Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.22.56 (caspase-3)
 35,750 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To clarify the controversy concerning whether the cell death of motor neurons in ALS is apoptosis, we investigated the expression of Apaf-1 and caspase-9 mRNA in spinal cord tissue obained at autopsy from patients with ALS and controls using RT-PCR; the presence of in situ nuclear DNA fragmentation in motor neurons by the TdT-mediated dUTP-biotin nick end-labeling (TUNEL) method; and immunocytochemical localization of Apaf-1 and caspase-3, which are known as promotors of apoptotic processes. Although Apaf-1 and caspase-9 mRNAs levels were increased in ALS, Apaf-1 immunoreactivity (IR) showed no significant difference between ALS and the control, and caspase-3 IR was not observed in ALS motoneurons, casting doubt on the notion that motor neurons in ALS undergo death by the classic apoptotic pathway. Although TUNEL-positive motor neurons were frequently observed in the anterior horn in ALS, these neurons always showed an atrophic cell body with a shrunken and pyknotic nucleus, indicating that they were at the terminal stage of degeneration. No apoptotic bodies were seen. These findings suggest that the mechanism of motor neuronal cell death in ALS might not be apoptosis, but some other as yet unidentified mechanism.
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PMID:Is motoneuronal cell death in amyotrophic lateral sclerosis apoptosis? 1638 89

EAAT2 is a high affinity, Na+-dependent glutamate transporter with predominant astroglial localization. It accounts for the clearance of the bulk of glutamate released at central nervous system synapses and therefore has a crucial role in shaping glutamatergic neurotransmission and limiting excitotoxicity. Caspase-3 activation and impairment in expression and activity of EAAT2 are two distinct molecular mechanisms occurring in human amyotrophic lateral sclerosis (ALS) and in the transgenic rodent model of the disease. Excitotoxicity caused by down-regulation of EAAT2 is thought to be a contributing factor to motor neuron death in ALS. In this study, we report the novel evidence that caspase-3 cleaves EAAT2 at a unique site located in the cytosolic C-terminal domain of the transporter, a finding that links excitotoxicity and activation of caspase-3 as converging mechanisms in the pathogenesis of ALS. Caspase-3 cleavage of EAAT2 leads to a drastic and selective inhibition of this transporter. Heterologous expression of mutant SOD1 proteins linked to the familial form of ALS leads to inhibition of EAAT2 through a mechanism that largely involves activation of caspase-3 and cleavage of the transporter. In addition, we found evidence in spinal cord homogenates of mutant SOD1 ALS mice of a truncated form of EAAT2, likely deriving from caspase-3-mediated proteolytic cleavage, which appeared concurrently to the loss of EAAT2 immunoreactivity and to increased expression of activated caspase-3. Taken together, our findings suggest that caspase-3 cleavage of EAAT2 is one mechanism responsible for the impairment of glutamate uptake in mutant SOD1-linked ALS.
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PMID:Caspase-3 cleaves and inactivates the glutamate transporter EAAT2. 1656 4

Minocycline, a clinically used tetracycline for over 40 years, crosses the blood-brain barrier and prevents caspase up-regulation. It reduces apoptosis in mouse models of Huntington's disease and familial amyotrophic lateral sclerosis (ALS) and is in clinical trial for sporadic ALS. Because apoptosis also occurs after brain and spinal cord (SCI) injury, its prevention may be useful in improving recovery. We analyzed minocycline's neuroprotective effects over 28 days following contusion SCI and found significant functional recovery compared to tetracycline. Histology, immunocytochemistry, and image analysis indicated statistically significant tissue sparing, reduced apoptosis and microgliosis, and less activated caspase-3 and substrate cleavage. Since our original report in abstract form, others have published both positive and negative effects of minocycline in various rodent models of SCI and with various routes of administration. We have since found decreased tumor necrosis factor-alpha, as well as caspase-3 mRNA expression, as possible mechanisms of action for minocycline's ameliorative action. These results support reports that modulating apoptosis, caspases, and microglia provide promising therapeutic targets for prevention and/or limiting the degree of functional loss after CNS trauma. Minocycline, and more potent chemically synthesized tetracyclines, may find a place in the therapeutic arsenal to promote recovery early after SCI in humans.
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PMID:Minocycline neuroprotects, reduces microgliosis, and inhibits caspase protease expression early after spinal cord injury. 1663 21

