bluuebunny/crossref_metadata_2025 huggingface dataset:
Caspase cleavage of members of the amyloid precursor family of proteins
Authors: Galvan Veronica , Chen Sylvia , Lu Daniel , Logvinova Anna , Goldsmith Paul , Koo Edward H. , Bredesen Dale E.
Published: March 2003

The synapse loss and neuronal cell death characteristic of Alzheimer's disease (AD) are believed to result in large part from the neurotoxic effects of β‐amyloid peptide (Aβ), a 40–42 amino acid peptide(s) derived proteolytically from β‐amyloid precursor protein (APP). However, APP is also cleaved intracellularly to generate a second cytotoxic peptide, C31, and this cleavage event occurs in vivo as well as in vitro and preferentially in the brains of AD patients (Lu et al. 2000). Here we show that APPC31 is toxic to neurons in primary culture, and that like APP, the APP family members APLP1 and possibly APLP2 are cleaved by caspases at their C‐termini. The carboxy‐terminal peptide derived from caspase cleavage of APLP1 shows a degree of neurotoxicity comparable to APPC31. Our results suggest that even though APLP1 and APLP2 cannot generate Aβ, they may potentially contribute to the pathology of AD by generating peptide fragments whose toxicity is comparable to that of APPC31.
Generation of C‐terminally truncated amyloid‐β peptides is dependent on γ‐secretase activity
Authors: Beher Dirk , Wrigley Jonathan D. J. , Owens Andrew P. , Shearman Mark S.
Published: March 2003

Aberrant production of amyloid‐β peptides by processing of the β‐amyloid precursor protein leads to the formation of characteristic extracellular protein deposits which are thought to be the cause of Alzheimer's disease. Therefore, inhibiting the key enzymes responsible for amyloid‐β peptide generation, β‐ and γ‐secretase may offer an opportunity to intervene with the progression of the disease. In human brain and cell culture systems a heterogeneous population of amyloid‐β peptides with various truncations is detected and at present, it is unclear how they are produced. We have used a combination of surface enhanced laser desorption/ionization time‐of‐flight mass spectrometry (SELDI‐TOF MS) and a specific inhibitor of γ‐secretase to investigate whether the production of all amyloid‐β peptide species requires the action of γ‐secretase. Using this approach, we demonstrate that the production of all truncated amyloid‐β peptides except those released by the action of the nonamyloidogenic α‐secretase enzyme or potentially beta‐site βAPP cleaving enzyme 2 depends on γ‐secretase activity. This indicates that none of these peptides are generated by a separate enzyme entity and a specific inhibitor of the γ‐secretase enzyme should havethe potential to block the generation of all amyloidogenicpeptides. Furthermore in the presence of γ‐secretase inhibitors, the observation of increased cleavage of the membrane‐bound βAPP C‐terminal fragment C99 by α‐secretase suggests that during its trafficking C99 encounters compartments in which α‐secretase activity resides.
1‐Methyl‐4‐phenylpyridinium ion, a toxin that can cause parkinsonism, alters branched structures of DNA
Authors: Iwaasa Mitsutoshi , Umeda Shuyo , Ohsato Takashi , Takamatsu Chihiro , Fukuoh Atushi , Iwasaki Hiroshi , Shinagawa Hideo , Hamasaki Naotaka , Kang Dongchon
Published: March 2003

During replication, human mitochondrial DNA (mtDNA) takes on a triple‐stranded structure known as a D‐loop, which is implicated in replication and transcription. 1‐Methyl‐4‐phenylpyridinium ion (MPP+), a toxin inducing parkinsonism, inhibits mtDNA replication, possibly by resolving the D‐loops. For initiation of mtDNA replication, mitochondria are thought to have another triple‐stranded structure, an R‐loop. The R‐loop, which is resolved by a bacterial junction‐specific helicase, RecG, is also resolved by MPP+. Because mitochondrial D‐loops are likewise resolved by RecG, the D‐ and R‐loops may share a similar branched structure. MPP+ resolves cruciform DNA in supercoiled DNA. MPP+ converts a stacked conformation to an extended conformation in a synthetic Holliday junction. This conversion is reversed by 1 mm Mg2+, as is the resolution of the D‐loops or cruciform DNA. These observations suggest that the junction structure of mitochondrial D‐ and R‐loops is affected by MPP+.
No correlation between aggregates of Cu/Zn superoxide dismutase and cell death in familial amyotrophic lateral sclerosis
Authors: Lee J. P. , Gerin C. , Bindokas V. P. , Miller R. , Ghadge G. , Roos R. P.
Published: March 2003

Aggregates of Cu/Zn superoxide dismutase (SOD) have been demonstrated in familial amyotrophic lateral sclerosis (FALS) and other neurodegenerative diseases; however, their role in disease pathogenesis is unclear. In this study, we investigated the presence of SOD aggregates in nerve growth factor (NGF)‐differentiated PC12 cells and cell viability following: (i) transduction with replication‐deficient recombinant adenoviruses (AdVs) expressing wild‐type SOD (SODWT) or mutant SOD (SODMT, V148G or A4V); (ii) transfection of yellow fluorescent protein‐tagged SODWT (SODWT‐YFP) or SODMT (SODA4V‐YFP, SODV148G‐YFP). SOD aggregates were more prominent in cells following transduction of AdSODMT than AdSODWT and following treatment with H2O2, suggesting that mutant SOD leads to oxidation of cellular components. In addition, cells expressing SODMT‐YFP yielded SOD aggregates that were significantly larger and more frequent than SOD aggregates in cells expressing SODWT‐YFP. Proteasome inhibitors, but not cathepsin B inhibitors, increased aggregate formation but did not increase cell death. In addition, treatments that increased cell viability did not significantly decrease SOD aggregates. Taken together, our data demonstrate that there is no association between SOD aggregates and cell death in FALS.
Deletion of the glutamate carboxypeptidase II gene in mice reveals a second enzyme activity that hydrolyzes N‐acetylaspartylglutamate
Authors: Bacich Dean J. , Ramadan Epolia , O'Keefe Denise S. , Bukhari Noreen , Wegorzewska Iga , Ojeifo Olumide , Olszewski Rafal , Wrenn Craige C. , Bzdega Tomasz , Wroblewska Barbara , Heston Warren D. W. , Neale Joseph H.
Published: March 2003

