bluuebunny/crossref_metadata_2025 huggingface dataset:
Alcohols, ethers, carbohydrates, and related compounds. IV. carbohydrates
Authors: Lii Jenn‐Huei , Chen Kuo‐Hsiang , Allinger Norman L.
Published: July 2003

Ab initio calculations [B3LYP/6‐311++G(2d,2p)] have been carried out on 84 conformations of 12 different sugars (hexoses), in both pyranose and furanose forms, with the idea of generating a data base for carbohydrate structural energies that may be used for developing the predictive value of molecular mechanics calculations for carbohydrates. The average value for the apparent gas phase anomeric effect for a series of 31 pairs of pyranose conformations was found to be 1.83 kcal/mol (vs. 2.67 kcal/mol with a smaller basis set used in earlier calculations). In developing MM4 to reproduce these data, it was necessary first to have good energies for simple alcohols and ethers, together with an adequate treatment of hydrogen bonding, and then to include the anomeric effect, and the ethylene glycol type system, as was previously recognized. It was also found that the so‐called delta‐2 effect, long recognized in carbohydrates, must be explicitly included, in order to obtain acceptable results. When a force field that included all of these items as developed from the small molecules based on the MM4 hydrocarbon force field was applied without any parameter adjustment to the set of hexopyranose and furanose conformations mentioned earlier, the Eβ − Eα was found to have an average value of 1.88 kcal/mol, versus 1.74 for the quantum calculations. The signed average and RMS deviations of the MM4 from the QM results were +0.15 and 0.87 kcal/mol. © 2003 Wiley Periodicals, Inc. J Comput Chem 24: 1504–1513, 2003
Possible molecular hydrogen formation mediated by the radical cations of anthracene and pyrene
Authors: Hirama Mutsumi , Ishida Toshimasa , Aihara Jun‐Ichi
Published: July 2003

Hydrogen molecules cannot be formed readily by the association of gaseous hydrogen atoms. Possible H2 formation mediated by the radical cations of typical polycyclic aromatic hydrocarbons (PAHs), anthracene and pyrene, was studied at the B3LYP/6‐31G** level of theory. We presumed that H2 is formed by way of two elementary reactions: the addition of an H atom to a PAH molecular cation, and the H abstraction from the resulting monohydro‐PAH cation (i.e., arenium ion) by a second H atom to yield H2. The first reaction takes place without any activation energy. The second reaction is also predicted to proceed along almost barrierless pathways, although it is far from being a typical ion–molecule reaction. There is a possibility that these reactions might constitute one of the mechanisms for H2 formation in extremely cold interstellar space. Deuterium enrichment in PAH cations is possibly accompanied by such H2 formation because deuteration lowers the energies of polyatomic PAH cations appreciably. © 2003 Wiley Periodicals, Inc. J Comput Chem 24: 1378–1382, 2003
Direct hydroxide attack is a plausible mechanism for amidase antibody 43C9
Authors: Chong Lillian T. , Bandyopadhyay Pradipta , Scanlan Thomas S. , Kuntz Irwin D. , Kollman Peter A.
Published: July 2003

Direct hydroxide attack on the scissile carbonyl of the substrate has been suggested as a likely mechanism for esterase antibodies elicited by phosphonate haptens, which mimic the transition states for the alkaline hydrolysis of esters.1 The unique amidase activity of esterase antibody 43C9 has been attributed to nucleophilic attack by an active‐site histidine residue.2 Yet, the active site of 43C9 is strikingly similar to those of other esterase antibodies, particularly 17E8. We have carried out quantum mechanical calculations, molecular dynamics simulations, and free energy calculations to assess the mechanism involving direct hydroxide attack for 43C9. Results support this mechanism and suggest that the mechanism is plausible for other antiphosphonate antibodies that catalyze the hydrolysis of (p‐nitro)phenyl esters. © 2003 Wiley Periodicals, Inc. J Comput Chem 24: 1371–1377, 2003
Solvent viscosity dependence of the folding rate of a small protein: Distributed computing study
Authors: Zagrovic Bojan , Pande Vijay
Published: July 2003

By using distributed computing techniques and a supercluster of more than 20,000 processors we simulated folding of a 20‐residue Trp Cage miniprotein in atomistic detail with implicit GB/SA solvent at a variety of solvent viscosities (γ). This allowed us to analyze the dependence of folding rates on viscosity. In particular, we focused on the low‐viscosity regime (values below the viscosity of water). In accordance with Kramers' theory, we observe approximately linear dependence of the folding rate on 1/γ for values from 1–10−1× that of water viscosity. However, for the regime between 10−4–10−1× that of water viscosity we observe power‐law dependence of the form k ∼ γ−1/5. These results suggest that estimating folding rates from molecular simulations run at low viscosity under the assumption of linear dependence of rate on inverse viscosity may lead to erroneous results. © 2003 Wiley Periodicals, Inc. J Comput Chem 24: 1432–1436, 2003
Electronic excitations of the green fluorescent protein chromophore in its protonation states: SAC/SAC‐CI study
Authors: Das Abhijit K. , Hasegawa Jun‐Ya , Miyahara Tomoo , Ehara Masahiro , Nakatsuji Hiroshi
Published: July 2003

