bluuebunny/crossref_metadata_2025 huggingface dataset:
Macromolecular complex formation between mycotoxin zearalenon and dophamine‐beta‐monooxygenase
Authors: Boyajyan A. S. , Karagyozyan M. K.
Published: July 2010

Micotoxin zearalenon (MZ) is produced by Fusarium and introduces the lactone of rezorcylic acid possessed the pronounced estrogenic effect on animal organism. Taking into consideration the influence of estrogens administration on catecholamine turnover rate of the brain we have investigated the influence of MZ on dopamine‐β‐monooxigenase (DBM) (EC 1. 14. 17. 1), which catalyzes the final reaction of noradrenalin formation from dopamine. Using lineweares‐Burk plots Km for tyramine and ascorbate and Vmax of DBM catalyzing reaction and high speed centrifugation we demonstrate the complex formation between MZ and DBM. The mechanisms of the proposed effect of MZ on DBM and the details of toxic action of MZ on biological systems of the organism are conditioned by abnormalities in different systems of metabolism especially phospholipids (PL), so far as it is known already that membrane‐bound DBM is phospholipid‐dependent enzymatic system. The activating effect of MZ on adrenaline formation is accompanied by the elevation of Ca ions concentration, which plays an important role in intensification of Ca2+‐dependent phospholipase A2 activity, and accelerates the deacylation of the neutral PL, especially phosphatidylcholines with the formation of high concentrations of lysophosphatidylcholines, unesterified fatty acids and products of their peroxidation. These compounds have a pronounced membranotoxic and membranolytic action, which are one of the main pathogenetic mechanisms at MZ poisoning. Using of combined treatment by sodium thiosulfate and Ca2+‐ds‐RNA leads to the normalization of MZ poisoning phenomena.
Poster Sessions CP12: Neurotrauma & Injury. Characterization of cathepsin B expression following contusion spinal cord injury (SCI)
Authors: Ellis R. C. , Earnhardt J. N. , Nick H. S. , Hayes R. L. , Anderson D. K.
Published: July 2010

An increase in protease activity is a hallmark event of the secondary injury cascade following contusion SCI. Elevated levels of protease activity result in the degradation of cytoskeletal components and myelin proteins essential for cellular function and survival. We have shown that a member of the cathepsin protease family is affected by SCI. The excessive release and activity of cathepsin B, a fairly ubiquitous lysosomal cysteine protease, has been implicated in several pathologies including tumor metastasis and progression, arthritis and Alzheimer's disease. Thus, our goal was to characterize any SCI‐induced changes in cathepsin B expression. Following a T12 laminectomy and a moderate contusion (NYU device), the gene and protein profiles of cathepsin B in rats were examined using real‐time PCR and immunoblots, respectively. Both the contusion injured animals and the time‐matched sham controls exhibited elevated pro‐enzyme protein levels (37 kDa form) at the lesion site, with significant differences between the two groups at 48 h, 72 h and 7 days post‐SCI. Furthermore, there was a surge in the active species of the protein with significant differences at 72 h and 7 days post‐SCI for the 30 kDa form and at 48 h. and 7 days for the 25 kDa form. Real‐time PCR revealed increases in cathepsin B mRNA levels following contusion SCI as early as 6 h postinjury. These data indicate that SCI causes an up‐regulation of cathepsin gene expression and protein levels, and suggest that this protease may be involved in the secondary injury cascade perhaps for as long as 1 week postinjury.
Estrogen as a neuroprotective agent in rat spinal cord injury
Authors: Banik N. L. , Sribnick E. A. , Wingrave J. M. , Matzelle D. D. , Wilford G. G. , Ray S. K.
Published: July 2010

Spinal cord injury (SCI) is a neurological problem affecting approximately 11 000 Americans each year. Several treatment agents have been proposed; however, only methylprednisolone has limited efficacy. Estrogen is a multiactive neuroprotectant with antioxidant and anti‐inflammatory properties and attenuates calcium (Ca2+) influx following neuronal injury. To examine the neuroprotective effects of estrogen in SCI, we induced SCI (40 g/cm injury) in rats. Treatment groups were sham (laminectomy only), SCI plus vehicle, and SCI plus estrogen. Injured rats were treated with either 4 mg/kg 17 β‐estradiol (estrogen group) or dimethylsulfoxide (vehicle group) at 15 min and 24 h following injury. All rats were killed at 48 h to analyze SCI segments for calpain content and Bax/Bcl‐2 ratio by Western blotting. Tissue was also examined using calcium green‐2 to measure intracellular [Ca2+], JC‐1 to measure mitochondrial membrane potential, and double immunofluorescence for macrophages and calpain. Calpain content in the lesion penumbra, adjacent to the injury, was higher in vehicle than sham and this increase was attenuated in estrogen treated rats. In the lesion penumbra, the Bax/Bcl‐2 ratio was increased in vehicle rats as compared to sham. This increase was attenuated in estrogen treated rats. Estrogen treated rats had less Ca2+ influx, less inflammatory cell infiltration, and increased maintenance of mitochondrial membrane potential compared to vehicle treated rats. Our preliminary data suggest that estrogen may be effective in decreasing Ca2+ influx, inflammatory cell infiltration, and Bax/Bcl‐2 ratio following SCI.