Recent investigations have indicated that the nucleocytoplasmic transport system is essential for maintaining cell viability and cellular functions and that its dysfunction could lead to certain disorders. To investigate the involvement of this system in the pathomechanisms of amyotrophic lateral sclerosis (ALS), we examined the immunohistochemical localization of proteins associated with nucleocytoplasmic transport in the lumbar spinal cord in a mutant SOD1 (G93A) transgenic mouse model of ALS. This model is widely used for ALS research, and the mutant mice are known to exhibit neuronal loss and Lewy body-like hyaline inclusions (LBHIs) in the anterior horns, similar to the pathology seen in familial ALS patients associated with an SOD1 mutation and in several other transgenic rodent models. Using antibodies against the importin beta family of proteins, the major carrier proteins of nucleocytoplasmic transport, and those against their adapter protein, importin alpha, we found that the immunoreactivities were decreased within the nuclei and increased within the cytoplasm of a subset of the surviving anterior horn cells of the transgenic mice. In addition, LBHIs were invariably reactive toward these antibodies. Furthermore, the immunoreactivities for histone H1 and beta-catenin, representative cargo proteins transported by importin beta-dependent and beta-independent nucleocytoplasmic transport pathways, respectively, showed distributions similar to those for importin beta family and importin alpha proteins. The altered distributions of these proteins were not associated with caspase-3 expression, suggesting that the findings are unlikely to be a manifestation of apoptotic processes. Chronological quantitative analysis of importin beta-immunostained sections from the transgenic mice revealed a statistically significant progressive decrease in the proportion of the anterior horn cells exhibiting a more intense reactivity for these proteins in the nucleus than in the cytoplasm. To the contrary, we found that the anterior horn cells with the immunoreactivity in their cytoplasm, being more pronounced than that in their nucleus, were significantly increased in number along with the disease progression. This is the first report investigating nucleocytoplasmic transport in the ALS model mouse, and our present results imply that its dysfunction could be involved in the pathomechanisms underlying ALS.
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PMID:Altered distributions of nucleocytoplasmic transport-related proteins in the spinal cord of a mouse model of amyotrophic lateral sclerosis. 1695 27

The purpose of this study is to evaluate neuroprotective effects of (-)-Epigallocatechin-3-gallate (EGCG) in a transgenic mouse model of Amyotrophic lateral sclerosis (ALS). SOD1-G93A transgenic mice and wild-type mice were randomly divided into EGCG-treated groups (10 mg/kg, p.o) and vehicle-treated control groups. Rotarod measurement was performed to assess the motor function of mice starting at the age of 70 days. Nissl staining to examine the number of motor neurons and CD11b immunohistochemical staining to evaluate activation of microglia in the lumbar spinal cords were conducted at the age of 120 days. In addition, for further observation of regulation of cell signaling pathways by EGCG, we used immunohistochemical analysis for nuclear factor kappa B (NF-kappaB) and cleaved caspase-3 as well as western blot analysis to determine the expression of nitric oxide synthase (iNOS) and NF-kappaB in the spinal cord. This study demonstrated that oral administration of EGCG beginning from a pre-symptomatic stage significantly delayed the onset of disease, and extended life span. Furthermore, EGCG-treated transgenic mice showed increased number of motor neurons, diminished microglial activation, reduced immunohistochemical reaction of NF-kappaB and cleaved caspase-3 as well as reduced protein level of iNOS and NF-kappaB in the spinal cords. In conclusion, this study provides further evidences that EGCG has multifunctional therapeutic effects in the mouse model of ALS.
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PMID:Neuroprotective effects of (-)-epigallocatechin-3-gallate in a transgenic mouse model of amyotrophic lateral sclerosis. 1702 48