Two ground‐state protonation forms causing different absorption peaks of the green fluorescent protein chromophore were investigated by the quantum mechanical SAC/SAC‐CI method with regard to the excitation energy, fluorescence energy, and ground‐state stability. The environmental effect was taken into account by a continuum spherical cavity model. The first excited state, HOMO‐LUMO excitation, has the largest transition moment and thus is thought to be the source of the absorption. The neutral and anionic forms were assigned to the protonation states for the experimental A‐ and B‐forms, respectively. The present results support the previous experimental observations. © 2003 Wiley Periodicals, Inc. J Comput Chem 24: 1421–1431, 2003
Flexible effective fragment QM/MM method: Validation through the challenging tests
Authors: Nemukhin A. V. , Grigorenko B. L. , Topol I. A. , Burt S. K.
Published: July 2003

A new version of the QM/MM method, which is based on the effective fragment potential (EFP) methodology [Gordon, M. et al., J Phys Chem A 2001, 105, 293] but allows flexible fragments, is verified through calculations of model molecular systems suggested by different authors as challenging tests for QM/MM approaches. For each example, the results of QM/MM calculations for a partitioned system are compared to the results of an all‐electron ab initio quantum chemical study of the entire system. In each case we were able to achieve approximately similar or better accuracy of the QM/MM results compared to those described in original publications. In all calculations we kept the same set of parameters of our QM/MM scheme. A new test example is considered when calculating the potential of internal rotation in the histidine dipeptide around the CαCβ side chain bond. © 2003 Wiley Periodicals, Inc. J Comput Chem 24: 1410–1420, 2003
Multireference perturbation theory with optimized partitioning. II. Applications to molecular systems
Authors: Witek Henryk A. , Nakano Haruyuki , Hirao Kimihiko
Published: July 2003

The second‐order multireference perturbation theory using an optimized partitioning, denoted as MROPT(2), is applied to calculations of various molecular properties—excitation energies, spectroscopic parameters, and potential energy curves—for five molecules: ethylene, butadiene, benzene, N2, and O2. The calculated results are compared with those obtained with second‐ and third‐order multireference perturbation theory using the traditional partitioning techniques. We also give results from computations using the multireference configuration interaction (MRCI) method. The presented results show very close resemblance between the new method and MRCI with renormalized Davidson correction. The accuracy of the new method is good and is comparable to that of second‐order multireference perturbation theory using Møller‐Plesset partitioning. © 2003 Wiley Periodicals, Inc. J Comput Chem 24: 1390–1400, 2003
Systematic and statistical error in histogram‐based free energy calculations
Authors: Kobrak Mark N.
Published: July 2003

A common technique for the numerical calculation of free energies involves estimation of the probability density along a given coordinate from a set of configurations generated via simulation. The process requires discretization of one or more reaction coordinates to generate a histogram from which the continuous probability density is inferred. We show that the finite size of the intervals used to construct the histogram leads to quantifiable systematic error. The width of these intervals also determines the statistical error in the free energy, and the choice of the appropriate interval is therefore driven by the need to balance the two sources of error. We present a method for the construction of the optimal histogram for a given system, and show that the use of this technique requires little additional computational expense. We demonstrate the efficacy of the technique for a model system, and discuss how the principles governing the choice of discretization interval could be used to improve extended sampling techniques. © 2003 Wiley Periodicals, Inc. J Comput Chem 24: 1437–1446, 2003
Immersion fixation with Carnoy solution for conventional immunohistochemical detection of particular N‐methyl‐D‐aspartate receptor subunits in murine hippocampus
Authors: Yoneyama Masanori , Kitayama Tomoya , Taniura Hideo , Yoneda Yukio
Published: July 2003

Immunoblotting analysis revealed heterologous distribution profiles of the N‐methyl‐D‐aspartate (NMDA) receptor subunits GluR ζ‐1, GluR ϵ‐1, and GluR ϵ‐2 in membrane preparations of different murine brain structures, with exclusive detection of GluR ϵ‐3 subunit in cerebellar preparations. Mice were anesthetized and perfused with 4% paraformaldehyde (PA), Zamboni, or Carnoy solution for subsequent immunohistochemical detection of these NMDA receptor subunits. In coronal brain sections from animals perfused with 4% PA and Zamboni solutions, high immunoreactivity was detected exclusively with GluR ζ‐1, GluR ϵ‐1, GluR ϵ‐2, and GluR ϵ‐3 subunits in the pyramidal and dentate granular layers, where the GluR ϵ‐3 subunit is supposed to be absent. By contrast, high immunoreactivity was detected with GluR ζ‐1, GluR ϵ‐1, and GluR ϵ‐2, but not GluR ϵ‐3, subunits in the strata oriens and radiatum of the CA1 subfield without immunoreactivity along the pyramidal and granular layers when sections were prepared by immersion fixation with Carnoy solution after dissection from brains of mice decapitated. On these sections, relatively high immunoreactivity was found also with GluR ζ‐1, GluR ϵ‐1, and GluR ϵ‐2 subunits in the stratum lacunosum‐moleculare of the CA1 region, the strata oriens, radiatum, and lacunosum‐moleculare of the CA3 region, and the stratum moleculare of the dentate gyrus, respectively. The systemic administration of kainate led to significant decreases in immunoreactivity to GluR ζ‐1, GluR ϵ‐1, and GluR ϵ‐2 subunits in the CA1 and CA3 subfields on brain sections prepared by immersion fixation with Carnoy solution. These results suggest that immersion fixation with Carnoy solution may be suitable and appropriate for reproducible and quantitative immunohistochemical detection of particular NMDA receptor subunits in murine hippocampus. © 2003 Wiley‐Liss, Inc.
Developmental and cell type‐specific expression of the neuronal marker NeuN in the murine cerebellum
Authors: Weyer Anja , Schilling Karl
Published: July 2003