Acknowledgements:  Supported in part by grants from NIH‐NINDS and South Carolina Electric and Gas.
Effect of neuronal NOS inhibitor, 7‐nitro‐indazole, on cytosolic caspase 9 activity in vitro
Authors: Mishra O. P.
Published: July 2010

Previous studies have shown that nitric oxide synthase (NOS) inhibitors prevent or reduce the hypoxia‐induced neuronal injury. The present study tests the hypothesis that neuronal NOS inhibitor, 7‐nitro‐indazole (7‐NI) will inhibit the cysteine protease, caspase 9, the initiator of programmed cell death. The effect of 7 NI on caspase 9 activity was determined in the cytosolic fraction of the cerebral cortex of normoxic (Nx, n = 4) and hypoxic (Hx, n = 4) newborn piglets. The animals were exposed to either 21% (Nx) or 7% (Hx) oxygen for 60 min. Cerebral tissue hypoxia was documented by the ATP and phosphocreatine levels. Cytosols were prepared and caspase 9 activity determined using a specific fluorogenic substrate (‐Ac‐Leu‐Glu‐His‐Asp‐AFC) for caspase 9 and monitored at 400 nm excitation and 505 nm emission wavelength. Caspase 9 activity in Nx group was 629 ± 49 as compared to 960 ± 105 pmoles AFC/min/mg protein (p < 0.002, 50.2% increase) in the Hx group. In presence of 10, 50 and 100 μm 7‐NI, caspase 9 activity decreased to 459 ± 41, 252 ± 36 and 256 ± 29 pmoles/min/mg protein, respectively, in Nx group. In Hx group, the activity decreased to 578 ± 49, 402 ± 37, 393 ± 35 pmoles/min/mg protein, respectively. The data demonstrate that 7‐NI inhibits caspase 9 in both the Nx and Hx groups. It is concluded that 7‐NI is an inhibitor of caspase 9. It is proposed that 7 NI administration will be neuroprotective due to its properties as inhibitor of nNOS as well as of caspase 9 and prevent the hypoxia‐induced neuronal death.

Acknowledgements:  Supported by NIH‐HD‐38079.
Superoxide activates synaptic particulate nitric oxide synthase: implications for superoxide signalling and neurotoxicity
Authors: Foley T. D. , McPherson D. B. , Kilker R. P.
Published: July 2010

Mounting evidence supports the view that nitric oxide may protect against superoxide‐induced oxidative damage by mechanisms involving nitrosylation, direct scavenging of oxygen radicals including superoxide, and up‐regulation of antioxidant enzymes. However, the ability of neurons to respond to superoxide by increasing nitric oxide synthesis is unknown. We report here that superoxide, added as potassium superoxide (KO2), produced a rapid activation (up to 12‐fold) of nitric oxide synthase (NOS), measured by the conversion of [3H]arginine to [3H]citrulline, and nitrite production in a synaptic particulate fraction from rat brain cerebral cortex. Stimulation of NOS by KO2 required the presence of ATP, which by itself inhibited basal NOS activity. Decayed KO2 or authentic hydrogen peroxide did not increase NOS activity and, importantly, activation of NOS by KO2 was blocked by addition of superoxide dismutase. In addition, stimulation of NOS activity by KO2 was abolished by the NOS inhibitor N‐monomethyl‐l‐arginine and EGTA but only partially reduced by the neuronal NOS inhibitor 7‐nitroindazole. These findings are the first to demonstrate directly that superoxide activates a synaptic NOS isoform. The nature of the particulate fraction, the experimental conditions, and the low sensitivity to 7‐nitroindazole strongly suggest that superoxide activated mitochondrial NOS (mtNOS). We propose that activation of the particulate NOS by superoxide reflects a novel mitochondrial response that may contribute to neuronal oxidant defenses although chronic activation of NOS under conditions of superoxide generation may produce toxic quantities of peroxynitrite and other reactive nitrogen oxides.
Induction of tissue transglutaminase in response to ischemic brain injury
Authors: Wang D. S. , Tolentino P. J. , Fan T. , Pike B. R. , Wang K. K. W. , Day A. L. , Hayes R. L.
Published: July 2010

Tissue transglutaminase (tTG) is a member of the transglutaminase family that catalyzes the post‐translational modification of proteins by inserting isopeptide bond within or between polypeptide chains in a Ca++ dependent manner. Although tTG is known to be involved in cell growth, apoptosis and neurodegeneration, the role of tTG in traumatic and ischemic brain injury (TBI and IBI) is largely unknown. Previously, we showed that tTG was up‐regulated at mRNA and protein levels after TBI in rats. Although TBI and IBI can result in some common pathological sequelae, there are many important differences. Thus, we hypothesized that tTG message and protein is up‐regulated after transient middle cerebral artery occlusion (MCAO), and that the magnitude and temporal expression differ from TBI. tTG protein and mRNA of ipsi‐cortex and hippocampus from rats of 0, 1, 3, 5 and 7 days after MCAO were quantified by Western blot and real time PCR with tissues from similar contra‐lateral areas as controls. Results showed that tTG mRNA in ipsi‐cortex increased steadily from 1 to 5 days (1.5–10 fold of control level) after MCAO and dropped to control level by 7 days. In ipsi‐hippocampus, tTG mRNA increased ∼2 fold of control at 1 day after MCAO and gradually returned to control level by 7 days. tTG protein levels for both ipsi‐cortex and ipsi‐hippocampus increased steadily during the experiment with a more significant increase in cortex (∼5.5 fold in ipsi‐cortex vs. 2 fold in ipsi‐hippocampus). Levels of tTG mRNA and protein of control tissues were unchanged. These results indicate that tTG after MCAO in cortex and hippocampus is up‐regulated in a similar pattern to those in TBI brains. Studies of functional significance of tTG after ischemia and TBI are underway.
A novel oxidative stress dependent apoptotic pathway in pesticide‐induced dopaminergic degeneration in PD models
Authors: Kanthasamy A. , Kitazawa M. , Kaul S. , Anantharam S. V. , Kanthasamy A. G.
Published: July 2010