Glutamate carboxypeptidase II (GCPII, EC 3.14.17.21) is a membrane‐bound enzyme found on the extracellular face ofglia. The gene for this enzyme is designated FOLH1 in humans and Folh1 in mice. This enzyme has been proposed to be responsible for inactivation of the neurotransmitter N‐acetylaspartylglutamate (NAAG) following synaptic release. Mice harboring a disruption of the gene for GCPII/Folh1 were generated by inserting into the genome a targeting cassette in which the intron–exon boundary sequences of exons 1 and 2 were removed and stop codons were inserted in exons 1 and 2. Messenger RNA for GCPII was not detected by northern blotting or RT‐PCR analysis of RNA from the brains of –/– mutant mice nor was GCPII protein detected on western blots of this tissue. These GCPII null mutant mice developed normally to adulthood and exhibited a normal range of neurologic responses and behaviors including mating, open field activity and retention of position in rotorod tests. No significant differences were observed among responses of wild type, heterozygous mutant and homozygous mutant mice on tail flick and hot plate latency tests. Glutamate, NAAG and mRNA for metabotropic glutamate receptor type 3 levels were not significantly altered in response to the deletion of glutamate carboxypeptidase II. A novel membrane‐bound NAAG peptidase activity was discovered in brain, spinal cord and kidney of the GCPII knock out mice. The kinetic values for brain NAAG peptidase activity in the wild type and GCPII nullmutant were Vmax = 45 and 3 pmol/mg/min and Km = 2650 nm and 2494 nm, respectively. With the exception of magnesium and copper, this novel peptidase activity had a similar requirement for metal ions as GCPII. Two potent inhibitors of GCPII, 4,4′‐phosphinicobis‐(butane‐1,3 dicarboxilic acid) (FN6) and 2‐(phosphonomethyl)pentanedioic acid (2‐PMPA) inhibited the residual activity. The IC50 value for 2‐PMPA was about 1 nm for wild‐type brain membrane NAAG peptidase activity consistent with its activity against cloned ratand human GCPII, and 88 nm for the activity in brain membranes of the null mutants.
Increased vulnerability of dopaminergic neurons in MPTP‐lesioned interleukin‐6 deficient mice
Authors: Bolin Laurel M. , Strycharska‐Orczyk Iwona , Murray Richard , Langston J. William , Di Monte Donato
Published: March 2003

To test the hypothesis that neuroinflammation contributes to dopaminergic neuron death in the MPTP‐lesioned mouse, we compared nigrostriatal degeneration in interleukin (IL)‐6 (+/+) with IL‐6 (–/–) mice. In the absence of IL‐6, a single injection of MPTP (30 mg/kg) resulted in significantly greater striatal dopamine depletion than that measured in IL‐6 (+/+) mice. The observed dopamine depletion was MPTP dose dependent. This loss of striatal dopamine and a significantly greater loss of TH+ cells in the substantia nigra pars compacta in IL‐6 (–/–) mice as compared with control IL‐6 (+/+) mice, suggest that IL‐6 is neuroprotective in the MPTP‐lesioned nigrostriatal system. Co‐localization experiments identified striatal astrocytes as the source of IL‐6 in IL‐6 (+/+) mice at 1 and 7 days postinjection of MPTP. The increased sensitivity of dopaminergic neurons to neurotoxicant in the absence of IL‐6, is compatible with a neuroprotective activity of IL‐6 in the injured nigrostriatal system.
Involvement of Gadd153 in the pathogenic action of presenilin‐1 mutations
Authors: Milhavet Ollivier , Martindale Jennifer L. , Camandola Simonetta , Chan Sic L. , Gary Devin S. , Cheng Aiwu , Holbrook Nikki J. , Mattson Mark P.
Published: March 2003

Mutations in the presenilin‐1 (PS1) gene cause early onset familial Alzheimer's disease (FAD) by a mechanism believed to involve perturbed endoplasmic reticulum (ER) function and altered proteolytic processing of the amyloid precursor protein. We investigated the molecular mechanisms underlying cell death and ER dysfunction in cultured cells and knock‐in mice expressing FAD PS1 mutations. We report that PS1 mutations cause a marked increase in basal protein levels of the pro‐apoptotic transcription factor Gadd153. PS1 mutations increase Gadd153 protein translation without affecting mRNA levels, while decreasing levels of the anti‐apoptotic protein Bcl‐2. Moreover, an exaggerated Gadd153 response to stress induced by ER stress agents was observed in PS1 mutant cells. Cell death in response to ER stress is enhanced by PS1 mutations, and this endangering effect is attenuated by anti‐sense‐mediated suppression of Gadd153 production. An abnormality in the translational regulation of Gadd153 may sensitize cells to the detrimental effects of ER stress and contribute to the pathogenic actions of PS1 mutations in FAD.
Signaling mechanisms that regulate actin‐based motility processes in the nervous system
Authors: Meyer Gary , Feldman Eva L.
Published: March 2003