NeuN is a 46/48‐kD nuclear protein antigen used widely to identify postmitotic neurons in both research and diagnostics. It is expressed by neurons throughout the nervous system of a variety of species, including birds, rodents, and man (Mullen et al. [1992] Development 116:201–211). When we sought to use NeuN to follow the developmental progression of murine cerebellar interneurons, we observed that expression of this antigen in the cerebellum was restricted to granule neurons and a small population of cells present in the lower molecular layer of the adult cerebellum. In an attempt to identify these cells, we combined immunostaining for NeuN with a panel of cell type‐specific markers to unambiguously identify neurons that express NeuN in the adult and developing cerebellum. In contrast to postmitotic granule neurons, NeuN was not expressed by any other immunocytochemically identified cerebellar interneurons, which comprised basket and stellate cells, Golgi neurons, unipolar brush cells, and Lugaro cells. NeuN‐positive cells in the molecular layer failed to express any cell type‐specific markers tested. They may represent ectopic granule cells; alternatively, they may represent a hitherto unknown population of cerebellar cells. In vitro experiments suggest that NeuN expression is related closely to granule cell axogenesis. This approach also revealed that the level of NeuN expression could be modulated by chronically depolarizing these cells. Thus, whereas NeuN expression per se is a reliable marker of proliferative capacity, levels of NeuN expression may also be indicative of the physiological status of a postmitotic neuron. © 2003 Wiley‐Liss, Inc.
Intravenous versus intrastriatal cord blood administration in a rodent model of stroke
Authors: Willing A.E. , Lixian J. , Milliken M. , Poulos S. , Zigova T. , Song S. , Hart C. , Sanchez‐Ramos J. , Sanberg P.R.
Published: July 2003

Human umbilical cord blood (hUCB) is a rich source of hematopoietic stem cells that have been used to reconstitute immune cells and blood lineages. Cells from another hematopoietic source, bone marrow, have been found to differentiate into neural cells and are effective in the treatment of stroke. In this study, we administered hUCB cells intravenously into the femoral vein or directly into the striatum and assessed which route of cell administration produced the greatest behavioral recovery in rats with permanent middle cerebral artery occlusion (MCAO). All animals were immunosuppressed with cyclosporine (CSA). When spontaneous activity was measured using the Digiscan automated system, it was found to be significantly less when hUCB was transplanted 24 hr after stroke compared with nontransplanted, stroked animals (P < 0.01). Furthermore, behavioral recovery was similar with both striatal and femoral hUCB delivery. This is in contrast to the step test, in which significant improvements were found only after femoral delivery of the hUCB cells. In the passive avoidance test, transplanted animals learned the task faster than nontransplanted animals (P < 0.05). Together, these results suggest that hUCB transplantation may be an effective treatment for brain injuries, such as stroke, or neurodegenerative disorders. In addition, intravenous delivery may be more effective than striatal delivery in producing long‐term functional benefits to the stroked animal. © 2003 Wiley‐Liss, Inc.
Human Aβ1–42 reduces iron‐induced toxicity in rat cerebral cortex
Authors: Bishop Glenda M. , Robinson Stephen R.
Published: July 2003

Senile plaques, the major neuropathological lesions of Alzheimer's disease (AD), are composed primarily of amyloid‐β (Aβ) peptide and contain high concentrations of iron (1.0 mM). We have previously shown that intracortical injections of 1.0 mM iron to adult rats produce significantly more neuronal loss than control injections of saline vehicle, whereas injections of Aβ do not. Because iron has been shown to increase the in vitro toxicity of Aβ, the present study was undertaken to determine whether iron can make Aβ neurotoxic in vivo. Aβ and 1.0 mM iron (as ferric ammonium citrate) were coinjected into rat cerebral cortex, and the neuronal loss was compared with that produced by pure Aβ or pure iron. The human and rat variants of Aβ1–42 were compared to determine whether they produce the same amount of neuronal loss when combined with iron. Coinjection of iron with either Aβ variant caused significantly more neuronal loss than Aβ peptide alone, suggesting that iron may contribute to the toxicity associated with senile plaques. Rat Aβ1–42 combined with iron was as toxic as iron alone, whereas iron combined with human Aβ1–42 was significantly less toxic. This latter finding indicates that fibrillar human Aβ is able to reduce iron‐induced neurotoxicity in vivo and raises the interesting possibility that senile plaques in AD may represent a neuroprotective response to the presence of elevated metal ions. © 2003 Wiley‐Liss, Inc.
Dopamine induces autophagic cell death and α‐synuclein increase in human neuroblastoma SH‐SY5Y cells
Authors: Gómez‐Santos Cristina , Ferrer Isidre , Santidrián Antonio F. , Barrachina Marta , Gil Joan , Ambrosio Santiago
Published: July 2003

