bluuebunny/crossref_metadata_2025 huggingface dataset:
Arachidonic acid mediates muscarinic inhibition and enhancement of N-type Ca
2+
current in sympathetic neurons
Authors: Liu Liwang , Rittenhouse Ann R.
Published: January 2003

N-type Ca
2+
channels participate in acute activity-dependent processes such as regulation of Ca
2+
-activated K
+
channels and in more prolonged events such as gene transcription and long-term depression. A slow postsynaptic M
1
muscarinic receptor-mediated modulation of N-type current in superior cervical ganglion neurons may be important in regulating these processes. This slow pathway inhibits N-type current by using a diffusible second messenger that has remained unidentified for more than a decade. Using whole-cell patch–clamp techniques, which isolate the slow pathway, we found that the muscarinic agonist oxotremorine methiodide not only inhibits currents at positive potentials but enhances N-type current at negative potentials. Enhancement was also observed in cell-attached patches. These findings provide evidence for N-type Ca
2+
-current enhancement by a classical neurotransmitter. Moreover, enhancement and inhibition of current by oxotremorine methiodide mimics modulation observed with direct application of a low concentration of arachidonic acid (AA). Although no transmitter has been reported to use AA as a second messenger to modulate any Ca
2+
current in either neuronal or nonneuronal cells, we nevertheless tested whether a fatty acid signaling cascade was involved. Blocking phospholipase C, phospholipase A
2
, or AA but not AA metabolism minimized muscarinic modulation of N-type current, supporting the participation of these molecules in the slow pathway. A role for the G protein G
q
was also confirmed by blocking muscarinic modulation of Ca
2+
currents with anti-G
qα
antibody. Our finding that AA participates in the slow pathway strongly suggests that it may be the previously unknown diffusible second messenger.
Crystal structure of saposin B reveals a dimeric shell for lipid binding
Authors: Ahn Victoria E. , Faull Kym F. , Whitelegge Julian P. , Fluharty Arvan L. , Privé Gilbert G.
Published: January 2003

Saposin B is a small, nonenzymatic glycosphingolipid activator protein required for the breakdown of cerebroside sulfates (sulfatides) within the lysosome. The protein can extract target lipids from membranes, forming soluble protein-lipid complexes that are recognized by arylsulfatase A. The crystal structure of human saposin B reveals an unusual shell-like dimer consisting of a monolayer of α-helices enclosing a large hydrophobic cavity. Although the secondary structure of saposin B is similar to that of the known monomeric members of the saposin-like superfamily, the helices are repacked into a different tertiary arrangement to form the homodimer. A comparison of the two forms of the saposin B dimer suggests that extraction of target lipids from membranes involves a conformational change that facilitates access to the inner cavity.
Natural selection shaped regional mtDNA variation in humans
Authors: Mishmar Dan , Ruiz-Pesini Eduardo , Golik Pawel , Macaulay Vincent , Clark Andrew G. , Hosseini Seyed , Brandon Martin , Easley Kirk , Chen Estella , Brown Michael D. , Sukernik Rem I. , Olckers Antonel , Wallace Douglas C.
Published: January 2003

Human mtDNA shows striking regional variation, traditionally attributed to genetic drift. However, it is not easy to account for the fact that only two mtDNA lineages (M and N) left Africa to colonize Eurasia and that lineages A, C, D, and G show a 5-fold enrichment from central Asia to Siberia. As an alternative to drift, natural selection might have enriched for certain mtDNA lineages as people migrated north into colder climates. To test this hypothesis we analyzed 104 complete mtDNA sequences from all global regions and lineages. African mtDNA variation did not significantly deviate from the standard neutral model, but European, Asian, and Siberian plus Native American variations did. Analysis of amino acid substitution mutations (nonsynonymous, Ka) versus neutral mutations (synonymous, Ks) (k
a
/k
s
) for all 13 mtDNA protein-coding genes revealed that the
ATP6
gene had the highest amino acid sequence variation of any human mtDNA gene, even though ATP6 is one of the more conserved mtDNA proteins. Comparison of the k
a
/k
s
ratios for each mtDNA gene from the tropical, temperate, and arctic zones revealed that ATP6 was highly variable in the mtDNAs from the arctic zone, cytochrome
b
was particularly variable in the temperate zone, and cytochrome oxidase I was notably more variable in the tropics. Moreover, multiple amino acid changes found in ATP6, cytochrome
b
, and cytochrome oxidase I appeared to be functionally significant. From these analyses we conclude that selection may have played a role in shaping human regional mtDNA variation and that one of the selective influences was climate.
Phenotypic effects of familial amyotrophic lateral sclerosis mutant
Sod
alleles in transgenic
Drosophila
Authors: Mockett Robin J. , Radyuk Svetlana N. , Benes Judith J. , Orr William C. , Sohal Rajindar S.
Published: January 2003