Actin‐based motility is critical for nervous system development. Both the migration of neurons and the extension of neurites require organized actin polymerization to push the cell membrane forward. Numerous extracellular stimulants of motility and axon guidance cues regulate actin‐based motility through the rho GTPases (rho, rac, and cdc42). The rho GTPases reorganize the actin cytoskeleton, leading to stress fiber, filopodium, or lamellipodium formation. The activity of the rho GTPases is regulated by a variety of proteins that either stimulate GTP uptake (activation) or hydrolysis (inactivation). These proteins potentially link extracellular signals to the activation state of rho GTPases. Effectors downstream of the rho GTPases that directly influence actin polymerization have been identified and are involved in neurite development. The Arp2/3 complex nucleates the formation of new actin branches that extend the membrane forward. Ena/VASP proteins can cause the formation of longer actin filaments, characteristic of growth cone actin morphology, by preventing the capping of barbed ends. Actin‐depolymerizing factor (ADF)/cofilin depolymerizes and severs actin branches in older parts of the actin meshwork, freeing monomers to be re‐incorporated into actively growing filaments. The signaling mechanisms by which extracellular cues that guide axons to their targets lead to direct effects on actin filament dynamics are becoming better understood.
Increased methamphetamine‐induced locomotor activity and behavioral sensitization in histamine‐deficient mice
Authors: Kubota Yasuhiko , Ito Chihiro , Sakurai Eiichi , Sakurai Eiko , Watanabe Takehiko , Ohtsu Hiroshi
Published: March 2003

We have recently suggested that the brain histamine has an inhibitory role on the behavioral effects of methamphetamine by pharmacological studies. In this study, we used the histidine decarboxylase gene knockout mice and measured the spontaneous locomotor activity, the changes of locomotion by single and repeated administrations of methamphetamine, and the contents of brain monoamines and amino acids at 1 h after a single administration of methamphetamine. In the histidine decarboxylase gene knockout mice, spontaneous locomotor activity during the dark period was significantly lower than in the wild‐type mice. Interestingly, methamphetamine‐induced locomotor hyperactivity and behavioral sensitization were facilitated more in the histidine decarboxylase gene knockout mice. In the neurochemical study, noradrenaline and O‐phosphoserine were decreased in the midbrain of the saline‐treated histidine decarboxylase gene knockout mice. On the other hand, single administration of methamphetamine decreased GABA content of the midbrain of the wild‐type mice, but did not alter that of histidine decarboxylase gene knockout mice. These results suggest that the histamine neuron system plays a role as an awakening amine in concert with the noradrenaline neuron system, whereas it has an inhibitory role on the behavioral effects of methamphetamine through the interaction with the GABAergic neuron system.
The neuropathogenic contributions of lysosomal dysfunction
Authors: Bahr Ben A. , Bendiske Jennifer
Published: March 2003

Multiple lines of evidence implicate lysosomes in a variety of pathogenic events that produce neurodegeneration. Genetic mutations that cause specific enzyme deficiencies account for more than 40 lysosomal storage disorders. These mostly pre‐adult diseases are associated with abnormal brain development and mental retardation. Such disorders are characterized by intracellular deposition and protein aggregation, events also found in age‐related neurodegenerative diseases including (i) Alzheimer's disease and related tauopathies (ii) Lewy body disorders and synucleinopathies such as Parkinson's disease, and (iii) Huntington's disease and other polyglutamine expansion disorders. Of particular interest for this review is evidence that alterations to the lysosomal system contribute to protein deposits associated with different types of age‐related neurodegeneration. Lysosomes are in fact highly susceptible to free radical oxidative stress in the aging brain, leading to the gradual loss of their processing capacity over the lifespan of an individual. Several studies point to this lysosomal disturbance as being involved in amyloidogenic processing, formation of paired helical filaments, and the aggregation of α‐synuclein and mutant huntingtin proteins. Most notably, experimentally induced lysosomal dysfunction, both in vitro and in vivo, recapitulates important pathological features of age‐related diseases including the link between protein deposition and synaptic loss.
Neuroprotective effects of bcl‐2 overexpression in hippocampal cultures: interactions with pathways of oxidative damage
Authors: Howard Sarah , Bottino Clement , Brooke Sheila , Cheng Elise , Giffard Rona G. , Sapolsky Robert
Published: March 2003

Overexpression of bcl‐2protects neurons from numerous necrotic insults, both in vitro and in vivo. While the bulk of such protection is thought to arise from Bcl‐2 blocking cytochrome c release from mitochondria, thereby blocking apoptosis, the protein can target other steps in apoptosis, and can protect against necrotic cell death as well. There is evidence that these additional actions may be antioxidant in nature, in that Bcl‐2 has been reported to protect against generators of reactive oxygen species (ROS), to increase antioxidant defenses and to decrease levels of ROS and of oxidative damage. Despite this, there are also reports arguing against either the occurrence, or the importance of these antioxidant actions. We have examined these issues in neuron‐enriched primary hippocampal cultures, with overexpression of bcl‐2 driven by a herpes simplex virus amplicon: (i) Bcl‐2 protected strongly against glutamate, whose toxicity is at least partially ROS‐dependent. Such protection involved reduction in mitochondrially derived superoxide. Despite that, Bcl‐2 had no effect on levels of lipid peroxidation, which is thought to be the primary locus of glutamate‐induced oxidative damage; (ii) Bcl‐2 was also mildly protective against the pro‐oxidant adriamycin. However, it did so without reducing levels of superoxide, hydrogen peroxide or lipid peroxidation; (iii) Bcl‐2 protected against permanent anoxia, an insult likely to involve little to no ROS generation. These findings suggest that Bcl‐2 can have antioxidant actions that may nonetheless not be central to its protective effects, can protect against an ROS generator without targeting steps specific to oxidative biochemistry, and can protect in the absence of ROS generation. Thus, the antioxidant actions of Bcl‐2 are neither necessary nor sufficient to explain its protective actions against these insults in hippocampal neurons.
High transcytosis of melanotransferrin (P97) across the blood–brain barrier
Authors: Demeule Michel , Poirier Julie , Jodoin Julie , Bertrand Yanick , Desrosiers Richard R. , Dagenais Claude , Nguyen Tran , Lanthier Julie , Gabathuler Reinhard , Kennard Malcolm , Jefferies Wilfred A. , Karkan Delara , Tsai Sam , Fenart Laurence , Cecchelli Roméo , Béliveau Richard
Published: March 2003

