompted our search for novel inhibitors of AMPK activity and zinc-induced neuronal death using cultured mouse cortex and a rat model of brain injury after middle cerebral artery occlusion (MCAO). In structure-based virtual screening, 118 compounds were predicted to bind the active site of AMPK α2, and 40 showed in vitro AMPK α2 inhibitory activity comparable to compound C (a well-known, potent AMPK inhibitor). In mouse cortical neuronal cultures, 7 of 40 compound reduced zinc-induced neuronal death at levels comparable to compound C. Ultimately, only agents 2G11 and 1H10 significantly attenuated various types of neuronal death, including oxidative stress, excitotoxicity, and apoptosis. When administered as intracerebroventricular injections prior to permanent MCAO in rats, 2G11 and 1H10 reduced brain infarct volumes, whereas compound C did not. Therefore, these novel AMPK inhibitors could be drug development candidates to treat stroke.

Carvacrol Attenuates Hippocampal Neuronal Death after Global Cerebral Ischemia via Inhibition of Transient Receptor Potential Melastatin 7.

Over the last two decades, evidence supporting the concept of zinc-induced neuronal death has been introduced, and several intervention strategies have been investigated. Vesicular zinc is released into the synaptic cleft, where it then translocates to the cytoplasm, which leads to the production of reactive oxygen species and neurodegeneration. Carvacrol inhibits transient receptor potential melastatin 7 (TRPM7), which regulates the homeostasis of extracellular metal ions, such as calcium and zinc. In the present study, we test whether carvacrol displays any neuroprotective effects after global cerebral ischemia (GCI), via a blockade of zinc influx. To test our hypothesis, we used eight-week-old male Sprague–Dawley rats, and a GCI model was induced by bilateral common carotid artery occlusion (CCAO), accompanied by blood withdrawal from the femoral artery. Ischemic duration was defined as a seven-minute electroencephalographic (EEG) isoelectric period. Carvacrol (50 mg/kg) was injected into the intraperitoneal space once per day for three days after the onset of GCI. The present study found that administration of carvacrol significantly decreased the number of degenerating neurons, microglial activation, oxidative damage, and zinc translocation after GCI, via downregulation of TRPM7 channels. These findings suggest that carvacrol, a TRPM7 inhibitor, may have therapeutic potential after GCI by reducing intracellular zinc translocation.

Prevention of hypoglycemia-induced hippocampal neuronal death by N-acetyl-L-cysteine (NAC).

Type 1 and type 2 diabetic patients who are treated with insulin or other blood glucose reducing agents for tight control of blood glucose levels are frequently at risk of experiencing severe hypoglycemia which can lead to seizures, loss of consciousness and death. Hypoglycemic neuronal cell death is not a simple result of low glucose supply to the brain, but, instead, results from a cell death signaling pathway that is started by the re-administration of glucose after glucose deprivation. Zinc is a biologically important element for physiological function of central nervous system. However, excessive zinc release from the presynaptic terminals and subsequent translocation into the postsynaptic neurons may contribute to neuronal death following hypoglycemia. N-acetyl-L-cysteine (NAC) acts as a zinc chelator that alleviates zinc-induced neuronal death processes. In addition, NAC restores levels of neuronal glutathione (GSH), a potent antioxidant, by providing a cell-permeable source of cysteine. Thus, we hypothesized that NAC treatment can reduce neuronal cell death, not only by increasing GSH concentration but also by zinc chelation. As a result, we found that NAC decreased the oxidative stress, zinc release and translocation, and improved the level of glutathione. Therefore, NAC administration alleviated hippocampal neuron death in hypoglycemia-induced rats.

AMP-activated protein kinase contributes to zinc-induced neuronal death via activation by LKB1 and induction of Bim in mouse cortical cultures.