Emerging epidemiological and case control studies suggest that environmental factors especially pesticides are dominant risk factors in the etiology of idiopathic Parkinson's disease (PD). In the present study, we characterized oxidative stress dependent cellular events by exposing rat mesencephalic dopaminergic cell line (N27) and PC 12 cells to dieldrin (a potential environmental risk factor for development of PD). Exposure of dieldrin (30–200 μm) resulted in a rapid increase in generation of reactive oxygen species within 30 min and was followed by release of mitochondrial cytochrome‐c into cytoplasm. Cytochrome‐c then activated a proapoptotic cysteine protease, caspase‐3 in a time‐ and dose‐dependent manner. Interestingly, we also found that dieldrin exposure induces a time‐ and dose‐dependent proteolytic cleavage of native protein kinase Cδ (PKCδ, 74 kDa) to yield 41‐kDa catalytically active and 38‐kDa regulatory fragments. Pretreatment with caspase inhibitors blocked dieldrin‐induced proteolytic cleavage of PKCδ, indicating that cleavage is mediated by caspase‐3. Attenuation of ROS generation, caspase‐3 activation or overexpression of mutant PKCδK376R (dominant negative) almost completely suppressed dieldrin‐induced DNA fragmentation. We also found that dieldrin exposure activates the redox sensitive transcription factor NF‐kB through PKCδ dependent manner. Together, these data demonstrate that caspase‐3 dependent proteolytic activation of PKCδ plays a key role in oxidative stress‐mediated apoptosis in dopaminergic cells following exposure to an environmental neurotoxic agent.

Acknowledgements:  Supported in part by ES 10586.
Proteins with pathogenic polyglutamine expansion inhibit fast axonal transport
Authors: Szebenyi G. , Morfini G. , Young M. , Sheridan S. , McPhaul M. J. , Brady S. T.
Published: July 2010

Several neurological diseases, including Huntington's and spinobulbar muscular atrophy (SBMA), are the result of pathogenic activities of expanded polyglutamine (polyQ) tracts. The reasons for the late onset of the disease and for the special susceptibility of certain populations of neurons remain unknown, as are the mechanisms of polyQ induced neurodegeneration. Based on the observation that Huntingtin can be associated with vesicles in transport, we examined the effects of polyQ on rates of axonal transport and on cellular functions that are strongly dependent on prompt delivery of cellular components. Pathogenic forms of Huntington inhibited both anterograde and retrograde axonal transport in the squid axoplasm, while normal Huntington protein had no effect. To test the activity of polyQ on cellular functions in intact neurons, cell lines stably expressing either wild type or SBMA pathogenic forms of the androgen receptor were examined. PolyQ expansion had no effect on rates of proliferation, but negatively affected process elongation. The same polyQ constructs that most severely affected the elaboration of cellular processes in SH‐SY5Y cells were the ones most effective at inhibiting axonal transport in the squid. Taken together, our data are consistent with polyQ proteins altering the regulation of microtubule dependent transport processes.
Oral Presentations OP02: Myelin and Diseases of Myelin. NPP2, A matricellular protein expressed during myelination, actively modulates oligodendrocyte adhesion
Authors: Fox M. A. , Bowlin G. , Fuss B.
Published: July 2010

NPP2, also known as phosphodiesterase‐I alpha/autotaxin, is a type‐II membrane protein that belongs to the nucleotide pyrophosphatase/phosphodiesterase family (NPP). We have recently demonstrated that NPP2 is expressed and released by differentiating oligodendrocytes during the critical stages of CNS myelination. The structural domains of this secreted macromolecule suggest a functional role in the regulation of oligodendrocyte adhesion. Here, we present data that demonstrates that NPP2 interferes with the ability of oligodendroglial cells to adhere to known CNS adhesion molecules present during the onset of myelination, such as fibronection, vitronectin, and merosin (laminin2). Responses to NPP2 appear to be regulated by a different mechanism depending on the developmental stage of the oligodendrocyte. Although the exact mechanisms for NPP2 mediated counter‐adhesion are unknown, our studies have implicated that an active signalling mechanism involving heterotrimeric G proteins is responsible for adhesion modulation. These studies clearly define a role of NPP2 as a matricellular protein modulating oligodendrocyte adhesion and suggest that NPP2 function may represent the first step of oligodendrocyte remodelling when differentiating oligodendrocytes are actively involved in the formation of the myelin sheath.
Astrocyte‐specific neurofibromatosis 1 gene deletion is insufficient for astrocytoma formation in mice
Authors: Gutmann D. H. , Bajenaru M. L. , Hedrick N. , Wu Y. , Fink L. , Zhu Y. , Parada L.
Published: July 2010