Gulf War illness (GWI) affects a significant percentage of veterans of the 1991 conflict, but its origin remains unknown. Associated with some cases of GWI are increased incidences of amyotrophic lateral sclerosis and other neurological disorders. Whereas many environmental factors have been linked to GWI, the role of the anthrax vaccine has come under increasing scrutiny. Among the vaccine's potentially toxic components are the adjuvants aluminum hydroxide and squalene. To examine whether these compounds might contribute to neuronal deficits associated with GWI, an animal model for examining the potential neurological impact of aluminum hydroxide, squalene, or aluminum hydroxide combined with squalene was developed. Young, male colony CD-1 mice were injected with the adjuvants at doses equivalent to those given to US military service personnel. All mice were subjected to a battery of motor and cognitive-behavioral tests over a 6-mo period postinjections. Following sacrifice, central nervous system tissues were examined using immunohistochemistry for evidence of inflammation and cell death. Behavioral testing showed motor deficits in the aluminum treatment group that expressed as a progressive decrease in strength measured by the wire-mesh hang test (final deficit at 24 wk; about 50%). Significant cognitive deficits in water-maze learning were observed in the combined aluminum and squalene group (4.3 errors per trial) compared with the controls (0.2 errors per trial) after 20 wk. Apoptotic neurons were identified in aluminum-injected animals that showed significantly increased activated caspase-3 labeling in lumbar spinal cord (255%) and primary motor cortex (192%) compared with the controls. Aluminum-treated groups also showed significant motor neuron loss (35%) and increased numbers of astrocytes (350%) in the lumbar spinal cord. The findings suggest a possible role for the aluminum adjuvant in some neurological features associated with GWI and possibly an additional role for the combination of adjuvants.
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PMID:Aluminum adjuvant linked to Gulf War illness induces motor neuron death in mice. 1711 26

Amyotrophic lateral sclerosis (ALS) is an adult-onset neurodegenerative disorder in which excitotoxicity has been implicated as a cause for cell death. To examine neurofilament (NF) aggregate-mediated sensitization of motor neurons to NMDA excitotoxicity, we examined NMDA receptor expression and the impact of NO donors (NOC12 or NOC5) or sodium cyanide (NaCN) on calcium influx and viability in dissociated motor neurons derived from wt and hNFL+/+ (NF aggregate-forming) mice. Alterations in intracellular calcium were assayed using Oregon Green calcium dye and the extent of apoptosis using active caspase-3 immunoreactivity. Although NF aggregate-bearing neurons demonstrated increased intracellular calcium levels and enhanced cell death in response to NMDA receptor activation, this was not associated with increased NMDA receptor expression. The down-regulation of the NMDA receptor using NO donors decreased calcium influx and caspase-3 activation in aggregate-bearing neurons, but had no effect on wt cultures. The converse was observed with NaCN in which intracellular calcium levels increased significantly in wt cultures in association with increased cell death. No effect was observed in aggregate-bearing neurons. These findings suggest that the presence of NF aggregates renders motor neurons more susceptible to NMDA-mediated excitotoxicity, and that this can be reversed by NO.
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PMID:Loss of nitric oxide-mediated down-regulation of NMDA receptors in neurofilament aggregate-bearing motor neurons in vitro: implications for motor neuron disease. 1715 1