BackgroundWe reported that zinc neurotoxicity, a key mechanism of ischemic neuronal death, was mediated by poly ADP-ribose polymerase (PARP) over-activation following NAD+/ATP depletion in cortical cultures. Because AMP-activated protein kinase (AMPK) can be activated by ATP depletion, and AMPK plays a key role in excitotoxicity and ischemic neuronal death, we examined whether AMPK could be involved in zinc neurotoxicity in mouse cortical neuronal cultures.ResultsCompound C, an AMPK inhibitor, significantly attenuated zinc-induced neuronal death. Activation of AMPK was detected beginning 2 h after a 10-min exposure of mouse cortical neurons to 300 μM zinc, although a significant change in AMP level was not detected until 4 h after zinc treatment. Thus, AMPK activation might not have been induced by an increase in intracellular AMP in zinc neurotoxicity. Furthermore, we observed that liver kinase B1 (LKB1) but not Ca2+/calmodulin-dependent protein kinase kinase β (CaMKKβ), was involved in AMPK activation. Although STO-609, a chemical inhibitor of CaMKKβ, significantly attenuated zinc neurotoxicity, zinc-induced AMPK activation was not affected, which suggested that CaMKKβ was not involved in AMPK activation. Knockdown of LKB1 by siRNA significantly reduced zinc neurotoxicity, as well as zinc-induced AMPK activation, which indicated a possible role for LKB1 as an upstream kinase for AMPK activation. In addition, mRNA and protein levels of Bim, a pro-apoptotic Bcl-2 family member, were noticeably increased by zinc in an AMPK-dependent manner. Finally, caspase-3 activation in zinc-induced neuronal death was mediated by LKB1 and AMPK activation.ConclusionsThe results suggested that AMPK mediated zinc-induced neuronal death via up-regulation of Bim and activation of caspase-3. Rapid activation of AMPK was detected after exposure of cortical neuronal cultures to zinc, which was induced by LKB1 activation but not increased intracellular AMP levels or CaMKKβ activation. Hence, blockade of AMPK in the brain may protect against zinc neurotoxicity, which is likely to occur after acute brain injury.

Ratiometric fluorescence imaging of lysosomal Zn2+ release under oxidative stress in neural stem cells.

Zinc dyshomeostasis is a major mechanism of neuronal death, which is involved in many different neuropathological conditions. Lysosomal membrane permeabilisation has an important function in zinc-induced neuronal death under oxidative stress. To investigate lysosomal zinc functions in neurons with high spatial and temporal reliability, we report a ratiometric probe, LysoZn-1. It is derived from the styryl-BODIPY-DPA scaffold with a lysosome-targeted 2-morpholinoethylamine moiety to allow localisation in lysosomes. The electron donor at the meso-position of the BODIPY fluorophore makes the present probe prefer complexing with Zn2+ rather than Cd2+, which can be explained by HSAB (Hard-Soft Acid-Base) theory and was confirmed by Gaussian calculation. Upon Zn2+ binding, LysoZn-1 exhibits obvious fluorescence enhancement (F578 nm) and ratio (F578 nm/F680 nm) changes. The emission intensities of LysoZn-1 and LysoZn-1+Zn2+ do not change significantly under lysosomal pH ranging from 4.5 to 6.0. Confocal imaging experiments indicate that LysoZn-1 is able to localise to lysosomes in neural stem cells (NSCs), MCF-7 and Hela cells and detect exogenous Zn2+ levels in NSCs and MCF-7 cells. LysoZn-1 function is not disturbed by chloroquine in living cells. Furthermore, increases in lysosomal Zn2+ concentration upon H2O2 stimulation in NSCs are observed using LysoZn-1.

D-Histidine and l-histidine attenuate zinc-induced neuronal death in GT1-7 cells

Although zinc (Zn) is an essential trace element, excess Zn causes neuronal death following transient global ischemia and plays a central role in the pathogenesis of vascular-type dementia. In this study, we developed a rapid and convenient screening system for substances that prevent Zn-induced neurotoxicity by using GT1-7 cells (immortalized hypothalamic neurons), with the aim of identifying a treatment for vascular-type dementia. Among tested, we found a protective substance in the extract of round herring (Etrumeus teres), and determined its structure as l-histidine. Analysis of the structure-activity relationship by using histidine analogues revealed that both l-histidine and d-histidine exhibit the same neuroprotective activity. Furthermore, we investigated the molecular mechanisms underlying the protective effect of histidine on Zn-induced neurotoxicity using Zn imaging and gene expression analysis, and found that histidine protects against Zn-induced neurotoxicity not by inhibiting Zn chelation, thereby preventing increases in intracellular Zn(2+). Moreover, it is also suggested that endoplasmic reticulum (ER) stress and activity-regulated cytoskeleton associated protein (Arc) are implicated in Zn-induced degeneration of neurons.

Protective Substances Against Zinc-Induced Neuronal Death after Ischemia:Carnosine as a Target for Drug of Vascular Type of Dementia

Recent studies have indicated the significance of zinc in neurodegeneration after transient global ischemia. After ischemia, excess glutamate and zinc, which are released in the synaptic clefts, cause the apoptotic death of the target neurons, and finally lead the pathogenesis of vascular type of dementia. Considering the removal of zinc using zinc-sensitive chelators was effective in the prevention of neuronal death after transient global ischemia, it is highly possible that substances which protect against zinc-induced neuronal death will become a candidate for drugs of vascular type of dementia. Based on this 'zinc hypothesis', we have searched for such substances among various agricultural products including fruits, vegetables, and fishes using our developed in vitro screening system. Among tested, we found that carnosine (beta-alanyl histidine) protected against zinc-induced death of cultured neurons, and have applied for the patent as a drug of ischemia-induced neuronal death and the treatment/prevention for vascular type of dementia (application No. 2006-145857) in Japan. Here, we review the perspective of protective substances of zinc-induced neuronal death as a drug of vascular type of dementia based on our studies and other numerous studies.