A subset of patients suffering from familial amyotrophic lateral sclerosis (FALS) exhibit point mutations in the gene encoding Cu-Zn superoxide dismutase [superoxide:superoxide oxidoreductase, EC
1.15.1.1
(SOD)]. The human wild-type and five FALS
Sod
mutant transgenes were introduced into the fruit fly,
Drosophila melanogaster
, in a Cu-Zn
Sod
null background.
Sod
null flies had dramatically decreased life span, glutathione and methionine content, fertility, locomotor activity, and resistance to hyperoxic stress, compared with wild-type controls. All of these phenotypic manifestations were rescued fully by a single human wild-type allele, expressing 5–10% of wild-type SOD activity. Full recovery of wild-type life span was also observed when human mutant and wild-type alleles were placed together in the fly
Sod
null background. The FALS
Sod
mutations alone caused a recessive phenotype, usually involving low or undetectable levels of SOD activity, in which: (
i
) full restoration of the wild-type phenotype was observed among young adults, and (
ii
) older adults exhibited a sudden increase in oxidative stress, accompanied by physiological impairment of abrupt onset, and followed by premature death. Thus, the minimal SOD activity associated with the FALS
Sod
mutations appears to determine longevity, not by chronically increasing oxidative stress, but by limiting the time in which a viable redox environment can be maintained. However, the dominant gain of function by mutant SOD, which occurs in human patients and in the transgenic mouse model of FALS, is not observed in
Drosophila
.
Mitotic clonal expansion: A synchronous process required for adipogenesis
Authors: Tang Qi-Qun , Otto Tamara C. , Lane M. Daniel
Published: January 2003

When induced to differentiate, growth-arrested 3T3-L1 preadipocytes synchronously reenter the cell cycle and undergo mitotic clonal expansion (MCE) followed by expression of genes that produce the adipocyte phenotype. The preadipocytes traverse the G
1
/S checkpoint synchronously as evidenced by the expression/activation of cdk2-cyclin-E/A, turnover of p27/kip1, hyperphosphorylation of Rb, translocation of cyclin D
1
from nuclei to cytoplasm and GSK-3β from cytoplasm to nuclei, and incorporation of [
3
H]thymidine into DNA. As the cells cross the G
1
/S checkpoint, C/EBPβ acquires DNA-binding activity, initiating a cascade of transcriptional activation that culminates in the expression of adipocyte proteins. The mitogen-activated protein kinase/extracellular signal-regulated kinase kinase (MEK) inhibitor PD98059 delays, but does not block, MCE and differentiation, the extent of the delay causing a comparable delay in the expression of cell-cycle markers, MCE, and adipogenesis. The more potent and specific MEK inhibitor UO126 and the cyclin-dependent kinase inhibitor roscovitine, which inhibit the cell cycle at different points, block MCE, expression of cell cycle and adipocyte markers, as well as adipogenesis. These results show that MCE is a prerequisite for differentiation of 3T3-L1 preadipocytes into adipocytes.
Chemical nature of nitric oxide storage forms in rat vascular tissue
Authors: Rodriguez Juan , Maloney Ronald E. , Rassaf Tienush , Bryan Nathan S. , Feelisch Martin
Published: January 2003

Endothelial NO production results in local formation of adducts that may act as storage forms of NO. Because little is known about their chemical nature, concentrations, and possible role in vascular biology, we sought to characterize those species basally present in rat aorta, using two independent approaches. In the first approach, tissue homogenates were analyzed by using chemiluminescence- and ion-chromatography-based techniques that allow trace-level quantification of NO-related compounds in complex biological matrices. In the second approach, NO stores were characterized by their ability to release NO when illuminated with light and subsequently relax vascular smooth muscle (photorelaxation). The latter included a careful assessment of action spectra for photorelaxation, taking into account the light-scattering properties of the tissue and the storage depletion rates induced by exposure to controlled levels of light. Biochemical analyses revealed that aortic tissues contained 10 ± 1 μM nitrite, 42 ± 7 μM nitrate, 40 ± 6 nM
S
-nitroso, and 33 ± 6 nM
N-
nitroso compounds (
n
= 4–8). The functional data obtained suggest that the NO photolytically released in the tissue originated from species with photophysical properties similar to those reported for low-molecular-weight
S
-nitrosothiols, as well as from nitrite. The relative contribution of these potential NO stores to the extent of photorelaxation was consistent with their concentrations detected biochemically in vascular tissue when their photoactivity was taken into account. We conclude that intravascular nitroso species and nitrite both have the potential to release physiologically relevant quantities of NO independent of enzymatic control by NO synthase.
Addition of the keto functional group to the genetic code of
Escherichia
coli
Authors: Wang Lei , Zhang Zhiwen , Brock Ansgar , Schultz Peter G.
Published: January 2003