The blood–brain barrier (BBB) performs a neuroprotective function by tightly controlling access to the brain; consequently it also impedes access of proteins as well as pharmacological agents to cerebral tissues. We demonstrate here that recombinant human melanotransferrin (P97) is highly accumulated into the mouse brain following intravenous injection and in situ brain perfusion. Moreover, P97 transcytosis across bovine brain capillary endothelial cell (BBCEC) monolayers is at least 14‐fold higher than that of holo‐transferrin, with no apparent intra‐endothelial degradation. This high transcytosis of P97 was not related to changes in the BBCEC monolayer integrity. In addition, the transendothelial transport of P97 was sensitive to temperature and was both concentration‐ and conformation‐dependent, suggesting that the transport of P97 is due to receptor‐mediated endocytosis. In spite of the high degree of sequence identity between P97 and transferrin, a different receptor than the one for transferrin is involved in P97 transendothelial transport. A member of the low‐density lipoprotein receptor protein family, likely LRP, seems to be involved in P97 transendothelial transport. The brain accumulation, high rate of P97 transcytosis and its very low level in the blood suggest that P97 could be advantageously employed as a new delivery system to target drugs directly to the brain.
Subtype‐specific regulation of the expression and function of muscarinic acetylcholine receptors in embryonic chicken retinal cells
Authors: Goin Juan C. , Nathanson Neil M.
Published: March 2003

We examined the effect of long‐term agonist exposure on muscarinic acetylcholine receptor expression and function in embryonic chicken retinal cells. Long‐term carbachol exposure induced a time‐ and concentration‐dependent decrease in M2, M3 and M4 muscarinic receptor numbers. Kinetic analyses revealed a first‐order process with similar rate constants for all three subtypes. Both the maximal decrease and the agonist potency for regulation of M3 were significantly higher than those for M2 and M4. Upon agonist removal, M2 and M4 numbers returned to control values, but M3 recovery after 24 h was no higher than 40%. Agonist treatment did not alter the levels of receptor mRNAs. Receptor inactivation with a covalent alkylating antagonist demonstrated that the partial M3 protein recovery was not due to a decreased intrinsic basal rate of synthesis, suggesting that it is induced by agonist treatment. Prolonged carbachol exposure induced concomitant decreases in muscarinic‐mediated inhibition of cyclic AMP accumulation which were completely reversed after agonist removal. Sustained receptor activation also promoted significant decreases in muscarinic receptor‐stimulated phosphoinositide turnover, which were only partially reversed after agonist removal. These data demonstrate subtype‐specific regulation of the expression and function of muscarinic receptors in the retina.
Antipsychotic drug treatment induces differential gene expression in the rat cortex
Authors: Kontkanen Outi , Törönen Petri , Lakso Merja , Wong Garry , Castrén Eero
Published: March 2003

Antipsychotic drug treatment is known to modulate gene expression in experimental animals. In this study, candidate target genes for antipsychotic drug action were searched using microarrays after acute clozapine treatment (1, 6 and 24 h) in the rat prefrontal cortex. Microarray data clustering with a self‐organizing map algorithm revealed differential expression of genes involved in presynaptic function following acute clozapine treatment. The differential expression of 35 genes most profoundly regulated in expression arrays was further examined using in situ hybridization following acute clozapine, and chronic clozapine and haloperidol treatments. Acute administration of clozapine regulated the expression of chromogranin A, synaptotagmin V and calcineurin A mRNAs in the cortex. Chronic clozapine treatment induced differential cortical expression of chromogranin A, son of sevenless (SoS) and Sec‐1. Chronic treatment with haloperidol regulated the mRNA expression of inhibitor of DNA‐binding 2 (ID‐2) and Rab‐12. Furthermore, the expression of visinin‐like proteins‐1, ‐2 and ‐3 was regulated by chronic drug treatments in various brain regions. Our data suggest that acute and chronic treatments with haloperidol and clozapine modulate the expression of genes involved in synaptic function and in regulation of intracellular Ca2+ in cortex.
CD44 deficiency in mice protects brain from cerebral ischemia injury
Authors: Wang Xinkang , Xu Lin , Wang Hugh , Zhan Yutian , Puré Ellen , Feuerstein Giora Z.
Published: March 2003