Children with neurofibromatosis 1 (NF1) develop low‐grade astrocytomas. To study the role of the NF1 gene product, neurofibromin, in astrocyte growth and NF1‐associated astrocytoma formation, we generated astrocyte‐specific NF1 conditional knockout mice (NF1GFAP CKO) using the Cre/LoxP recombination system. Astrocyte‐specific Nf1 conditional knockout mice (NF1 CKO mice) were generated by crossing GFAP‐Cre‐IRES‐LacZ mice with LoxP targeted NF1 mice. NF1GFAP CKO mice were monitored for astrocytoma formation and neurological abnormalities for 20 months and no tumors were detected. Increased numbers of astrocytes were found in several regions of the brains of NF1GFAPCKO mice. Cultured astrocytes derived from NF1GFAP CKO mice show a 1.8‐fold increase in proliferation, but do not grow in soft agar or serum free media. In addition, embryonic day 13 astrocyte cultures from conventional NF1 knockout mice were generated. We also deleted the NF1 gene in vitro in postnatal day 2 NF1 CKO astrocytes with Cre adenovirus. Embryonic and postnatal NF1−/− astrocytes show a growth increase in vitro, but they do not grow in soft agar or serum free media, and do not form tumors when injected in the brains of immunocompromised nude (nu/nu) mice. In addition, NF1‐null cultured astrocytes were injected into wild type and NF1+/−; mouse brains to determine the effect of the brain genetic environment of astrocytoma formation. Collectively, our results suggest that loss of neurofibromin might not be sufficient for astrocytoma formation in the mouse and that other genetic or environmental factors may influence tumorigenesis.
Symposium 3: Function and Pathogenesis in Parkinson’s Disease. Synuclein: the third amyloid in Alzheimer's disease
Authors: Trojanowski J. Q.
Published: July 2010

A dramatic paradigm shift in understanding Parkinson's disease (PD) has emerged with implications for Alzheimer's disease (AD) because: (1) Mutations in the alpha‐synuclein (AS) gene cause familial PD, (2) Antibodies to AS detect Lewy bodies (LBs) and dystrophic Lewy neurites in PD, dementia with LBs (DLB), sporadic AD and the LB variant of AD (LBVAD), (3) Insoluble AS filaments are recovered from DLB brains and purified LBs, (4) Recombinant AS assembles into LB‐like filaments and residues 71–82 are essential for filament assembly, (5) AS transgenic mice and flies develop a PD‐like phenotype, (6) Cortical LBs detected with antibodies to AS correlate with dementia in PD, DLB and LBVAD, (7) Antibodies to AS detect LBs in 50% of familial AD, sporadic AD and Down's syndrome brains, (8) AS forms glial cytoplasmic inclusions (GCIs) in multiple system atrophy, (9) Epitopes throughout AS in LBs and GCIs, (10) Filamentous AS aggregates in LBs, GCIs and related lesions contain nitrated tyrosines, (11) Cells transfected with AS and treated with nitric oxide generators develop LB‐like AS inclusions, (12) Bigenic mice overexpressing mutant human APP and AS show an augmentation in AS inclusions. Thus, neurodegenerative diseases characterized by AS pathologies are synucleinopathies, and the filamentous AS lesions in these disorders may result in part from oxidative/nitrative damage to AS. Abnormal interactions of brain proteins may underlie synucleinopathies and other neurodegenerative disorders. Acknowledgements:

Supported by NIA/NIH and Alzheimer's Association.
Toward a normal function for the synuclein proteins
Authors: George J. M.
Published: July 2010

α‐Synuclein has received much attention in recent years because of its association with neurodegenerative disease. Mutations in the α‐synuclein gene are linked to some rare cases of early onset Parkinson's disease, and the α‐synuclein protein itself is abundant in intracellular inclusions typical of sporadic and familial Parkinson's disease, dementia with Lewy bodies, Down's syndrome, and multiple system atrophy. Our lab has focused on the biophysical attributes of α‐synuclein as a means to elucidate its normal cellular function. α‐Synuclein and its relatives, β‐ and γ‐synuclein, are abundant proteins of the nervous system, and their primary sequences are extremely well conserved, suggesting that specific molecular interactions may constrain their evolution. One candidate interaction is that which occurs between synucleins and phospholipid membranes. Each of the proteins possesses an apolipoprotein‐like N‐terminal domain of ∼ 100 amino acid which adopts a random coil conformation in solution, but folds into an amphipathic α‐helix upon association with phospholipid bilayers. We find that a similar conformational change can be driven by fatty acid micelles, and that association with α‐synuclein protects the fatty acids against oxidative damage. Drawing on these observations and on the published literature, this presentation will attempt to formulate a general hypothesis for synuclein function which accounts for the observed similarities between the synucleins and the exchangeable apolipoproteins.
Environmental risk factors and Parkinson's disease: the role of interactions between neurotoxicants and alpha‐synuclein
Authors: Di Monte D. A.
Published: July 2010

Several lines of clinical and experimental evidence suggest that environmental risk factors are likely to play a role in the pathogenesis of Parkinson's disease (PD). The two pathological hallmarks of PD are selective nigrostriatal degeneration and formation of alpha‐synuclein‐containing intraneuronal inclusions (called Lewy bodies). The goal of these studies was to test the hypothesis that environmental toxicants may be able to reproduce these parkinsonian features in experimental animal models. A mouse model recently developed in our laboratory was characterized by the selective loss of dopaminergic neurons in the substantia nigra pars compacta after repeated exposures to the herbicide paraquat. As with human parkinsonism, paraquat‐induced neurodegeneration in mice was accompanied by gliosis. Another intriguing feature of this model was an up‐regulation of alpha‐synuclein that consistently followed each paraquat administration. Although organized structures resembling Lewy bodies were not observed, signs of protein aggregation were evident in the form of intraneuronal accumulation of thioflavine S‐positive deposits. Co‐localization of alpha‐synuclein within these deposits provided further evidence that paraquat injections triggered alpha‐synuclein aggregation in the mouse brain. Taken together, these results suggest that up‐regulation of alpha‐synuclein may be part of the response of neurons to toxic insults. They also provide important evidence in support of a relationship between environmental exposures, alpha‐synuclein–toxicant interactions and the development of parkinsonian pathology.
GM1 ganglioside in the treatment of Parkinson's disease: long‐term clinical experience
Authors: Schneider J. S.
Published: July 2010