Although the keto group is the most versatile of the functional groups in organic chemistry, it is absent in the genetically encoded amino acids. To overcome this natural limitation on protein biosynthesis, we have evolved an orthogonal tRNA-synthetase pair that makes possible the efficient incorporation of a keto amino acid,
p
-acetyl-
l
-phenylalanine, into proteins in
E. coli
with high translational fidelity in response to the amber nonsense codon. To demonstrate the utility of this keto amino acid, we have used it to modify a protein selectively with a small molecule fluorophore and biotin derivative. This additional genetically encoded amino acid should greatly expand our ability to manipulate protein structure and function both
in vitro
and in living cells.
Positive and negative regulation of APP amyloidogenesis by sumoylation
Authors: Li Yonghong , Wang Hui , Wang Su , Quon Diana , Liu Yu-Wang , Cordell Barbara
Published: January 2003

Amyloid β peptide (Aβ) generated from amyloid precursor protein (APP) is central to Alzheimer's disease (AD). Signaling pathways affecting APP amyloidogenesis play critical roles in AD pathogenesis and can be exploited for therapeutic intervention. Here, we show that sumoylation, covalent modification of cellular proteins by small ubiquitin-like modifier (SUMO) proteins, regulates Aβ generation. Increased protein sumoylation resulting from overexpression of SUMO-3 dramatically reduces Aβ production. Conversely, reducing endogenous protein sumoylation with dominant-negative SUMO-3 mutants significantly increases Aβ production. We also show that mutant SUMO-3, K11R, which can only be monomerically conjugated to target proteins, has an opposite effect on Aβ generation to that by SUMO-3, which can form polymeric chains on target proteins. In addition, SUMO-3 immunoreactivity is predominantly detected in neurons in brains from AD, Down's syndrome, and nondemented humans. Therefore, polysumoylation reduces whereas monosumoylation or undersumoylation enhances Aβ generation. These findings provide a regulatory mechanism in APP amyloidogenesis and suggest that components in the sumoylation pathway may be critical in AD onset or progression.
Identification of bile acid precursors as endogenous ligands for the nuclear xenobiotic pregnane X receptor
Authors: Goodwin Bryan , Gauthier Karine C. , Umetani Michihisa , Watson Michael A. , Lochansky Matthew I. , Collins Jon L. , Leitersdorf Eran , Mangelsdorf David J. , Kliewer Steven A. , Repa Joyce J.
Published: January 2003

Sterol 27-hydroxylase (CYP27A1) is required for bile acid synthesis by both the classical and alternate pathways.
Cyp27a1
−/−
mice exhibit a dramatic increase in the activity of cytochrome P450 3A (CYP3A), which catalyzes side-chain hydroxylations of bile acid intermediates, thereby facilitating their excretion in the bile and urine. We examine the role of the nuclear xenobiotic receptor PXR (pregnane X receptor) in this process. We demonstrate that expression of
Cyp3a11
and other established PXR target genes is increased in the
Cyp27a1
−/−
mice. When
Cyp27a1
−/−
mice are fed a diet containing either cholic acid or chenodeoxycholic acid, expression of CYP7A1, which catalyzes the rate-limiting step in bile acid biosynthesis, is strongly suppressed. In parallel, the induction of
Cyp3a11
observed in these mice is reversed, suggesting that bile acid intermediates serve as PXR activators. In support of this hypothesis, three potentially toxic sterols (7α-hydroxy-4-cholesten-3-one, 5β-cholestan-3α,7α,12α-triol, and 4-cholesten-3-one), including two that are known to accumulate in
Cyp27a1
−/−
mice, are efficacious activators of mouse PXR. All three compounds are more potent activators of mouse PXR than of human PXR, which may explain in part why humans who lack functional CYP27A1 do not display a corresponding increase in CYP3A activity and are stricken with the disease cerebrotendinous xanthomatosis. Taken together, these results reveal the existence of a feedforward regulatory loop by which potentially toxic bile acid intermediates activate PXR and induce their own metabolism. In addition, this study demonstrates that animal models with alterations in gene expression can be used to identify endogenous ligands for orphan nuclear receptors.
Nitric oxide-induced cellular stress and p53 activation in chronic inflammation
Authors: Hofseth Lorne J. , Saito Shin'ichi , Hussain S. Perwez , Espey Michael G. , Miranda Katrina M. , Araki Yuzuru , Jhappan Chamelli , Higashimoto Yuichiro , He Peijun , Linke Steven P. , Quezado Martha M. , Zurer Irit , Rotter Varda , Wink David A. , Appella Ettore , Harris Curtis C.
Published: January 2003