Free cytoplasmic dopamine may be involved in the genesis of neuronal degeneration in Parkinson's disease and other such diseases. We used SH‐SY5Y human neuroblastoma cells to study the effect of dopamine on cell death, activation of stress‐induced pathways, and expression of α‐synuclein, the characteristic protein accumulated in Lewy bodies. We show that 100 and 500 μM dopamine causes a 40% and 60% decrease of viability, respectively, and triggers autophagy after 24 hr of exposure, characterized by the presence of numerous cytoplasmic vacuoles with inclusions. Dopamine causes mitochondrial aggregation in adherent cells prior to the loss of functionality. Plasma membrane and nucleus also maintain their integrity. Cell viability is protected by the dopamine transporter blocker nomifensine and the antioxidants N‐acetylcysteine and ascorbic acid. Dopamine activates the stress‐response kinases, SAPK/JNK and p38, but not ERK/MAPK or MEK, and increases α‐synuclein expression. Both cell viability and the increase in α‐synuclein expression are prevented by antioxidants; by the specific inhibitors of p38 and SAPK/JNK, SB203580 and SP600125, respectively; and by the inhibitor of autophagy 3‐methyladenine. This indicates that oxidative stress, stress‐activated kinases, and factors involved in autophagy up‐regulate α‐synuclein content. The results show that nonapoptotic death pathways are triggered by dopamine, leading to autophagy. These findings should be taken into account in the search for strategies to protect dopaminergic neurons from degeneration. © 2003 Wiley‐Liss, Inc.
Developmental expression of N‐methyl‐D‐aspartate glutamate receptor 1 splice variants in the chick retina
Authors: Lee‐Rivera I. , Zarain‐Herzberg A. , López‐Colomé A.M.
Published: July 2003

Glutamate is the major excitatory neurotransmitter in the vertebrate retina. The N‐methyl‐D‐aspartate glutamate receptor (NMDAR) is assembled as a tetramer containing NR1 and NR2, and possibly NR3 subunits, NR1 being essential for the formation of the ion channel. The NMDAR1 (NR1) gene encodes for mRNAs that generate at least eight functional variants by alternative splicing of exon 5 (cassette N1), 21 (cassette C1), or 22 (cassettes C2 or C2′). NR1 splice variants were identified in the mature chick retina, and their variation during embryonic development (ED) was analyzed. NR1 was shown to lack N1 in early ED, shifting to N1‐containing variants in the mature retina, which could contribute to explaining the distinct biochemical properties of retinal NMDARs compared with the CNS. Sequence analysis of C‐terminal variants containing C1 and C2 cassettes suggests a membrane‐targeting mechanism for avian NMDARs distinct from that in mammals. An NR1 variant containing a novel alternative C‐terminal splice exon named C3 was found, which encodes six amino acids containing a predicted casein kinase II phosphorylation site. This new variant is expressed in the retina during a restricted period of ED, coincident with the generation of spontaneous calcium activity waves, which precedes synapse formation in the retina, suggesting its participation in this process. © 2003 Wiley‐Liss, Inc.
γ‐Aminobutyric acid‐synthesizing cells in the retina of the chameleon Chamaeleo chameleon
Authors: Bennis M. , Versaux‐Botteri C. , Repérant J. , Armengol J.A. , Ward R.
Published: July 2003

Antibodies directed against γ‐aminobutyric acid (GABA) and L‐glutamic acid decarboxylases 65 and 67 kDa (GAD65 and ‐67) were used to study the GABAergic cell populations of the chameleon retina. GABA immunoreactivity was found in the two main types of retinal interneurons, amacrine and horizontal cells. Amacrine, displaced amacrine, and intra‐ and interplexiform cells displayed the strongest GABA immunoreactivity of all the retinal cell types. Horizontal cells formed a continuous GABA‐immunoreactive cell layer lying against the outermost portion of the inner nuclear layer. In contrast to previous studies (Quesada et al. [1996] Cell Biol. Int. 20:395–400; [1999] Eur. J. Anat. 3:13–25), the present results demonstrate that the horizontal cells of the chameleon retina are GABA immunoreactive and that a subpopulation of these is immunolabelled by an antibody against GAD65. These results indicate that GABAergic synaptic transmission plays a key role in the outer plexiform layer of the vertebrate retina. © 2003 Wiley‐Liss, Inc.
Transforming growth factor‐β and tumor necrosis factor‐α cooperate to induce apoptosis in the oligodendroglial cell line OLI‐neu
Authors: Schuster Norbert , Bender Herdis , Rössler Oliver G. , Philippi Anja , Dünker Nicole , Thiel Gerald , Krieglstein Kerstin
Published: July 2003

As shown previously, transforming growth factor‐β (TGF‐β) plays an important role during the period of developmental cell death in the nervous system. As with neurons, oligodendrocytes are generated in excess and eliminated by apoptosis. The present study was aimed at investigating the possible interaction of TGF‐β with tumor necrosis factor‐α (TNF‐α) in the regulation of cell death in oligodendroglial precursor cells and analyzing the underlying signaling mechanisms. We show that both factors induce apoptosis independently, but cooperate when applied together. The investigation of the signaling events revealed an important role of the JNK pathway during induction of apoptosis. TGF‐β seemed to be more efficient at inducing a release in cytochrome c from mitochondria than TNF‐α. This might be the consequence of decreased Bcl‐xL levels observed in cells treated with TGF‐β but not with TNF‐α. Both factors stimulated caspase‐3 activity, which could be inhibited by caspase‐8 or caspase‐9 inhibitors. Therefore, we conclude that TNF‐α and TGF‐β affect partially common pathways but also regulate different steps in the apoptotic cascade. © 2003 Wiley‐Liss, Inc.
Olanzapine protects PC12 cells from oxidative stress induced by hydrogen peroxide
Authors: Wei Zelan , Bai Ou , Richardson J. Steven , Mousseau Darrell D. , Li Xin‐Min
Published: July 2003