CD44 is a transmembrane glycoprotein known to be involved in endothelial cell recognition, lymphocyte trafficking, and regulation of cytokine gene expression in inflammatory diseases. In the present study, we demonstrated that the expression of CD44 mRNA was induced in a mouse model of cerebral ischemia. A potential role of CD44 in ischemic brain injury was investigated using CD44‐deficient (CD44–/–) mice. Over 50% (p < 0.05, n = 14) and 78% (p < 0.05, n = 10) reduction in ischemic infarct was observed in CD44–/– mice compared with that of wild‐type mice following transient (30 min ischemia) and permanent (24 h) occlusion of the middle cerebral artery (MCAO), respectively. Similarly, significant improvement was observed in neurological function in CD44–/– mice as evidenced by spontaneous and forced motor task scores. The marked protection from ischemic brain injury in CD44–/– mice was associated with normal physiological parameters, cytokine gene expression, astrocyte and microglia activation as compared with wild‐type mice. However, in CD44–/– mice, significantly lower expression of soluble interleukin‐1β protein was noted after brain ischemia. Our data provide new evidence on the potential role of CD44 in brain tissue in response to ischemia and may suggest that this effect might be associated with selective reduction in inflammatory cytokines such as interleukin‐1β.
The anti‐inflammatory activity of estrogen in glial cells is regulated by the PKC‐anchoring protein RACK‐1
Authors: Viviani Barbara , Corsini Emanuela , Binaglia Marco , Lucchi Laura , Galli Corrado L. , Marinovich Marina
Published: March 2003

It has recently been suggested that estrogen inhibits glial activation and the release of neurotoxic mediators. The mechanisms involved in this anti‐inflammatory effect are unclear. We found that an nM concentration of 17‐β estradiol inhibits protein kinaseC‐βII translocation induced by lipopolysaccharide in primary astrocytes. Estradiol treatment did not change the total content of kinaseC‐βII or of lipopolysaccharide receptor, but dose‐dependently reduced the levels of receptors for activated C kinases‐1 (RACK‐1), the anchoring protein involved in protein kinase C (PKC) shuttling. This decrease could thus account for the defective protein kinaseC‐βII activation. Pre‐treatment with 1 nmβ‐estradiol, which reduced by ∼35% the expression of RACK‐1, prevented the lipopolysaccharide‐induced expression of tumour necrosis factor‐α mRNA and of the inducible form of nitric oxide (NO) synthase. As a consequence, the production of tumour necrosis factor‐α and NO were decreased. An antisense oligonucleotide for RACK‐1 also reduced tumour necrosis factor‐α and nitric oxide production on lipopolysaccharide stimulation. These results demonstrate that estrogen reduction of the RACK‐1 expression, leading to a defective protein kinase‐C activation counteracts the inflammatory response in astrocytes.
Differences in multidrug resistance phenotype and matrix metalloproteinases activity between endothelial cells from normal brain and glioma
Authors: Régina Anthony , Demeule Michel , Bérubé Alexandra , Moumdjian Robert , Berthelet France , Béliveau Richard
Published: March 2003

Endothelial cells (ECs) are new targets for tumor therapy. In this work, we purified endothelial cells from intracerebral and subcutaneous experimental gliomas as well as from normal brain in order to define some of the phenotypical differences between angiogenic and quiescent brain vasculature. We show that the multidrug resistance genes encoding drug efflux pumps at the brain endothelium are expressed differently in normal and tumoral vasculature. We also show that ECs from gliomas present increased activity of gelatinase B (MMP9), key enzyme in the angiogenic process. Importantly, we observe a different phenotype between ECs in the intracerebral and subcutaneous models. Our results provide molecular evidence of phenotypic distinction between tumoral and normal brain vasculature and indicate that the EC phenotype depends on interactions both with tumor cells and also with the microenvironment.
The immunoglobulin‐superfamily molecule basigin is a binding protein for oligomannosidic carbohydrates: an anti‐idiotypic approach
Authors: Heller Martin , von der Ohe Maren , Kleene Ralf , Mohajeri M. Hasan , Schachner Melitta
Published: March 2003

Recognition molecules that carry carbohydrate structures regulate cell interactions during development and play important roles in synaptic plasticity and regeneration in the adult. Glycans appear to be involved in these interactions. We have searched for binding proteins for oligomannosidic structures using the L3 antibody directed against high mannose‐type glycans in an anti‐idiotypic approach. A selected monoclonal anti‐idiotype antibody was used for affinity chromatography and identified basigin as a binding protein from mouse brain detergent lysates. Basigin was found to bind to high mannose‐carrying cell recognition molecules, such as myelin‐associated glycoprotein, L1, the β2‐subunit of Na+/K+‐ATPase and an oligomannosidic neoglycolipid. Furthermore, basigin was involved in outgrowth of astrocytic processes in vitro. A striking homology between the first immunoglobulin (Ig)‐like domain of basigin and the fourth Ig‐like domain of NCAM, previously shown to bind to oligomannosidic glycans, and the lectin domain of the mannose receptor confirms that basigin is an oligomannose binding lectin. To our knowledge this is the first report that anti‐idiotypic antibodies can be used to identify binding partners for carbohydrates.
HIV‐Tat protein induces oxidative and inflammatory pathways in brain endothelium
Authors: Toborek Michal , Lee Yong Woo , Pu Hong , Malecki Andrzej , Flora Govinder , Garrido Rosario , Hennig Bernhard , Bauer Hans‐Christian , Nath Avindra
Published: March 2003