Free radical-induced cellular stress contributes to cancer during chronic inflammation. Here, we investigated mechanisms of p53 activation by the free radical, NO. NO from donor drugs induced both ataxia-telangiectasia mutated (ATM)- and ataxia-telangiectasia mutated and Rad3-related-dependent p53 posttranslational modifications, leading to an increase in p53 transcriptional targets and a G
2
/M cell cycle checkpoint. Such modifications were also identified in cells cocultured with NO-releasing macrophages. In noncancerous colon tissues from patients with ulcerative colitis (a cancer-prone chronic inflammatory disease), inducible NO synthase protein levels were positively correlated with p53 serine 15 phosphorylation levels. Immunostaining of HDM-2 and p21
WAF1
was consistent with transcriptionally active p53. Our study highlights a pivotal role of NO in the induction of cellular stress and the activation of a p53 response pathway during chronic inflammation.
Canonical histidine kinase activity of the transmitter domain of the ETR1 ethylene receptor from
Arabidopsis
is not required for signal transmission
Authors: Wang Wuyi , Hall Anne E. , O'Malley Ronan , Bleecker Anthony B.
Published: January 2003

Ethylene signaling in plants is mediated by a family of receptors related to bacterial two-component histidine kinases. Of the five members of the
Arabidopsis
ethylene receptor family, members of subfamily I (ETR1 and ERS1) contain completely conserved histidine kinase domains, whereas members of subfamily II (ETR2, EIN4, and ERS2) lack conserved residues thought to be necessary for kinase activity. To examine the role of the conserved histidine kinase domain in receptor signaling,
ers1;etr1
loss-of-function double mutants were generated. The double mutants exhibited a severe constitutive ethylene response phenotype consistent with the negative regulator model for receptor function. The adult
ers1-2;etr1-6
and
ers1-2;etr1-7
phenotypes included miniature rosette size, delayed flowering, and both male and female sterility, whereas etiolated-seedling responses were less affected. Chimeric transgene constructs in which the
ETR1
promoter was used to drive expression of cDNAs for each of the five receptor isoforms were transferred into the
ers1-2;etr1-7
double-mutant plants. Subfamily I constructs restored normal growth, whereas subfamily II constructs failed to rescue the double mutant, providing evidence for a unique role for subfamily I in receptor signaling. However, transformation of either the
ers1-2;etr1-6
or
ers1-2;etr1-7
mutant with a kinase-inactivated
ETR1
genomic clone also resulted in complete restoration of normal growth and ethylene responsiveness in the double-mutant background, leading to the conclusion that canonical histidine kinase activity by receptors is not required for ethylene receptor signaling.
Sequence variations in the public human genome data reflect a bottlenecked population history
Authors: Marth Gabor , Schuler Greg , Yeh Raymond , Davenport Ruth , Agarwala Richa , Church Deanna , Wheelan Sarah , Baker Jonathan , Ward Ming , Kholodov Michael , Phan Lon , Czabarka Eva , Murvai Janos , Cutler David , Wooding Stephen , Rogers Alan , Chakravarti Aravinda , Harpending Henry C. , Kwok Pui-Yan , Sherry Stephen T.
Published: January 2003

Single-nucleotide polymorphisms (SNPs) constitute the great majority of variations in the human genome, and as heritable variable landmarks they are useful markers for disease mapping and resolving population structure. Redundant coverage in overlaps of large-insert genomic clones, sequenced as part of the Human Genome Project, comprises a quarter of the genome, and it is representative in terms of base compositional and functional sequence features. We mined these regions to produce 500,000 high-confidence SNP candidates as a uniform resource for describing nucleotide diversity and its regional variation within the genome. Distributions of marker density observed at different overlap length scales under a model of recombination and population size change show that the history of the population represented by the public genome sequence is one of collapse followed by a recent phase of mild size recovery. The inferred times of collapse and recovery are Upper Paleolithic, in agreement with archaeological evidence of the initial modern human colonization of Europe.
Up-regulation of neuronal calcium sensor-1 (NCS-1) in the prefrontal cortex of schizophrenic and bipolar patients
Authors: Koh Phil Ok , Undie Ashiwel S. , Kabbani Nadine , Levenson Robert , Goldman-Rakic Patricia S. , Lidow Michael S.
Published: January 2003

The delineation of dopamine dysfunction in the mentally ill has been a long-standing quest of biological psychiatry. The present study focuses on a recently recognized group of dopamine receptor-interacting proteins as possible novel sites of dysfunction in schizophrenic and bipolar patients. We demonstrate that the dorsolateral prefrontal cortex in schizophrenia and bipolar cases from the Stanley Foundation Neuropathology Consortium display significantly elevated levels of the D2 dopamine receptor desensitization regulatory protein, neuronal calcium sensor-1. These levels of neuronal calcium sensor-1 were not influenced by age, gender, hemisphere, cause of death, postmortem period, alcohol consumption, or antipsychotic and mood stabilizing medications. The present study supports the hypothesis that schizophrenia and bipolar disorder may be associated with abnormalities in dopamine receptor-interacting proteins.
Convergence and divergence in the mechanism of SNARE binding by Sec1/Munc18-like proteins
Authors: Dulubova Irina , Yamaguchi Tomohiro , Araç Demet , Li Hongmei , Huryeva Iryna , Min Sang-Won , Rizo Josep , Südhof Thomas C.
Published: January 2003