Neuroanatomical studies suggest that neuronal atrophy and destruction occur over the course of many years in neurodegenerative conditions such as schizophrenia and Alzheimer's disease. In schizophrenia, early intervention with atypical neuroleptics such as olanzapine has been shown to prevent development of some of the more serious and debilitating symptoms in many patients. The mechanisms whereby olanzapine slows or prevents symptom progression in schizophrenia remain unclear. A previous study found that olanzapine increased mRNA for the copper/zinc isoform of the superoxide dismutase enzyme (SOD‐1). We investigated the effects of olanzapine in PC12 cells exposed to hydrogen peroxide. We measured cell viability, observed evidence of necrosis and apoptosis, checked the SOD‐1 mRNA by Northern blot analyses, and determined SOD‐1 enzyme activity. We found that: 1) the decrease in cell viability induced by hydrogen peroxide was attenuated in PC12 cells pretreated with olanzapine; 2) olanzapine increased SOD enzyme activity in PC12 cells; 3) inhibiting SOD activity with diethyldithiocarbamic acid prevented the cytoprotective actions of olanzapine; and 4) the decrease in SOD‐1 mRNA level induced by hydrogen peroxide was blocked by pretreatment with olanzapine. These data indicate that the neuroprotective action of olanzapine includes the upregulation of SOD. © 2003 Wiley‐Liss, Inc.
Caspase‐independent cell death by low concentrations of nitric oxide in PC12 cells: Involvement of cytochrome C oxidase inhibition and the production of reactive oxygen species in mitochondria
Authors: Yuyama Kohei , Yamamoto Hideko , Nishizaki Itone , Kato Takeshi , Sora Ichiro , Yamamoto Toshifumi
Published: July 2003

We reported previously that low levels of nitric oxide (NO) induced cell death with properties of apoptosis, including chromatin fragmentation and condensation in undifferentiated PC12 pheochromocytoma cells. The present study demonstrates that cytotoxicity of low concentrations of NO is mediated by inhibition of mitochondrial cytochrome c oxidase and generation of reactive oxygen species (ROS). An NO donor, (±)‐(E)‐4‐ethyl‐2‐[(E)‐hydroxyimino]‐5‐nitro‐3‐hexenamide (NOR3) induced cell death even at low concentrations (10–100 μM), whereas peroxynitrite and a peroxynitrite generator, 3‐(4‐morpholinyl)‐sydnonimine (SIN‐1), did not have a significant effect on cell viability up to a concentration of 0.5 mM. The NOR3‐induced cell death was unaffected by pretreatment with superoxide dismutase (SOD) or its mimetic peroxynitrite scavenger, manganese(III) tetrakis(benzoic acid)porphyrin chloride (Mn‐TBAP), or with uric acid. These findings indicate that peroxynitrite does not contribute to this cell death. Furthermore, neither the release of cytochrome c from mitochondrial membranes, the cleavage of poly‐ADP ribose polymerase (PARP), nor the activation of caspase‐3‐like activities was observed. Inhibitors of PARP, benzamide, and aminobenzamide, had no effect on the NOR3‐induced cell death. In addition, pretreatment with general or selective caspase inhibitors, benzyloxy‐carbonyl‐Val‐Ala‐Asp‐fluoromethylketone (Z‐VAD‐fmk), N‐acetyl‐Asp‐Glu‐Val‐Asp‐aldehyde (Ac‐DEVD‐CHO), and benzyloxycarbonyl‐Asp‐2,6‐dichlorobenzoyloxymethylketone (Z‐Asp‐Ch2‐DCB) did not prevent NOR3‐induced cell death. Taken together, these findings suggest that cell death induced by NOR3 occurs by a caspase‐independent mechanism. In contrast, we found an early increase in mitochondrial H2O2 production during NOR3 exposure using the fluorescent dye 2′,7′‐dichlorofluorescin‐diacetate (DCFH‐DA) and dihydrorohdamine123 (DHR123), and these events were accompanied by strong inhibition of cytochrome c oxidase activity in the cells. Furthermore, we observed that several antioxidants, such as ascorbate, glutathione (GSH), cysteine, tetrahydrobiopterin, and dithiothreitol (DTT), all effectively prevented the NOR3‐induced cell death. NOR3 treatment decreased the level of total intracellular GSH, but did not affect the activities of antioxidant enzymes SOD, GSH‐peroxidase (GPX), and catalase. These results suggest that cell death induced at physiologically low concentrations of NO is mediated by ROS production in mitochondria, most likely resulting from the inhibition of cytochrome c oxidase, with ROS acting as an initiator of caspase‐independent cell death. © 2003 Wiley‐Liss, Inc.
Enhanced neurogenesis following stroke
Authors: Felling Ryan J. , Levison Steven W.
Published: July 2003

Each year hundreds of thousands of people must cope with the severe neurological consequences of a stroke. Current therapeutic strategies for stroke focus on acute treatment and neuroprotection. Unfortunately, these practices do little to reduce the long‐term morbidity associated with the injury. To develop effective therapies that promote regeneration, we must have an understanding of the cellular and molecular events involved in the recovery from an insult. Neural stem and progenitor cells are likely to be affected during this period. Here we review how the proliferation, migration, and maturation of these precursors are affected by ischemia. Furthermore, we summarize data available on the underlying mechanisms and the therapeutic implications of these studies. The studies that we review provide compelling evidence that neural precursors resident in the brain initiate a compensatory response to stroke that results in the production of new neurons. Moreover, administration of growth factors can enhance this compensatory response. Based on these encouraging results, we may eventually be able to manipulate these precursors to improve recovery of function in individuals afflicted by this devastating injury. © 2003 Wiley‐Liss, Inc.
Citicoline decreases phospholipase A2 stimulation and hydroxyl radical generation in transient cerebral ischemia
Authors: Adibhatla Rao Muralikrishna , Hatcher James F.
Published: July 2003