Impaired function of the brain vasculature might contribute to the development of HIV‐associated dementia. For example, injury or dysfunction of brain microvascular endothelial cells (BMEC) can lead to the breakdown of the blood–brain barrier (BBB) and thus allow accelerated entry of the HIV‐1 virus into the CNS. Mechanisms of injury to BMEC during HIV‐1 infection are not fully understood, but the viral gene product Tat may be, at least in part, responsible for this effect. Tat can be released from infected perivascular macrophages in the CNS of patients with AIDS, and thus BMEC can be directly exposed to high concentrations of this protein. To study oxidative and inflammatory mechanisms associated with Tat‐induced toxicity, BMEC were exposed to increasing doses of Tat1−72, and markers of oxidative stress, as well as redox‐responsive transcription factors such as nuclear factor‐κB (NF‐κB) and activator protein‐1 (AP‐1), were measured. Tat1−72 treatment markedly increased cellular oxidative stress, decreased levels of intracellular glutathione and activated DNA binding activity and transactivation of NF‐κB and AP‐1. To determine if Tat1−72 can stimulate inflammatory responses in brain endothelium in vivo, expression of monocyte chemoattractant protein‐1 (MCP‐1), an NF‐κB and AP‐1‐dependent chemokine, was studied in brain tissue in mice injected with Tat1−72 into the right hippocampus. Tat1−72 markedly elevated the MCP‐1 mRNA levels in brain tissue. In addition, a double immunohistochemistry study revealed that MCP‐1 protein was markedly overexpressed on brain vascular endothelium. These data indicate that Tat1−72 can induce redox‐related inflammatory responses both in in vitro and in vivo environments. These changes can directly lead to disruption of the BBB. Thus, Tat can play an important role in the development of detrimental vascular changes in the brains of HIV‐infected patients.
Corticotropin‐releasing hormone induces proliferation and TNF‐α release in cultured rat microglia via MAP kinase signalling pathways
Authors: Wang Wei , Ji Ping , Dow Kimberly E.
Published: March 2003

We have previously demonstrated that corticotropin‐releasing hormone (CRH) receptor 1 (CRH‐R1) is functionally expressed in rat microglia. In the present study, we show that CRH, acting on CRH‐R1, promoted cell proliferation and tumour necrosis factor‐α (TNF‐α) release in cultured rat microglia. Exogenous CRH resulted in an increase in BrdU incorporation compared with control cells, which was observed in a range of concentrations of CRH between 10 and 500 nm, with a maximal response at 50 nm. The effect of CRH on BrdU incorporation was inhibited by a CRH antagonist astressin but not by a cAMP‐dependent protein kinase inhibitor H89. Exposure of microglial cells to CRH resulted in a transient and rapid increase in TNF‐α release in a dose–dependent manner. In the presence of astressin, the effects of CRH on TNF‐α release were attenuated. CRH effects on TNF‐α release were also inhibited by specific inhibitors of MEK, the upstream kinase of the extracellular signal‐regulated protein kinase (ERK) (PD98059) or p38 mitogen‐activated protein kinase (SB203580), but not by H89. Furthermore, CRH induced rapid phosphorylation of ERK and p38 kinases. Astressin, PD98059, and SB230580 were able to inhibit CRH‐induced kinase phosphorylation. These results suggest that CRH induces cell proliferation and TNF‐α release in cultured microglia via MAP kinase signalling pathways, thereby providing insight into the interactions between CRH and inflammatory mediators.
Alterations in inducible 72‐kDa heat shock protein and the chaperone cofactor BAG‐1 in human brain after head injury
Authors: Seidberg Neal A. , Clark Robert S. B. , Zhang Xiaopeng , Lai Yichen , Chen Minzhi , Graham Steven H. , Kochanek Patrick M. , Watkins Simon C. , Marion Donald W.
Published: March 2003

The stress response in injured brain is well characterized after experimental ischemic and traumatic brain injury (TBI); however, the induction and regulation of the stress response in humans after TBI remains largely undefined. Accordingly, we examined injured brain tissue from adult patients (n = 8) that underwent emergent surgical decompression after TBI, for alterations in the inducible 72‐kDa heat shock protein (Hsp70), the constitutive 73‐kDa heat shock protein (Hsc70), and isoforms of the chaperone cofactor BAG‐1. Control samples (n = 6) were obtained postmortem from patients dying of causes unrelated to CNS trauma. Western blot analysis showed that Hsp70, but not Hsc70, was increased in patients after TBI versus controls. Both Hsp70 and Hsc70 coimmunoprecipitated with the cofactor BAG‐1. The 33 and 46, but not the 50‐kDa BAG‐1 isoforms were increased in patients after TBI versus controls. The ratio of the 46/33‐kDa isoforms was increased in TBI versus controls, suggesting negative modulation of Hsp70/Hsc70 protein refolding activity in injured brain. These data implicate induction of the stress response and its modulation by the chaperone cofactor and Bcl‐2 family member BAG‐1, after TBI in humans.
Characterization of amyloid β peptides from brain extracts of transgenic mice overexpressing the London mutant of human amyloid precursor protein
Authors: Pype Stefan , Moechars Dieder , Dillen Lieve , Mercken Marc
Published: March 2003

Alzheimer's disease (AD) is marked by the presence of neurofibrillary tangles and amyloid plaques in the brain of patients. To study plaque formation, we report on further quantitative and qualitative analysis of human and mouse amyloid β peptides (Aβ) from brain extracts of transgenic mice overexpressing the London mutant of human amyloid precursor protein (APP). Using enzyme‐linked immunosorbant assays (ELISAs) specific for either human or rodent Aβ, we found that the peptides from both species aggregated to form plaques. The ratios of deposited Aβ1–42/1–40 were in the order of 2–3 for human and 8–9 for mouse peptides, indicating preferential deposition of Aβ42. We also determined the identity and relative levels of other Aβ variants present in protein extracts from soluble and insoluble brain fractions. This was done by combined immunoprecipitation and mass spectrometry (IP/MS). The most prominent peptides truncated either at the carboxyl‐ or the amino‐terminus were Aβ1–38 and Aβ11–42, respectively, and the latter was strongly enriched in the extracts of deposited peptides. Taken together, our data indicate that plaques of APP‐London transgenic mice consist of aggregates of multiple human and mouse Aβ variants, and the human variants that we identified were previously detected in brain extracts of AD patients.
Chromaffin cell death induced by 6‐hydroxydopamine is independent of mitochondrial swelling and caspase activation
Authors: Galindo María F. , Jordán Joaquín , González‐García Carmen , Ceña Valentín
Published: March 2003