The monosialoganglioside GM1 is a glycosphingolipid with putative neuroprotective and neurorestorative properties. GM1 has been shown to at least partially reverse neurochemical, neuropathological and behavioral deficits in various lesion models including experimental models of parkinsonism in rodents, felines and nonhuman primates. Based on these experimental findings, we conducted a randomized double blind placebo controlled trial of GM1 in Parkinson's disease (PD). The results of this study showed significant improvements in Unified Parkinson's Disease Rating Scale (UPDRS) motor ratings as well as in other indices of motor function and activities of daily living assessments in PD patients that received 16 weeks of GM1 treatment (Schneider et al. Neurology 1998; 50: 1630–1636). At the conclusion of that study, patients were allowed to receive GM1 in an open extension trial. Fifteen patients chose to receive GM1 and remained in the open trial for approximately 4 years; 11 patients completed the full 5 years open trial. Chronic daily use of GM1 ganglioside by Parkinson's disease patients, over a period of 4–5 years is safe and has led to significant and sustained improvements in motor functioning, in most cases. In addition, we have observed a lower than expected frequency of motor complications of levodopa therapy (i.e. levodopa‐induced dyskinesias) in this patient group. Although the precise mechanisms of action of GM1 ganglioside are still unclear, animal and in vitro studies suggest that GM1 may interfere with apoptotic and to a lesser extent, necrotic cell death and stimulate cell survival and perhaps fiber/terminal sprouting through a variety of cellular/molecular processes.
Sphingolipids – roads still to be taken
Authors: Schengrund C. L.
Published: July 2010

Despite advances already made towards understanding the biological role(s) of sphingolipids (SLs) there are still a number of questions that need to be addressed.

For example, while glycosphingolipids (GSLs) are known to be a component of lipid rafts, where they can provide multivalent ligands for carbohydrate–carbohydrate interactions, their actual function in the rafts is unknown. When gangliosides are added to cells and associate with their plasma membranes are they found in rafts or do they distribute in a relatively even fashion across the entire cell membrane? What happens to raft formation in cells made GSL minus? How do subtle changes in a cell's total SL composition, or in the ceramide or polar portion of a specific SL, affect its behavior? What are the specific functions of the different carbohydrate moieties associated with GSLs? In addition to their known role as ligands for certain pathogens, do they function as specific ligands for endogenous molecules? How does substitution of a deoxy for a hydroxyl group on a sugar molecule in a GSL affect their function? Does replacement of a ganglioside sialic acid with a different, negatively charged, component (e.g. sulfate) alter its behavior? How are GSLs transported from the site of injection to their putative site of action when used in animal or human trials to enhance neuronal recovery? Do they associate with albumin? Are they cleared from the circulation by specific galactose binding proteins? Can their clearance by the liver be reduced by modifying their carbohydrate portion? Each of the above questions describes a road currently in need of more exploration.
Agonists specific for the transcription factor PPARdelta accelerate differentiation of oligodendrocytes
Authors: Skoff R. P. , Bessert D. , Saluja I. , Granneman J.
Published: July 2010

Peroxisome proliferator activated receptors (PPARs) are transcription factors belonging to the nuclear hormone receptor superfamily that regulate key genes involved in lipid metabolism. PPARδ is ubiquitously expressed at low levels in many tissues and its function has remained elusive. However, we have shown that PPARδ is abundantly expressed in oligodendrocytes (Ols), suggesting this receptor plays a critical role in oligodendrocyte differentiation (Granneman et al. 1998 J. Neurosci. Res51, 563). We first investigated the effects of PPAR agonists on proliferation and differentiation of Ols in tissue culture. Primary glial and enriched Ol cultures were treated with ligands that specifically activate PPARγ and PPARδ (Berger et al. 1999 J. Biol. Chem. 274, 6717). PPARδ but not PPARγ agonists increased the size of OL membrane sheets within 24 h of application. The increase in membrane sheet size was mirrored by increases in MBP and PLP mRNA's. In enriched Ol cultures, the number of Ols was increased 70% with the PPARδ agonist but not the PPARγ agonist (Saluja et al. 2001 Glia33, 191). In vivo injections of PPARδ agonist into P2 and P3 mice show an increase of total macroglia in the ventral and dorsal funiculi of the spinal cord of 20–40% compared to controls. Preliminary observations suggest the Ols in agonist treated cultures are larger and more densely stained than controls. Our results show for the first time that a specific ligand for a transcription factor is capable of activating the program of Ol differentiation.

Acknowledgements: Supported by NMSS.
Colloquium 9: Structural and Molecular Dissection of the Node of Ranvier. Sodium channel β subunits: multifunctional molecules at the node of Ranvier
Authors: Isom L. L.
Published: July 2010

Voltage‐gated sodium channels are unique in that they combine action potential conduction with cell adhesion. Mammalian sodium channels are heterotrimers, composed of a central, pore‐forming α subunit and two auxiliary β subunits. The α subunits are members of a large gene family containing the voltage‐gated sodium, potassium, and calcium channels. Sodium channel α subunits form a gene subfamily with at least 11 members. Mutations in sodium channel α subunit genes have been linked to paroxysmal disorders such as epilepsy, long QT syndrome (LQT), and hyperkalemic periodic paralysis in humans, and motor endplate disease and cerebellar ataxia in mice. Three genes encode the sodium channel β subunits with at least one alternative splice product. Unlike the pore‐forming α subunits, the sodium channel β subunits are not structurally related to β subunits of calcium and potassium channels. Sodium channel β subunits are multifunctional. They modulate channel gating and regulate the level of channel expression at the plasma membrane. We have shown that β subunits also function as cell adhesion molecules (CAMs) in terms of interaction with extracellular matrix molecules, regulation of cell migration, cellular aggregation, and interaction with the cytoskeleton. A mutation in SCN1B has been shown to cause GEFS + 1 epilepsy in human families. We propose that the sodium channel signalling complex at nodes of Ranvier involves β subunits as channel modulators as well as CAMs, other CAMs such as neurofascin and contactin, RPTPβ, and extracellular matrix molecules such as tenascin.
The myelin membrane influences the organization of molecules on the axonal surface
Authors: Pedraza L. , Huang J. K. , Svenningsen A. F. , Colman D. R.
Published: July 2010