Sec1/Munc18-like (SM) proteins functionally interact with soluble
N
-ethylmaleimide-sensitive factor attachment protein receptors (SNARE) in membrane fusion, but the mechanisms of these interactions differ. In vertebrates, SM proteins that mediate exocytosis (Munc18-1, 18-2, and 18c) bind to the closed conformation of syntaxins 1–4, which requires the N-terminal H
abc
domains and SNARE motifs of these syntaxins. In contrast, SM proteins that mediate Golgi and endoplasmic reticulum fusion (Sly1 and Vps45) bind only to short N-terminal sequences of syntaxins 5, 16, or 18, independently of their H
abc
domains and SNARE motifs. We now show that Munc18-1, Sly1, and Vps45 interact with cognate syntaxins via similar, autonomously folded N-terminal domains, but the syntaxin 5-binding surface of the Sly1 N-terminal domain is opposite to the syntaxin 1-binding surface of the Munc18-1 N-terminal domain. In transfected cells, the N-terminal domain of Sly1 specifically disrupts the structure of the Golgi complex, supporting the notion that the interaction of Sly1 with syntaxin 5 is essential for fusion. These data, together with previous results, suggest that a relatively small N-terminal domain of SM proteins is dedicated to mechanistically distinct interactions with SNAREs, leaving the remaining large parts of SM proteins free to execute their as yet unknown function as effector domains.
Catalysis of electron transfer during activation of O
2
by the flavoprotein glucose oxidase
Authors: Roth Justine P. , Klinman Judith P.
Published: January 2003

Two prototropic forms of glucose oxidase undergo aerobic oxidation reactions that convert FADH
−
to FAD and form H
2
O
2
as a product. Limiting rate constants of
k
cat
/
K
M
(O
2
) = (5.7 ± 1.8) × 10
2
M
−1
⋅s
−1
and
k
cat
/
K
M
(O
2
) = (1.5 ± 0.3) × 10
6
M
−1
⋅s
−1
are observed at high and low pH, respectively. Reactions exhibit oxygen-18 kinetic isotope effects but no solvent kinetic isotope effects, consistent with mechanisms of rate-limiting electron transfer from flavin to O
2
. Site-directed mutagenesis studies reveal that the pH dependence of the rates is caused by protonation of a highly conserved histidine in the active site. Temperature studies (283–323 K) indicate that protonation of His-516 results in a reduction of the activation energy barrier by 6.0 kcal⋅mol
−1
(0.26 eV). Within the context of Marcus theory, catalysis of electron transfer is attributed to a 19-kcal⋅mol
−1
(0.82 eV) decrease in the reorganization energy and a much smaller 2.2-kcal⋅mol
−1
(0.095 eV) enhancement of the reaction driving force. An explanation is advanced that is based on changes in outer-sphere reorganization as a function of pH. The active site is optimized at low pH, but not at high pH or in the H516A mutant where rates resemble the uncatalyzed reaction in solution.
Estradiol differentially regulates lipocalin-type prostaglandin D synthase transcript levels in the rodent brain: Evidence from high-density oligonucleotide arrays andin situhybridization
Authors: Mong Jessica A. , Devidze Nino , Frail Donald E. , O'Connor Lawrence T. , Samuel Manjo , Choleris Elena , Ogawa Sonoko , Pfaff Donald W.
Published: January 2003

Microarrays comprise an efficient approach to discovering large numbers of differentially expressed mRNA transcripts in the CNS resulting from changes in hormonal milieu. We used high-density oligonucleotide microarrays to examine the short- and long-term actions of estradiol (E2) on the transcriptomes from the medial basal hypothalamus and other brain regions of E2-treated (10 μg) adult female mice. Our results have revealed several unanticipated gene regulations. Most striking is lipocalin prostaglandin D2synthase (L-PGDS), which catalyzes the conversion of prostaglandin (PG) H2to PGD2, a neuromodulator involved in a variety of functions, including sleep, pain, and odor responses.In situhybridization revealed significant increases in L-PGDS expression in the arcuate and ventromedial nucleus of the medial basal hypothalamus compared with vehicle controls. The magnitude of these changes is ≈2-fold and suggests a modulatory role for PGD2in E2-controlled neuroendocrine secretions and behaviors. Surprisingly, L-PGDS gene expression is reduced 2-fold after E2treatment in the ventrolateral preoptic area (VLPO), the suspected site of action for the sleep-promoting effects of PGD2. Finally, whereas L-PGDS has been reported to be expressed primarily in oligodendrocytes of the adult rodent brain, we demonstrate, immunocytochemically, that L-PGDS is also expressed in a population of VLPO neurons. Thus, our data suggest the intriguing possibility that E2modulation of L-PGDS plays a role in the regulation of sleep–wake states through hitherto unknown mechanisms in VLPO neurons and through hormone-dependent neuronal-glial cooperation.
Identification of vaccinia virus epitope-specific HLA-A*0201-restricted T cells and comparative analysis of smallpox vaccines
Authors: Drexler Ingo , Staib Caroline , Kastenmüller Wolfgang , Stevanović Stefan , Schmidt Burkhard , Lemonnier François A. , Rammensee Hans-Georg , Busch Dirk H. , Bernhard Helga , Erfle Volker , Sutter Gerd
Published: January 2003