Our results provide evidence that 6‐hydroxydopamine induced, after auto‐oxidation, toxic levels of hydrogen peroxide (H2O2) that caused bovine chromaffin cell toxicity and death. 6‐Hydroxydopamine (6‐OHDA) treatment markedly reduced, in a dose–response fashion, chromaffin cell viability. Cell death was accompanied by cell shrinkage, nuclear condensation and DNA degradation. Under our experimental conditions, 6‐OHDA auto‐oxidation formed quinones and reactive oxygen species (ROS) that mainly contributed to 6‐OHDA‐induced cytotoxicity in bovine chromaffin cells. Accordingly, different antioxidants, including catalase, vitamin E, Mn(IIItetrakis(4‐benzoic acid)porphyrin chloride (MnTBAP) or ascorbic acid, provided protection against 6‐OHDA‐induced toxicity. Further evidence that 6‐OHDA induces oxidative stress is provided by the fact that this compound decreased total mitochondrial reduced NAD(P)H levels. Our results also suggest that mitochondrial swelling and caspase activation do not play a direct role in 6‐OHDA‐induced death in bovine chromaffin cells.
Pattern of levodopa‐induced striatal changes is different in normal and MPTP‐lesioned mice
Authors: Gross Christian E. , Ravenscroft Paula , Dovero Sandra , Jaber Mohamed , Bioulac Bernard , Bezard Erwan
Published: March 2003

While levodopa‐induced neurochemical changes have been studied in animal models of Parkinson's disease, very little is known regarding the effects of levodopa administration in normal animals. The present study investigates the effects normal and MPTP‐lesioned mice chronically treated with two different doses of levodopa. We assess changes in striatal dopamine (DA) receptor binding, striatal DA receptor mRNA levels and striatal neuropeptide precursor levels (preproenkephalin‐A [PPE‐A]; preprotachykinin [PPT]; preproenkephalin‐B [PPE‐B]). The extent of the lesion was measured by striatal DA transporter binding and stereological estimation of the number of tyrosine hydroxylase immunoreactive neurones in the substantia nigra pars compacta (SNc). In non‐lesioned animals, chronic levodopa treatment induced an increase in PPE‐A mRNA, whereas both D3R binding and PPE‐B mRNA levels were dramatically increased in the lesioned animals in a dose dependent manner. The present results show that chronic levodopa administration may induce pathophysiological changes, even in the absence of a lesion of the nigro‐striatal pathway, suggesting that the sensitization process involves predominantly the indirect striatofugal pathway in non‐lesioned animals, whereas the direct pathway is primarily involved in lesioned animals.
Site‐directed mutagenesis of rabbit LAT1 at amino acids 219 and 234
Authors: Boado Ruben J. , Li Jian Yi , Pardridge William M.
Published: March 2003

The availability of amino acids in the brain is regulated by the blood–brain barrier (BBB) large neutral amino acid transporter type 1 (LAT1) isoform, which is characterized by a high affinity (low Km) for substrate large neutral amino acids. The hypothesis that brain amino acid transport activity can be altered with single nucleotide polymorphisms was tested in the present studies with site‐directed mutagenesis of the BBB LAT1. The rabbit has a high Km LAT1 large neutral amino acid transporter, as compared to the low Km neutral amino acid transporter at the human or rat BBB. The rabbit LAT1 was cloned from a rabbit brain capillary cDNA library. Alignment of the amino acid sequences of rabbit, human, and rat LAT1 revealed two radical amino acid residues that differ in the rabbit relative to the rat or human LAT1. The G219D mutation had a modest effect on the Km and Vmax of tryptophan transport via cloned rabbit LAT1 in frog oocytes, but the W234L variant reduced the Km by 64% and the Vmax by 96%. Conversely, LAT1 transport of either tryptophan or phenylalanine was nearly normalized when the double mutation W234L/G219D variant was produced. These studies show that marked changes in the affinity and capacity of the LAT1 are caused by single nucleotide polymorphisms and that phenotype can be restored with a double mutation.
cAMP‐induced differentiation of human neuronal progenitor cells is mediated by nuclear fibroblast growth factor receptor‐1 (FGFR1)
Authors: Stachowiak E. K. , Fang X. , Myers J. , Dunham S. , Stachowiak M. K.
Published: March 2003