The paranodal axoglial junction, a highly unique and specialized vertebrate structure, is the most likely domain at which axon‐glial communication takes place. The molecular components of this structure are largely unknown. The first identified protein that was localized specifically to the axoglial junction was caspr/paranodin, on the axonal side. The second was neurofascin 155 (NF155), a member of the immunoglobulin superfamily of cell adhesion molecules, on the glial cell side. In the mature myelinating glial cell, NF155 clusters in apposition to axonal caspr at the paranodal junction. These proteins are believed to be important in the establishment and stabilization of the axo‐glial contact and in the restriction of sodium channels to the nodal region. We have studied the spatial and temporal organization of the myelin membrane and the way myelin influences the organization of the axon, in particular of membrane proteins at the nodal and paranodal level. We observed that there is a focal increase of caspr along axonal segments that are in contact with oligodendrocyte membrane expansions. This up‐regulation or redistribution of caspr on the axonal side is mirrored by an increased of NF155 on the oligodendrocyte side. Therefore, caspr and NF155 codistribute from the time of initial contact, until they reach their final distribution at the paranodal junction. This implicates NF155 and caspr in axoglial recognition and early communication, as well as in the establishment of membrane compartments or domains, where molecules are restricted from freely diffusing in the lateral plane of the axonal membrane.
Putative
Arabidopsis
THO/TREX mRNA export complex is involved in transgene and endogenous siRNA biosynthesis
Authors: Yelina Nataliya E. , Smith Lisa M. , Jones Alexandra M. E. , Patel Kanu , Kelly Krystyna A. , Baulcombe David C.
Published: July 2010

RNA silencing in plants and some animals has a non–cell-autonomous effect due to an RNA signal that moves between cells or organs. To identify unique factors involved in this process, we analyzed a group of
Arabidopsis
mutants with defective spread of RNA silencing from a transgene expressed specifically in the phloem. These mutants accumulated reduced amounts of small interfering (si)RNA from the transgene locus and from endogenous loci
TAS1
,
TAS2
, and an inverted repeat locus
IR71
. The defect in
TAS1
and
TAS2
siRNA biogenesis is in the processing of a long siRNA precursor. We mapped the mutations to a gene encoding the
Arabidopsis
homolog of a protein, TEX1, which is involved in intracellular transport of RNA in animals. TEX1 is a component of the THO/TREX complex, and we show that the
Arabidopsis
TEX1 interacts with other predicted components of a THO/TREX complex. Correspondingly, we found at least two other components of the
Arabidopsis
THO core complex that are involved in RNA silencing. To reconcile the effect of these mutations on transgene and endogenous gene siRNA, we propose a mechanism in which THO/TREX processes or transports a long RNA molecule so that it can be a template for secondary siRNA production.
Regulation of NF-κB activity and inducible nitric oxide synthase by regulatory particle non-ATPase subunit 13 (Rpn13)
Authors: Mazumdar Tuhina , Gorgun F. Murat , Sha Youbao , Tyryshkin Alexey , Zeng Shenyan , Hartmann-Petersen Rasmus , Jørgensen Jakob Ploug , Hendil Klavs B. , Eissa N. Tony
Published: July 2010

Human Rpn13, also known as adhesion regulating molecule 1 (ADRM1), was recently identified as a novel 19S proteasome cap-associated protein, which recruits the deubiquitinating enzyme UCH37 to the 26S proteasome. Knockdown of Rpn13 by siRNA does not lead to global accumulation of ubiquitinated cellular proteins or changes in proteasome expression, suggesting that Rpn13 must have a specialized role in proteasome function. Thus, Rpn13 participation in protein degradation, by recruiting UCH37, is rather selective to specific proteins whose degradation critically depends on UCH37 deubiquitination activity. The specific substrates for the Rpn13/UCH37 complex have not been determined. Because of a previous discovery of an interaction between Rpn13 and inducible nitric oxide synthase (iNOS), we hypothesized that iNOS is one of the substrates for the Rpn13/UCH37 complex. In this study, we show that Rpn13 is involved in iNOS degradation and is required for iNOS interaction with the deubiquitination protein UCH37. Furthermore, we discovered that IκB-α, a protein whose proteasomal degradation activates the transcription factor NF-κB, is also a substrate for the Rpn13/UCH37 complex. Thus, this study defines two substrates, with important roles in inflammation and host defense for the Rpn13/UCH37 pathway.
Core Components for Infection Prevention and Control Programs: A World Health Organization Network Report
Authors: Seto Wing Hong , Otaíza Fernando , Pessoa-Silva Carmen L. ,   World Health Organization Infection Prevention and Control Network
Published: July 2010

Under the leadership of the World Health Organization (WHO), the core components necessary for national and local infection prevention and control programs are identified. These components were determined by a network of international experts who are representatives from WHO regional offices and relevant WHO programs. The respective roles of the national authorities and the local healthcare facilities are delineated.
The Hypersensitive Induced Reaction and Leucine-Rich Repeat Proteins Regulate Plant Cell Death Associated with Disease and Plant Immunity
Authors: Choi Hyong Woo , Kim Young Jin , Hwang Byung Kook
Published: July 2010