Despite worldwide eradication of naturally occurring variola virus, smallpox remains a potential threat to both civilian and military populations. New, safe smallpox vaccines are being developed, and there is an urgent need for methods to evaluate vaccine efficacy after immunization. Here we report the identification of an immunodominant HLA-A*0201-restricted epitope that is recognized by cytotoxic CD8
+
T cells and conserved among
Orthopoxvirus
species including variola virus. This finding has permitted analysis and monitoring of epitope-specific T cell responses after immunization and demonstration of the identified T cell specificity in an A*0201-positive human donor. Vaccination of transgenic mice allowed us to compare the immunogenicity of several vaccinia viruses including highly attenuated, replication-deficient modified vaccinia virus Ankara (MVA). MVA vaccines elicited levels of CD8
+
T cell responses that were comparable to those induced by the replication-competent vaccinia virus strains. Finally, we demonstrate that MVA vaccination is fully protective against a lethal respiratory challenge with virulent vaccinia virus strain Western Reserve. Our data provide a basis to rationally estimate immunogenicity of safe, second-generation poxvirus vaccines and suggest that MVA may be a suitable candidate.
Comparative Prevalence, Multiplicity, and Progression of Spontaneous and Vinyl Carbamate-Induced Liver Lesions in Five Strains of Male Mice
Authors: Takahashi Kimimasa , Dinse Gregg E. , Foley Julie F. , Hardisty Jerry F. , Maronpot Robert R.
Published: January 2003

The overall and age-specific prevalences and multiplicities of spontaneous and chemically induced hepatocellular neoplasia were compared among male B6D2F1, B6C3F1, C3H (C3H/HeNCrl MTV-), B6CF1, and C57BL/6 (C57BL/6NCrl) mice following a single intraperitoneal injection of 0.03 μM vinyl carbamate (VC)/g body weight or vehicle alone at 15 days of age. Additional groups of B6C3F1, C3H, and C57BL/6 males received 0.15 μM VC/g body weight at 15 days of age. For male B6D2F1, B6C3F1, C3H, B6CF1, and C57BL/6 mice, the estimated overall prevalences (and multiplicities) of hepatocellular adenomas or carcinomas in vehicle controls were 14.1% (0.19), 12.3% (0.15), 8.2% (0.10), 7.2% (0.09), and 2.4% (0.02), respectively. The analogous estimates in the low-dose group were 59.2% (1.19), 72.9% (4.07), 48.6% (1.99), 22.8% (0.29), and 43.9% (0.82). Analogous estimates for B6C3F1, C3H, and C57BL/6 mice in the high-dose group were 45.3% (4.29), 59.7% (6.63), and 46.8% (1.74), respectively. Age-specific multiplicity estimates suggested a progression from altered hepatocellular foci (AHF) to hepatocellular neoplasms. Further evidence of progression was provided by the temporal occurrence of hepatocellular adenomas before carcinomas, and the apparent origination of carcinomas within adenomas. Pulmonary metastases were observed in many of the mice with hepatocellular carcinomas. These findings confirm previous observations of strain differences in liver neoplasm response, suggest a progressive development from AHF to adenomas, and ultimately to carcinomas, and show sensitivity to VC-induced hepatocarcinogenesi s in all 5 strains.
Apparent Mesonephric Duct (Rete Anlage) Origin for Cysts and Proliferative Epithelial Lesions in the Mouse Ovary
Authors: Long Gerald G.
Published: January 2003