Activation of cAMP signaling pathway and its transcriptional factor cyclic AMP response element binding protein (CREB) and coactivator are key determinants of neuronal differentiation and plasticity. We show that nuclear fibroblast growth factor receptor‐1 (FGFR1) mediates cAMP‐induced neuronal differentiation and regulates CREB and CREB binding protein (CBP) function in α‐internexin‐expressing human neuronal progenitor cells (HNPC). In proliferating HNPC, FGFR1 was associated with the cytoplasm and plasma membrane. Treatment with dB‐cAMP induced nuclear accumulation of FGFR1 and caused neuronal differentiation, accompanied by outgrowth of neurites expressing MAP2 and neuron‐specific neurofilament‐L protein and enolase. HNPC transfected with nuclear/cytoplasmic FGFR1 or non‐membrane FGFR1(SP‐/NLS), engineered to accumulate exclusively in the cell nucleus, underwent neuronal differentiation in the absence of cAMP stimulation. In contrast, FGFR1/R4, with highly hydrophobic transmembrane domain of FGFR4, was membrane associated, did not enter the nucleus and failed to induce neuronal differentiation. Transfection of tyrosine kinase‐deleted dominant negative receptor mutants, cytoplasmic/nuclear FGFR1(TK‐) or nuclear FGFR1(SP‐/NLS)(TK‐), prevented cAMP‐induced neurite outgrowth. Nuclear FGFR1 localized in speckle‐like domains rich in phosphorylated histone 3 and splicing factors, regions known for active RNA transcription and processing, and activated the neurofilament‐L gene promoter. FGFR1(SP‐/NLS) transactivated CRE, up‐regulated phosphorylation and transcriptional activity of CREB and stimulated the activity of CBP several‐fold. Thus, cAMP‐induced nuclear accumulation of FGFR1 provides a signal that triggers molecular events leading to neuronal differentiation.
Interaction between adult patients’ family members and nursing staff on a hospital ward
Authors: Åstedt‐Kurki Päivi , Paavilainen Eija , Tammentie Tarja , Paunonen‐Ilmonen Marita
Published: March 2003

Interaction between adult patients’ family members and nursing staff on a hospital ward

The purpose of this study was to generate knowledge of the interaction between an adult patient’s family members and nursing staff from the staff’s perspective.

Data were collected from nursing staff (n=155) working on the wards and out‐patient departments for pulmonary, rheumatic, neurological and gastroenterological diseases at a university hospital by using a new questionnaire based on earlier research and the literature. The questions explored the staff’s views of interaction with the adult patient’s family members. In this study, interaction is seen as an umbrella concept which encompasses giving information to relatives, discussion, contacts between staff and significant others and working together. The instrument included questions about personal and telephone discussions, the provision of written instructions and factors facilitating and complicating interaction. The response rate was 55%. The data were analysed using SPSS software and examined using frequency and percentage distributions and cross‐tabulation. The open‐ended questions were analysed using qualitative content analysis by reducing, grouping and abstracting the data inductively.

Discussions with relatives while they visited the patient in hospital were the commonest form of interaction. The majority of respondents perceived the interaction with the patient and knowing his or her family members as important. Less than one‐fourth of the respondents started discussion with family members, while the majority expected family members to initiate interaction. The majority of respondents perceived the patient’s presence in discussion as important, but sometimes they thought it was necessary to discuss with family members without the patient. The staff discussed with family members mainly in the ward office or in patient rooms, which were, however, not perceived as peaceful. Discussions primarily pertained to the patient’s condition, discharge from hospital and planning of continued treatment.

The majority of respondents reported that the opportunity for discussion offered by staff, the patient’s positive attitude towards discussion with the family and family members’ own interest facilitated interaction. The staff’s haste and shift‐work, family members’ shyness of approaching the staff and the absence of a peaceful place for discussion serve to complicate interaction.

Family members are primarily seen as informants and recipients of information. The staff’s ability to recognize the importance of family members to successful patient care is relatively adequate. However, insufficient attention is paid to family members and their need for support.
Active learning of geriatric rehabilitation: deliberations of an undergraduate occupational therapy programme
Authors: Chung Jenny C.C.
Published: March 2003

Active learning of geriatric rehabilitation: deliberations of an undergraduate occupational therapy programme

Occupational therapists working in geriatric rehabilitation must possess skills of self‐directed learning and critical thinking, but conventional teaching–learning strategies are limited in acquiring the said skills. This action research aimed at developing in students an ability for active learning and problem solving. An inquiry‐based learning (IBL) methodology was introduced to two geriatric‐related subjects of an undergraduate occupational therapy (OT) programme. Students worked on four real‐life problems in tutorials and participated in service learning. Lectures were carefully structured to consolidate theoretical knowledge generated from the problems. This study was evaluated qualitatively through interviews of both students and clinical educators, and reflective journals. Students found the problems and service learning stimulating enough to encourage active learning and develop basic study skills. Likewise, clinical educators noted students to be more competent when working with geriatric clients than former students who learned through a conventional curriculum. However, the IBL methodology was stressful in areas related to consistency of teaching–learning strategies across curriculum, workload and time constraints. Curriculum review, development of resource files and provision of systematic support are some of the recommendations to the challenges encountered. To conclude, the IBL approach provides appropriate learning environments that foster in students a self‐directed learning attitude in the study of geriatric rehabilitation.
The accuracy of patient records in Swedish nursing homes: congruence of record content and nurses’ and patients’ descriptions
Authors: Ehrenberg Anna , Ehnfors Margareta
Published: March 2003

The accuracy of patient records in Swedish nursing homes: congruence of record content and nurses’ and patients’ descriptions

Data from patient records will increasingly be used for care planning, quality assessment, research, health planning and allocation of resources. Knowledge about the accuracy of such secondary data, however, is limited and only a few studies have been conducted on the accuracy of nursing recording. The aim of this study was to analyse the concordance between the nursing documentation in nursing homes and descriptions of some specific problems of nurses and patients. Comparisons were made between wards where nurses had received training in structured recording based on the nursing process (study group) and wards where no intervention had taken place (reference group). Data were collected from the patient records of randomly selected nursing home residents (n=85). The methods used were audits of patient records and structured interviews with residents and nurses. The study revealed considerable deficiencies in the accuracy of the patient records when the records were compared with the reports from nurses and residents. The overall agreement between the interview data from nurses and from the patient records was low. Concordance was better in the study group as compared with the reference group in which the recorded data were structured only following chronological order. The study unequivocally demonstrates that there are major limitations in using records as a data source for the evaluation, planning and development of care.