Pathogen-induced programmed cell death (PCD) is intimately linked with disease resistance and susceptibility. However, the molecular components regulating PCD, including hypersensitive and susceptible cell death, are largely unknown in plants. In this study, we show that pathogen-induced Capsicum annuum hypersensitive induced reaction 1 (CaHIR1) and leucine-rich repeat 1 (CaLRR1) function as distinct plant PCD regulators in pepper plants during Xanthomonas campestris pv. vesicatoria infection. Confocal microscopy and protein gel blot analyses revealed that CaLRR1 and CaHIR1 localize to the extracellular matrix and plasma membrane (PM), respectively. Bimolecular fluorescent complementation and coimmunoprecipitation assays showed that the extracellular CaLRR1 specifically binds to the PM-located CaHIR1 in pepper leaves. Overexpression of CaHIR1 triggered pathogen-independent cell death in pepper and Nicotiana benthamiana plants but not in yeast cells. Virus-induced gene silencing (VIGS) of CaLRR1 and CaHIR1 distinctly strengthened and compromised hypersensitive and susceptible cell death in pepper plants, respectively. Endogenous salicylic acid levels and pathogenesis-related gene transcripts were elevated in CaHIR1-silenced plants. VIGS of NbLRR1 and NbHIR1, the N. benthamiana orthologs of CaLRR1 and CaHIR1, regulated Bax- and avrPto-/Pto-induced PCD. Taken together, these results suggest that leucine-rich repeat and hypersensitive induced reaction proteins may act as cell-death regulators associated with plant immunity and disease.
A Putative RNA-Binding Protein Positively Regulates Salicylic Acid–Mediated Immunity in Arabidopsis
Authors: Qi Yiping , Tsuda Kenichi , Joe Anna , Sato Masanao , Nguyen Le V. , Glazebrook Jane , Alfano James R. , Cohen Jerry D. , Katagiri Fumiaki
Published: July 2010

RNA-binding proteins (RBP) can control gene expression at both transcriptional and post-transcriptional levels. Plants respond to pathogen infection with rapid reprogramming of gene expression. However, little is known about how plant RBP function in plant immunity. Here, we describe the involvement of an RBP, Arabidopsis thaliana RNA-binding protein-defense related 1 (AtRBP-DR1; At4g03110), in resistance to the pathogen Pseudomonas syringae pv. tomato DC3000. AtRBP-DR1 loss-of-function mutants showed enhanced susceptibility to P. syringae pv. tomato DC3000. Overexpression of AtRBP-DR1 led to enhanced resistance to P. syringae pv. tomato DC3000 strains and dwarfism. The hypersensitive response triggered by P. syringae pv. tomato DC3000 avrRpt2 was compromised in the Atrbp-dr1 mutant and enhanced in the AtRBP-DR1 overexpression line at early time points. AtRBP-DR1 overexpression lines showed higher mRNA levels of SID2 and PR1, which are salicylic acid (SA) inducible, as well as spontaneous cell death in mature leaves. Consistent with these observations, the SA level was low in the Atrbp-dr1 mutant but high in the overexpression line. The SA-related phenotype in the overexpression line was fully dependent on SID2. Thus, AtRBP-DR1 is a positive regulator of SA-mediated immunity, possibly acting on SA signaling-related genes at a post-transcriptional level.
Lack of Galactose or Galacturonic Acid in Bradyrhizobium japonicum USDA 110 Exopolysaccharide Leads to Different Symbiotic Responses in Soybean
Authors: Quelas Juan Ignacio , Mongiardini Elías J. , Casabuono Adriana , López-García Silvina L. , Althabegoiti M. Julia , Covelli Julieta M. , Pérez-Giménez Julieta , Couto Alicia , Lodeiro Aníbal R.
Published: July 2010

Exopolysaccharide (EPS) and lipopolysaccharide (LPS) from Bradyrhizobium japonicum are important for infection and nodulation of soybean (Glycine max), although their roles are not completely understood. To better understand this, we constructed mutants in B. japonicum USDA 110 impaired in galactose or galacturonic acid incorporation into the EPS without affecting the LPS. The derivative LP 3010 had a deletion of lspL-ugdH and produced EPS without galacturonic acid whereas LP 3013, with an insertion in exoB, produced EPS without galactose. In addition, the strain LP 3017, with both mutations, had EPS devoid of both galactosides. The missing galactosides were not replaced by other sugars. The defects in EPS had different consequences. LP 3010 formed biofilms and nodulated but was defective in competitiveness for nodulation; and, inside nodules, the peribacteroid membranes tended to fuse, leading to the merging of symbiosomes. Meanwhile, LP 3013 and LP 3017 were unable to form biofilms and produced empty pseudonodules but exoB suppressor mutants were obtained when LP 3013 plant inoculation was supplemented with wild-type EPS. Similar phenotypes were observed with all these mutants in G. soja. Therefore, the lack of each galactoside in the EPS has a different functional effect on the B. japonicum–soybean symbiosis.
Antibiotic‐associated Bloody Diarrhea in Infants: Clinical, Endoscopic, and Histopathologic Profiles
Authors: Barakat Maha , El‐Kady Zeinab , Mostafa Mohamed , Ibrahim Naglaa , Ghazaly Hamdy
Published: July 2010

Objective:

Antibiotic‐associated diarrhea constitutes 1 of the most frequent side effects of antimicrobial therapy with widely varying clinical presentations; however, little is known about its antibiotic‐associated bloody diarrhea (AABD) form, particularly in very young children. The aim of this study was to describe the clinical, endoscopic, and histopathologic profiles of community‐acquired AABD in infants.