Overexpression of DSCR1 blocks zinc-induced neuronal cell death through the formation of nuclear aggregates

The Down syndrome (DS) candidate region gene 1 (DSCR1) is localized near DS critical region on chromosome 21 and is overexpressed in the brains of DS patients. Although DSCR1 was known for a modulator of calcineurin, the overexpression of DSCR1 is thought to play a role in neuronal cell death. Zinc, one of the most abundant transition metals in the brain, may also contribute to selective neuronal cell death when present in excessive amounts. In the present study, we investigated the effect of DSCR1 overexpression on zinc-induced cell death in hippocampal neuroprogenitor cells. The overexpression of DSCR1 caused apoptotic cell death without an apparent formation of intracellular protein inclusions. Upon exposure to zinc, soluble DSCR1 levels were significantly decreased and insoluble levels were enhanced to a similar extent, which were partially caused by the zinc-induced inhibition of proteasomal activity and a consequently diminished degradation of DSCR1. Furthermore, zinc treatment induced the formation of nuclear DSCR1 aggregates, which blocked zinc-induced cell death. These findings indicate that, although the up-regulation of DSCR1 levels exerts a cytotoxic effect, the addition of zinc leads to the formation of cytoprotective nuclear aggregates in neuronal cells.

Zinc induces cell death in immortalized embryonic hippocampal cells via activation of Akt-GSK-3β signaling

Zinc is an essential catalytic and structural element of many proteins and a signaling messenger that is released by neuronal activity at many central excitatory synapses. Excessive synaptic release of zinc followed by entry into vulnerable neurons contributes severe neuronal cell death. We have previously observed that zinc-induced neuronal cell death is accompanied by Akt activation in embryonic hippocampal progenitor (H19-7) cells. In the present study, we examined the role of Akt activation and its downstream signaling events during extracellular zinc-induced neuronal cell death. Treatment of H19-7 cells with 10 μM of zinc plus zinc ionophore, pyrithione, led to increased phosphorylation of Akt at Ser-473/Thr-308 and increased Akt kinase activity. Zinc-induced Akt activation was accompanied by increased Tyr-phosphorylated GSK-3β as well as increased GSK-3β kinase activity. Transient overexpression of a kinase-deficient Akt mutant remarkably suppressed GSK-3β activation and cell death. Furthermore, tau phosphorylation, but not the degradation of β-catenin, was dependent upon zinc-induced GSK-3β activation and contributed to cell death. The current data suggest that, following exposure to zinc, the sequential activation of Akt and GSK-3β plays an important role directing hippocampal neural precursor cell death.

Protective activity of mango (Mangifera indica L.) fruit against a zinc-induced neuronal cell death is independent of its antioxidant activity

Protective activity of mango (<i>Mangifera indica</i> L.) fruit against a zinc-induced neuronal cell death is independent of its antioxidant activity

In vitro neuronal cytotoxicity of latex and nonlatex orthodontic elastics

Although the toxicity of many dental materials has been thoroughly investigated, the toxicity of orthodontic elastic materials has not been extensively tested. We evaluated the neurotoxicity of 3 latex and 3 nonlatex orthodontic elastics in murine cerebral cortical cell cultures. Standard-sized pieces of each material from 3 manufacturers (American, Masel, and GAC) were placed on culture well inserts, allowing the material to be exposed to the culture bathing media without causing physical disruption of the cells. Cell death was quantified by assaying the release of the cytosolic enzyme lactate dehydrogenase. Exposure of cortical cultures to the nonlatex elastics did not cause significant neuronal death, but exposure to each of the latex elastics resulted in significant neuronal death. The neuronal death induced by each of the latex elastics was blocked by adding the metal chelator, EDTA (calcium disodium ethylenediaminetetraacetate). Because many latexes use zinc-containing compounds in the prevulcanization process, and the death induced had characteristics similar to zinc-induced neuronal death, it seems likely that the toxicity of latex elastics was mediated by zinc release. Because ingestion of zinc is not a health risk, the results suggest that, in spite of the finding that latex elastics have higher in vitro cytotoxicity than nonlatex elastics, the use of latex elastics in orthodontics is acceptable.