Neuroprotection by citicoline (CDP‐choline) in transient cerebral ischemia has been demonstrated previously. Citicoline has undergone several Phase III clinical trials for stroke, and is being evaluated for treatment of Alzheimer's and Parkinson's diseases. Phospholipid degradation and generation of reactive oxygen species (ROS) are major factors causing neuronal injury in CNS trauma and neurodegenerative diseases. Oxidative metabolism of arachidonic acid (released by the action of phospholipases) contributes to ROS generation. We examined the effect of citicoline on phospholipase A2 (PLA2) activity in relation to the attenuation of hydroxyl radical (OH·) generation after transient forebrain ischemia of gerbil. PLA2 activity (requires mM Ca2+) increased significantly (P < 0.05) in both membrane (50.2 ± 2.2 pmol/min/mg protein compared to sham 35.9 ± 3.2) and mitochondrial fractions (77.0 ± 1.2 pmol/min/mg protein compared to sham 33.9 ± 1.2) after cerebral ischemia and 2 hr reperfusion in gerbil, which was significantly attenuated (P < 0.01) by citicoline (membrane, 39.9. ± 2.2 and mitochondria, 41.9 ± 3.2 pmol/min/mg protein). In vitro, citicoline and its components cytidine and choline had no effect on PLA2 activity, and thus citicoline as such is not a PLA2 inhibitor. Ischemia/reperfusion resulted in significant OH· generation (P < 0.01) and citicoline significantly (P < 0.01) attenuated their formation (expressed as 2,3‐dihydroxybenzoic acid/salicylate ratio; ischemia/24 hr reperfusion, 6.30 ± 0.23; sham, 2.56 ± 0.27; ischemia/24 hr reperfusion + citicoline, 4.85 ± 0.35). These results suggest that citicoline affects PLA2 stimulation and decreases OH· generation after transient cerebral ischemia. © 2003 Wiley‐Liss, Inc.
Coexpression of functional P2X and P2Y nucleotide receptors in single cerebellar granule cells
Authors: Hervás Cristina , Pérez‐Sen Raquel , Miras‐Portugal Ma Teresa
Published: July 2003

The present study describes the presence and expression of functional nucleotide receptors, both ionotropic and metabotropic, in highly purified cultures of cerebellar granule neurons. Microfluorimetric experiments have been carried out to record specific [Ca2+]itransients in individual granule neurons after challenge with diverse nucleotides. Although great heterogeneity was found in nucleotide responses in single cells, these responses all became modified during the course of granule cell differentiation, not only at the level of the number of responding cells, but also in the magnitude of the response to nucleotides. These in vitro developmental changes were more significant in metabotropic responses to pyrimidine nucleotides, UTP and UDP, which were down‐ and upregulated, respectively, during the time in culture. At least two types of ADP‐specific receptors seem expressed in different granule cell subpopulations responding to 2MeSADP, as the specific P2Y1antagonist MRS‐2179 inhibited Ca2+responses in only one of these populations. The great diversity of metabotropic responses observed was confirmed by the RT‐PCR expression of different types of P2Y receptors in granule cell cultures: P2Y1, P2Y4, P2Y6, and P2Y12. Similarly, ionotropic nucleotide responses were confirmed by the presence of specific messengers for different P2X subunits, and by immunolabeling studies (P2X1, P2X2, P2X3, P2X4and P2X7). Immunolabeling reflected great variety in the P2X subunit distribution along the granule neuron cytoarchitecture, with P2X2, P2X3and P2X4present at somatodendritic locations, and P2X1, P2X7, and P2X3, located at the axodendritic prolongations. The punctuated labeling pattern obtained for P2X3and P2X7subunits is particularly notable, as it presents a high degree of colocalization with synaptophysin, a specific marker of synaptic vesicles, suggesting specialized localization and function in granule neurons. © 2003 Wiley‐Liss, Inc.
Tracking superparamagnetic iron oxide labeled monocytes in brain by high‐field magnetic resonance imaging
Authors: Zelivyanskaya Marina L. , Nelson Jay A. , Poluektova Larissa , Uberti Mariano , Mellon Melissa , Gendelman Howard E. , Boska Michael D.
Published: July 2003

Inflammatory cells, most notably mononuclear phagocytes (MP; macrophages and microglia), play a critical role in brain homeostasis, repair and disease. One important event in cellular biodynamics is how MP move in and throughout the nervous system. Prior studies have focused principally on cell migration across the blood–brain barrier during neuroinflammatory processes with little work done on cell movement within the brain. During the past decade our laboratories have studied the role of MP in HIV‐1‐associated dementia (HAD). In HAD MP incite sustained glial inflammatory reactions causing significant neuronal damage. To extend these works we investigated cell movement in brain and its influence for disease in a novel co‐registration system integrating neuropathology with high‐field magnetic resonance imaging (MRI). Human monocytes labeled with superparamagnetic iron oxide particles were injected into the brain of severe combined immunodeficient (SCID) mice. MRI was recorded 1, 7, and 14 days after cell injection. MRI co‐registered with histology verified that the MRI signal modification was due to the labeled cells. MRI showed human monocyte‐derived macrophages along the injection site, the corpus callosum, the ventricular system and in other brain sites. These data support the idea that cell migration can be monitored in vivo and provides an opportunity to assess monocyte mobility in brain and its affects on neurodegenerative processes and notably HAD. © 2003 Wiley‐Liss, Inc.
Biology of the basic multicellular unit and the pathophysiology of osteoporosis
Authors: Jilka Robert L.
Published: July 2003