Patients and Methods:

The study included 23 infants referred with bloody diarrhea that developed a few days after receiving antibiotics on an outpatient basis for watery diarrhea (18), respiratory tract infections (4), or urinary tract infection (1). Detailed clinical assessment, videosigmoidoscopy, and histopathologic examination of endoscopic biopsies were performed for all.

Results:

Clinically, on presentation, bloody diarrhea was acute in all except 1 patient with a prolonged course (for 25 days) and stopped in all 2 to 6 days after discontinuation of antibiotics. Fever and/or leukocytosis were present only in 8 (34.8%). Sigmoidoscopy revealed varying types of erythema (patchy, ring, diffuse) and ulcers (aphthoid, diffuse) in 18 and pseudomembranes in 5. Histopathologically, only 3 showed the characteristic mushroom‐like pseudomembranes, whereas all of the other infants had nonspecific colitis.

Conclusions:

Community‐acquired AABD is not uncommon in infants presenting with acute or chronic forms even without fever or leukocytosis. When suspected, discontinuation of antibiotics is a good policy if facilities for bacterial culture with cytotoxin assays are limited. The characteristic endoscopic or histopathologic pseudomembranes are encountered only in a small percentage (26%). Rational use of antibiotics should be adhered to particularly in cases of watery diarrhea that is mostly of viral origin.
Increased Number of Regulatory T Cells in Children With Eosinophilic Esophagitis
Authors: Fuentebella Judy , Patel Anup , Nguyen Tammie , Sanjanwala Bharati , Berquist William , Kerner John A , Bass Dorsey , Cox Kenneth , Hurwitz Melissa , Huang Jennifer , Nguyen Christine , Quiros J Antonio , Nadeau Kari
Published: July 2010

Objectives:

There are limited data on the role of regulatory T cells (Treg) in the disease pathology of eosinophilic esophagitis (EoE). We tested the differences in Treg in subjects with EoE compared with those with gastroesophageal reflux disease (GERD) and healthy controls (HC).

Patients and Methods:

Pediatric patients evaluated by endoscopy were recruited for our study. Participants were categorized into 3 groups: EoE, GERD, and HC. RNA purified from esophageal biopsies were used for real‐time quantitative polymerase chain reaction assays and tested for forkhead box P3 (FoxP3) mRNA expression. Treg were identified as CD4+CD25hiCD127lo cells in peripheral blood and as CD3+/FoxP3+cells in esophageal tissue.

Results:

Forty‐eight subjects were analyzed by real‐time quantitative polymerase chain reaction: EoE (n = 33), GERD (n = 7), and HC (n = 8). FoxP3 expression was higher by up to 1.5‐fold in the EoE group compared with the GERD and HC groups (P < 0.05). Protein levels of FoxP3 in blood and tissue were then investigated in 21 subjects: EoE (n = 10), GERD (n = 6), and HC (n = 5). The percentage of Treg and their subsets in peripheral blood were not significant between groups (P > 0.05). The amount of Treg in esophageal tissue was significantly greater in the EoE group (mean 10.7 CD3+/FoxP3+cells/high power field [HPF]) compared with the other groups (GERD, mean 1.7 CD3+/FoxP3+cells/HPF and HC, mean 1.6 CD3+/FoxP3+cells/HPF) (P < 0.05).

Conclusions:

We show that Treg are increased in esophageal tissue of EoE subjects compared with GERD and HC subjects. The present study illustrates another possible mechanism involved in EoE that implicates impairment of immune homeostasis.
Increased HLA‐DR Expression on Tissue Eosinophils in Eosinophilic Esophagitis
Authors: Patel Anup J , Fuentebella Judy , Gernez Yael , Nguyen Tammie , Bass Dorsey , Berquist William , Cox Kenneth , Sibley Eric , Kerner John , Nadeau Kari
Published: July 2010

Objective:

The aim of the study was to investigate whether eosinophils have increased human leukocyte antigen (HLA)‐DR expression in subjects with eosinophilic esophagitis (EoE) compared with controls.

Patients and Methods:

Patients who were undergoing an upper endoscopy with biopsies for suspected gastroesophageal reflux disease (GERD) or EoE at Lucile Packard Children's Hospital were enrolled. In total, the blood and tissue samples of 10 healthy controls (HC), 11 subjects with GERD, and 10 with EoE were studied. Multiple tissue staining to identify eosinophils (via eosinophil cationic protein‐clone EG2) and major histocompatibility complex class II cell surface receptors (via HLA‐DR) was performed via immunohistochemistry. The peripheral blood was analyzed using flow cytometry to detect eosinophil HLA‐DR expression among these subjects.

Results:

In the tissue, a greater proportion of eosinophils expressed HLA‐DR among the subjects with EoE (mean 0.83 ± 0.14, n = 9) relative to those with GERD (mean 0.18 ± 0.19, n = 8, P < 0.01) and HC (mean 0.18 ± 0.13, n = 6, P < 0.01). In total, 6 participants (4 HC subjects and 2 subjects with GERD) did not have any eosinophils identified on tissue staining and were unable to be included in the present statistical analysis. In the blood, there was no statistically significant difference in eosinophil HLA‐DR expression among HC subjects (mean 415 ± 217, n = 6), subjects with GERD (mean 507 ± 429, n = 2), and those with EoE (mean 334 ± 181, n = 6).

Conclusions:

These data demonstrate that the eosinophils from the esophagus of subjects with EoE have increased HLA‐DR expression within this tissue.