Attenuation of zinc-induced neuronal death by the interaction of growth inhibitory factor with Rab3A in rat hippocampal neurons

We examined the protective effect of growth inhibitory factor (GIF) against zinc-induced neuronal death in rat hippocampal neurons. In an in vitro cell culture system, 300 μM Zn 2+ readily induced death of hippocampal neuronal cells, which was characterized by massive necrosis and a minor degree of apoptosis. Neither the addition of recombinant GIF nor Rab3A alone could rescue these cells from death. However, the combination of GIF with Rab3A could significantly enhance the survival of the hippocampal neurons. This result was supported by both Annexin -V FITC/propidium dual staining and chromosomal DNA analysis. These findings suggest that GIF may inhibit Zn 2+-induced neuronal death via its interaction with Rab3A.

Involvement of poly ADP ribosyl polymerase‐1 in acute but not chronic zinc toxicity

We have previously suggested that zinc-induced neuronal death may be mediated in part by inhibition of the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH), secondary to depletion of the essential cosubstrate NAD+. Given convergent evidence implicating the NAD+-catabolizing enzyme, poly ADP ribosyl polymerase (PARP) in mediating ATP depletion and neuronal death after excitotoxic and ischemic insults, we tested the specific hypothesis that the neuronal death induced by exposure to toxic levels of extracellular zinc might be partly mediated by PARP. PARP was activated in cultured mouse cortical astrocytes after a toxic acute Zn2+ exposure (350 microm Zn2+ for 15 min), but not in cortical neurons or glia after exposure to a toxic chronic Zn2+ exposure (40 microm Zn2+ for 1-4 h), an exposure sufficient to deplete NAD+ and ATP levels. Furthermore, the neurotoxicity induced by acute, but not chronic, Zn2+ exposure was reduced in mixed neuronal-glial cultures prepared from mutant mice lacking the PARP gene. These data suggest PARP activation may contribute to more fulminant forms of Zn2+-induced neuronal death.

Nerve Growth Factor-Dependent Regulation of NADE-Induced Apoptosis

The p75 neurotrophin receptor (p75NTR) is a member of the tumor necrosis factor receptor (TNFR) superfamily, and can mediate both cell survival and cell death in response to nerve growth factor (NGF). Based on the structural and functional differences between p75NTR and the related receptors Fas or TNFR, it has been suggested that these receptors have distinct signaling functions. NADE (p75NTR-associated cell death executor) is a p75NTR-associated protein that mediates apoptosis in response to NGF by interacting with the cell death domain of p75NTR. NADE has at least four isoforms, designated as NADE2, NADE3, NADE4/Bex1, and NADE5/Bex2. NADE plays a role in NGF-induced apoptosis in oligodendrocytes and in zinc-induced neuronal death. In this review, we focus on the proapoptotic actions of NADE that regulate p75NTR signaling in response to NGF.

Co-Induction of p75NTRand p75NTR-Associated Death Executor in Neurons After Zinc Exposure in Cortical Culture or Transient Ischemia in the Rat

Recently, a 22 kDa protein termed p75(NTR)-associated death executor (NADE) was discovered to be a necessary factor for p75(NTR)-mediated apoptosis in certain cells. However, the possible role for p75(NTR)/NADE in pathological neuronal death has yet been undetermined. In the present study, we have examined this possibility in vivo and in vitro. Exposure of cortical cultures to zinc induced both p75(NTR) and NADE in neurons, whereas exposure to NMDA, ionomycin, iron, or H(2)O(2) induced neither. In addition, zinc exposure increased neuronal NGF expression and its release into the medium. A function-blocking antibody of p75(NTR) (REX) inhibited association between p75(NTR) and NADE as well as neuronal death induced by zinc. Conversely, NGF augmented zinc-induced neuronal death. Caspase inhibitors reduced zinc-induced neuronal death, indicating that caspases were involved. Because reduction of NADE expression with cycloheximide or NADE antisense oligonucleotides attenuated zinc-induced neuronal death, NADE appears to contribute to p75(NTR)-induced cortical neuronal death as shown in other cells. Because zinc neurotoxicity may be a key mechanism of neuronal death after transient forebrain ischemia, we next examined this model. After ischemia, p75(NTR) and NADE were induced in degenerating rat hippocampal CA1 neurons. There was a close correlation between zinc accumulation and p75(NTR)/NADE induction. Suggesting the role of zinc here, injection of a metal chelator, CaEDTA, into the lateral ventricle completely blocked the induction of p75(NTR) and NADE. Our results suggest that co-induction of p75(NTR) and NADE plays a role in zinc-triggered neuronal death in vitro and in vivo.