We have developed an organotypic culture technique that uses slices of chick embryo spinal cord, in which trophic requirements for long-term survival of mature motoneurons (MNs) were studied. Slices were obtained from E16 chick embryos and maintained for up to 28 days in vitro (DIV) in a basal medium. Under these conditions, most MNs died. To promote MN survival, 14 different trophic factors were assayed. Among these 14, glial cell line-derived neurotrophic factor (GDNF) and vascular endothelial growth factor were the most effective. GDNF was able to promote MN survival for at least 28 DIV. K(+) depolarization or caspase inhibition prevented MN death but also induced degenerative-like changes in rescued MNs. Agents that elevate cAMP levels promoted the survival of a proportion of MNs for at least 7 DIV. Examination of dying MNs revealed that, in addition to cells exhibiting a caspase-3-dependent apoptotic pattern, some MNs died by a caspase-3-independent mechanism and displayed autophagic vacuoles, an extremely convoluted nucleus, and a close association with microglia. This organotypic spinal cord slice culture may provide a convenient model for testing conditions that promote survival of mature-like MNs that are affected in late-onset MN disease such as amyotrophic lateral sclerosis.
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PMID:Survival and death of mature avian motoneurons in organotypic slice culture: trophic requirements for survival and different types of degeneration. 1729 60

We have previously shown that the co-localization of neuronal nitric oxide synthase (nNOS) with neurofilament (NF) aggregates in motor neurons derived from transgenic mice over-expressing the human low molecular weight NF protein (hNFL+/+) is associated with a deregulation of calcium influx via the N-methyl-d-aspartate (NMDA) receptor, resulting in apoptosis. Because the absence of the GluR2 subunit of the alpha-amino-3-hydroxy-5-methyl-4-isoazolepropionic acid (AMPA) receptor confers calcium permeability and has been implicated in the process of excitotoxicity in ALS, we have examined the role of the AMPA receptor in this model. GluR2 protein expression and mRNA were examined in hNFL+/+ and wild-type motor neurons (wt). Live cell calcium imaging was performed using Oregon-Green Bapta and Fura-2 calcium dyes. For apoptotic studies, neurons were treated with glutamate, with or without glutamate receptor antagonists [6-cyano-7-nitroquinoxaline-2, 3-dione (CNQX) or (+)-5-methyl-10, 11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801)] and examined for active caspase-3 or phospholipid inversion. We observed that although both GluR2 mRNA and protein levels were decreased in hNFL+/+ motor neurons compared to wt, there was no appreciable calcium influx via the AMPA receptor. These studies demonstrate that calcium mediated excitotoxicity in NF aggregate-bearing neurons is NMDA receptor dependant.
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PMID:Calcium mediated excitotoxicity in neurofilament aggregate-bearing neurons in vitro is NMDA receptor dependant. 1736 87

Mutations in copper/zinc superoxide dismutase (SOD1) have been implicated in the pathogenesis of familial amyotrophic lateral sclerosis (ALS). Mutant SOD1 protein likely gains a novel cytotoxic property, leading to the death of motor neurons. We therefore investigated whether caspase-mediated apoptosis is associated with novel cytotoxic properties in a rodent model for familial ALS (G93A SOD1 transgenic mice). Caspase-9 (an effecter in the mitochondrial apoptotic pathway), caspase-8 (an effecter in the Fas apoptotic pathway), and caspase-3 (an executioner of both pathways) proteins were all present in nonactive forms in the spinal cords of wild-type mice during the early stage of the disease (8 weeks), at which time the mice had not yet exhibited motor paralysis. In transgenic mice, however, these proteins were present in their active forms, and their mRNA levels were significantly upregulated in the represent to this conversion from nonactive to active forms. During the advanced stage of the disease (16 weeks), when paralysis was evident, the active caspase levels were further elevated. On the other hand, the mRNA and protein levels of survivin, a counteraction protein against caspases, were significantly suppressed during the early stage, and sharply increased during the advanced stage. Although the mRNA and protein levels of X-linked inhibitor of apoptosis protein (XIAP) remained at the same levels as those seen in the control (wild-type mice) during the early stage, they were significantly depressed at an age of 16 weeks. These findings were observed exclusively in the spinal cord, the region responsible for the disease, and not in the cerebellum, a non-responsible region. We conclude that conditions facilitating the apoptotic process during the early stage of the disease play causative roles in the pathogenesis of ALS and that the suppression of XIAP levels during the advanced stage could contribute to disease expression and/or progression.
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PMID:Dysequilibrium between caspases and their inhibitors in a mouse model for amyotrophic lateral sclerosis. 1739 13


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