Bone structural integrity is maintained by removal of old bone by osteoclasts and synthesis of new bone in its place by osteoblasts. This process, called bone remodeling, is accomplished by assembly of osteoclasts and osteoblasts into discrete temporary anatomic structures called basic multicellular units (BMUs). The lifespan of osteoclasts and osteoblasts is short compared to the lifespan of the BMU; therefore, they must be continually replenished for BMU progression to occur. Osteoclasts develop from hematopoeitic progenitors; and osteoblasts are derived from mesenchymal stem cells, also known as marrow stromal fibroblasts. Circulating hormones together with locally produced cytokines and growth factors modulate the replication and differentiation of osteoclast and osteoblast progenitors. The most important locally produced pro‐osteoclastogenic cytokine is receptor activator of NF‐κB (RANK) ligand. It is expressed by stromal/osteoblastic cells, and binds to its receptor (RANK) on osteoclast progenitors to stimulate differentiation. The development of mature osteoblasts is promoted by growth factors released from the bone matrix during resorption, as well as by growth factors produced by osteoblast progenitors themselves. Many of these same locally produced factors and cytokines also govern the lifespan of osteoclasts and osteoblasts by their effects on apoptosis. The orderly genesis and apoptosis of both osteoclasts and osteoblasts is essential for normal bone homeostasis during bone remodeling. Bone loss caused by sex steroid deficiency or glucocorticoid excess is caused by alteration of bone cell production, and by prolonging osteoclast lifespan and shortening osteoblast lifespan. Therapies that prevent or reverse osteoporosis act at least in part by preventing osteoblast apoptosis and/or stimulating osteoclast apoptosis. Med Pediatr Oncol 2003;41:182–185, Published 2003 Wiley‐Liss, Inc.
Cisplatin nephrotoxicity affects magnesium and calcium metabolism
Authors: Goren Marshall P.
Published: July 2003

Cisplatin is not directly toxic to bone, but cisplatin nephrotoxicity leading to magnesium wasting may affect magnesium and calcium metabolism, both of which contribute to bone integrity. The specificity of the magnesium lesion suggests that cisplatin may have an affinity for proteins that regulate magnesium absorption. Sulfhydryls such as amifostine can reduce the toxicity of cisplatin in adults, but current pediatric data do not indicate a role for sulfhydryl therapy to reduce cisplatin toxicity in children. Med Pediatr Oncol 2003;41:186–189, © 2003 Wiley‐Liss, Inc.
Chronic ifosfamide nephrotoxicity in children
Authors: Skinner R.
Published: July 2003

Nephrotoxicity is a frequent complication of treatment with ifosfamide in children. Although renal damage may be acute and reversible, chronic toxicity may develop with potentially serious consequences. Chronic nephrotoxicity due to ifosfamide may lead to a wide variety of subclinical and clinical manifestations. Proximal tubular dysfunction is the commonest presentation, and may lead to a Fanconi syndrome, including hypophosphataemic rickets and proximal renal tubular acidosis. Glomerular impairment is also common, whilst distal tubular impairment has been described but is relatively rare. Although full reversibility has been described occasionally, there is no information about the very long‐term outcome of chronic ifosfamide nephrotoxicity. We studied a cohort of 12 children 1 and 10 years after completion of ifosfamide treatment. There was no statistically significant change in either glomerular or tubular toxicity in the group as a whole over this time period. However, marked improvements were seen in some aspects of toxicity in some patients, and deterioration was observed in others. We concluded that considerable nephrotoxicity is still present 10 years after completion of ifosfamide treatment, but that the outcome varies between individual patients. Although several risk factors for the development of chronic nephrotoxicity have been described, total ifosfamide dose, patient age at treatment, previous or concurrent cisplatin treatment, and unilateral nephrectomy are the most important. Nevertheless, it remains difficult to predict the occurrence of this toxicity with confidence. The pathogenesis of ifosfamide nephrotoxicity is poorly understood. There is an urgent need for the development and clinical investigation of nephroprotective strategies. Med Pediatr Oncol 2003;41:190–197, © 2003 Wiley‐Liss, Inc.
Assessment of bone health in children and adolescents with cancer: Promises and pitfalls of current techniques
Authors: Leonard Mary B.
Published: July 2003

During childhood and adolescence, skeletal development is characterized by gender‐, face‐, and maturation‐specific increases in cortical dimensions and trabecular density. Children with cancer have multiple risk factors for impuired bone mineralization, including delayed growth and maturation, sex hormone deficiencies, decreasal physical activity and biomechanical loading of the skeleton, glucocorticoid and other immunosuppressive therapies, growth hormone deficiency, and malnutrition. This review outlines the expected gains in bone dimensions, mineral content and strength during childhood and adolescence. Varied threats to bone health in the child with cancer are summarized, with special attention to potential effects on bone formation and resorption in the growing skeleton. The strengths and limitations of dual energy x‐ray absorptiometry (DXA) and quantitative computed tomography (QCT) techniques in the assessment of the different disease‐related effects on bone strength are discussed, and alternative analytic approaches explored. Med Pediatr Oncol 2003;41:198–207. © 2003 Wiley‐Liss, Inc.
Radiation‐therapy effects on bone density
Authors: Hopewell John W.
Published: July 2003