Ovaries and adjacent parovarian (mesovarial) tissues of CD-1 mice of various ages were examined to characterize cystic and proliferative epithelial lesions of the ovary and parovarian tissues. Ovaries and adjacent tissues from 6 mice each at approximately 3 and 8 months of age were processed for light microscopy and step-sections (50- μ intervals) of the entire tissue were examined. Tissues were collected from 40 mice each at 16 through 24 months of age and 3 step sections per mouse were examined. Mesonephric duct remnants were found in all mice at 3 and 8 months. Ducts were usually in the mesovarial adipose tissue and near or within the ovarian hilus and were often loosely associated with smooth muscle of the mesovarial ligament. The epithelium of the ducts varied from low cuboidal to columnar, occasional individual cells were ciliated, and small papillary confi gurations of epithelium were occasionally present. Ducts were dilated (> 1 mm) more often in mice at 8 months compared to 3 months, and some were continuou s with cystic spaces within the ovaries. As mice aged (16—24 months), dilation and cystic change in ducts and associated compression of ovarian tissue away from the ovarian hilus became common. The epithelium of dilated ducts was generally fl attened, but foci of cuboidal to columnar epithelium and/or occasional ciliated cells were present. Many ducts contained foci of hyperplastic and/or hypertrophic epithelium, and papillary projections of epithelium were occasionally found. Some of the latter lesions were consistent with a diagnosis of papillary cystadenoma. Hyperplasia of associated fibromuscular stroma was limited to a few apparently extraovarian ducts. The results of this study indicate that remnant mesonephric ductular structures are common in and adjacent to the ovaries of CD-1 mice. As mice age, these ducts become dilated and, in some, the epithelium becomes hyperplastic and/or hyperptrophic. These mesonephric duct remnants appear to be a common source of ovarian and parovarian cyts cysts and epithelial neoplasms of the ovary in mice.
Longitudinal Magnetic Resonance Imaging Quantitation of Rat Liver Regeneration After Partial Hepatectomy
Authors: Hockings Paul D. , Roberts Toby , Campbell Simon P. , Reid David G. , Greenhill Robert W. , Polley Stephen R. , Nelson Paul , Bertram Timothy A. , Kramer Kerstin
Published: January 2003

This report demonstrates the advantages of using a noninvasive soft tissue imaging technique—magnetic resonance imaging (MRI)—to monitor liver regeneration after 70% partial hepatectomy in the rat in a longitudinal manner. Six animals were scanned prior to and on 6 subsequent occasions up to 9 days after surgical removal of the median and left lateral lobes. Within the observed time frame liver volumes were restored to approximately 88% of presurgery values. Final liver volumes correlated well with postmortem liver weights ( R = 0.93). Regeneration is well-quantified empirically by a 4 parameter logistic equation: The rate of regeneration was maximal at 1.5 days, which coincided with the maximum increase of Mitotic Index—a measure of cell proliferation, determined in a subsequent study. Pre- and postpartial hepatectomy measurements remove two potentially confounding unknowns—the presurgery liver volume, and the amount of liver actually excised. 3D reconstructions of the liver effectively illustrate the morphological changes associated with the procedure, and the regrowth of liver tissue.
Characterization of Uterine Leiomyomas in CD-1 Mice Following Developmental Exposure to Diethylstilbestrol (DES)
Authors: Newbold Retha R. , Moore Alicia B. , Dixon Darlene
Published: January 2003

Experimental animal and clinical studies have well established the association of prenatal exposure to diethylstilbestrol (DES) and the subsequent development of reproductive tract abnormalities, including poor reproductive outcome and neoplasia. Overwhelmingly, the focus has been on DES-induced epithelial lesions, particularly vaginal adenosis and adenocarcinoma; however, uterine smooth muscle cells are also recognized as cellular targets of DES. This descriptive report characterizes uterine leiomyoma s that occur in outbred CD-1 mice following exposure to DES prenatally on days 9 to 16 of gestation or on neonatal days 1 to 5. These DES-induced uterine leiomyomas have typical histomorphologic, and some immunohistochemica l characteristics of spontaneously occurring uterine smooth muscle tumors of B6C3F1 mice previously described in our laboratory, and they are also similar to uterine leiomyomas (fibroids) commonly observed in premenopausal women.
An Autopsy Case of Minamata Disease (Methylmercury Poisoning)—Pathological Viewpoints of Peripheral Nerves
Authors: Eto Komyo , Tokunaga Hidehiro , Nagashima Kazuo , Takeuchi Tadao
Published: January 2003