Only limited data are available on the effects of radiation‐therapy on the mineral content of the bone of children treated for malignancy. The incidence of osteopenia varies between 8 and 23%, but confounding factors were the use of chemotherapy and the effects of prophylactic cranial irradiation. The factors influencing bone atrophy are no more clearly defined in adults treated for cancer by high dose local radiation‐therapy. Pathological observed in patient tissues, indicates a clear role for vascular changes in the development of osteopenia, although there remains some uncertainty as to the effects of osteoblast cell loss. Reduced blood flow in bone is clearly dose‐related in experimental animal studies and after single doses of >20 Gy changes in bone mineral content have been found. However, this was only at late times (≥30 weeks) after irradiation. The relationship between these changes and bone strength remains unproven because of the limited nature of many of the animal studies. Radiation dose fractionation data for rib‐fracture in breast cancer patients suggests an α/β ratio is in the range 1.8–2.8 Gy, comparable to values obtained for other late responding normal human tissues. Med Pediatr Oncol 2003;41:208–211. © 2003 Wiley‐Liss, Inc.
Effect of glucocorticoids on bone density
Authors: Rehman Qaiser , Lane Nancy E.
Published: July 2003

Glucocorticoid therapy is the most common cause of secondary iatrogenic osteoporosis. The bone loss occurs predominantly due to a decrease in bone formation, although increased bone resorption also occurs. Glucocorticoids induce osteoblast apoptosis and increase osteoclast survival and activity. Most of these effects are mediated through the RANKL‐OPG system but perturbations in gonadal hormone activity and calcium balance may also contribute significantly to bone loss. Future therapies in the treatment and prevention of glucocorticoid‐induced osteoporosis may be targeted at restoring the hormonal and cytokine balance to nullify the apoptotic effect of glucocorticoids on bone forming cells. Med Pediatr Oncol 2003;41:212–216. © 2003 Wiley‐Liss, Inc.
Role of estrogen and androgen in pubertal skeletal physiology
Authors: Frank Graeme R.
Published: July 2003

Since both estrogen and androgen are present in each sex, it has been difficult to discern the exact role that each sex steroid plays in skeletal physiology. However, studying clinical syndromes in which there is either only estrogen or androgen action has allowed us to gain insight into the unique role that each sex steroid plays in the growing skeleton. In complete androgen insensitivity syndrome (AIS) the only functional sex steroid receptor is that for estrogen. Effected XY females have a pubertal growth spurt that is typical of normal females, both in magnitude and timing. Individuals with AIS have a mild reduction in bone density but it is difficult to distinguish whether this is the result of androgen resistance or estrogen deficiency. These observations suggest that estrogen action only is sufficient to induce a normal pubertal growth spurt, epiphyseal maturation, and near normal bone mineral accretion in women. Until recently, the skeletal effects of estrogen were not thought to be of importance in the male. Conventional wisdom dictated that, in the male, testosterone mediated these skeletal changes. The notion that estrogen is of little importance in the male has been challenged by the recent discovery of two human syndromes in which estrogen action is lacking. In males with either estrogen resistance (inability to respond to circulating estrogen) or aromatase deficiency (inability to synthesize estradiol), as a result of the lack of estrogen action, a pubertal growth spurt does not appear to occur. Furthermore, complete epiphyseal maturation does not take place allowing for continued growth in adulthood and resultant tall stature. Finally normal bone mineral accretion does not take place resulting in severe osteoporosis. These findings indicate that estrogen plays a critical role in skeletal physiology of males as well as females. Med Pediatr Oncol 2003;41:217–221. © 2003 Wiley‐Liss, Inc.
Management of osteoporosis due to ovarian failure
Authors: Eastell Richard
Published: July 2003

The management of oestrogen deficiency bone loss needs to include general measures to protect against osteoporosis, the identification and treatment of other reversible causes of bone loss, and the use of proven agents for the treatment of osteoporosis. The general measures include improved physical activity, adequate diet (paying particular attention to calcium and vitamin D), and avoidance of behaviours that promote bone loss, such as smoking and alcohol abuse. The diseases that should be identified, other than estrogen‐deficiency, include primary hyperparathyroidism, thyrotoxicosis and celiac disease. The treatments that are proven to prevent fractures in women with estrogen deficiency, include hormone replacement therapy, raloxifene, nasal calcitonin, bisphosphonates, (alendronate and risedronate) and parathyroid hormone. The most appropriate therapy in the younger woman is HRT, although the trial‐based evidence that HRT prevents fractures is not strong. There is a wide choice of preparations and the use of continuous combined preparations avoids regular menstrual periods, one of the limitations to the use of HRT. Raloxifene has less effect on bone mineral density than HRT, but a similar effect on vertebral fractures and does not result in menstrual bleeding or increased risk of breast cancer. There is recent evidence suggesting that the beneficial effects on lipids translate into reduced risk of cardiovascular disease. Bisphosphonates are the standard treatment for the older woman with osteoporosis. Alendronate has been found to reduce the risk of spine, hip, and wrist fractures and has approval for a once weekly regimen, an approach that appears to prevent GI side effects. Risedronate reduces the risk of spine and non‐vertebral fractures within the first year of treatment and has been shown to reduce the risk of hip fracture. It has not been associated with an excess of GI side effects. Parathyroid hormone therapy results in increases in BMD that are even greater than estrogen and the bisphosphonates and to an even greater reduction in the risk of fractures, particularly non‐vertebral fractures. It works by stimulation of bone formation rather than by inhibition of bone resorption. However, it has to be given by daily injection. Thus, we have a wide choice of therapies for the woman with osteoporosis due to ovarian failure. Med Pediatr Oncol 2003;41:222–227. © 2003 Wiley‐Liss, Inc.