The outbreak of methylmercury poisoning in the geographic areas around Minamata Bay, Kumamoto, Japan in the 1950s has become known as Minamata disease. Based on earlier reports and extensive pathological studies on autopsied cases at the Kumamoto University School of Medicine, destructive lesions in the anterior portion of the calcarine cortex and depletion predominantly of granular cells in the cerebellar cortex came to be recognized as the hallmark and diagnostic yardstick of methylmercury poisoning in humans. As the number of autopsy cases of Minamata disease increased, it became apparent that the cerebral lesion was not restricted to the calcarine cortex but was relatively widespread. Less severe lesions, believed to be responsible for the motor symptoms of Minamata patients, were often found in the precentral, postcentral, and lateral temporal cortices. These patients also frequently presented with signs of sensory neuropathy affecting the distal extremities. Because of few sufficiently comprehensive studies, peripheral nerve degeneration has not been universally accepted as a cause of the sensory disturbances in Minamata patients. The present paper describes both biopsy and autopsy findings of the peripheral nerves in a male fisherman who died at the age of 64 years and showed the characteristic central nervous system lesions of Minamata disease at autopsy. A sural nerve biopsy with electron microscopy performed 1 month prior to his death showed endoneurial fibrosis and regenerated myelin sheaths. At autopsy the dorsal roots and sural nerve showed endoneurial fibrosis, loss of nerve fibers, and presence of Büngner's bands. The spinal cord showed Wallerian degeneration of thefasciculus gracilis (Goll's tract) with relative preservation of neurons in sensory ganglia. These findings support the contention that there is peripheral nerve degeneration in Minamata patients due to toxic injury from methylmercury.
Epithelial-Induced Intrapulpal Denticles in B6C3F1 Mice
Authors: Long Philip H. , Herbert Ronald A.
Published: January 2003

Multiple intrapulpal denticles were observed in maxillary incisors of control and treated B6C3F1 mice used in a chronic inhalation study. Histologically, the denticles originated from multiple small budlike projections emanating from the epithelial sheath, immediately adjacent to the pulp chamber. The denticles were round to ovoid in shape with a central cavity surrounded by tubular dentin. Immature denticles contained epithelial cells within the central cavity, whereas mature denticles were either devoid of cells or contained cell fragments. A layer of columnar odontoblasts surrounded the outer surface of each denticle. Denticles advanced in a coronal direction as the incisors grew. With continued incisor growth, some denticles impacted the tooth wall and were associated with defects in dentinogenesis, altered tooth shape, and microfractures. Some denticles became partly or entirely incorporated into the dentin of the tooth. Intrapulpal denticle formation may represent a previously unidentified alteration that could contribute to the development of dental dysplasia in mice by interfering with normal tooth development and predisposing affected teeth to malformation and biomechanical failure with fracture.
Use of the Spontaneous Tsc2 Knockout (Eker) Rat Model of Hereditary Renal Cell Carcinoma for the Study of Renal Carcinogens
Authors: Mcdorman Kevin S. , Wolf Douglas C.
Published: January 2003

The kidney is a frequent site for chemically induced cancers in rodents and among the 10 most frequent sites for cancer in human patients. Renal cell carcinoma (RCC) is the most frequent upper urinary tract cancer in humans and accounts for 80—85% of malignant renal tumors. Hereditary RCC occurs in Eker rats that are heterozygou s for an insertion mutation in the Tsc2 tumor suppressor gene. The germline mutation renders heterozygous mutants highly susceptible to renal carcinogens. The utility of this model in studying potential renal carcinogens is due to an ordered progression of proliferative renal lesions that can be identified and counted microscopically. The quantitative nature of the model allows for the production of statistically powerful data to understand the relative degree and potency of chemical effects and allow analysis of genetic alterations that may be chemical specific.
A 13-Week Subchronic Intravaginal Toxicity Study of the Novel Broad-Spectrum Anti-HIV and Spermicidal Agent, N-[2-(1-cyclohexenyl)ethyl]-N'-[2-(5-bromopyridyl)]-thiourea (PHI-346) in Mice
Authors: D'Cruz Osmond J. , Waurzyniak Barbara , Uckun Fatih M.
Published: January 2003

The nonnucleoside inhibitor (NNI) of HIV-1 reverse transcriptase, PHI-346 ( N -[2-(1-cyclohexenyl)ethyl]- N'-[2-(5-bromopyridyl)]-thiourea), is a dual-function spermicidal agent with potent anti-HIV activity against drug-sensitive as well as drug-resistant HIV-1 strains with genotypic and phenotypic NNI resistance. PHI-346 was formulated via a lipophilic gel-microemulsion for intravaginal use as a potential dual-function microbicide. To evaluate the toxicity potential of short-term intravaginal exposure to PHI-346, groups of 15 female B6C3F 1 and CD-1 mice were exposed intravaginally to a gel-microemulsion containing 0, 0.5, 1.0, or 2.0% PHI-346, 5 days per week for 13 consecutive weeks. On a molar basis, these concentrations of PHI-346 are 350 to 1,400-times higher than its spermicidal EC50 and nearly 5 × 106 to 2 × 107 times higher than its in vitro anti-HIV IC50. After 13 weeks of intravaginal treatment, B6C3F 1 mice were evaluated for survival, body weight gain, absolute and relative organ weights. Blood was analyzed for hematology and clinical chemistry profiles. Microscopic examination was performed on hematoxylin and eosin-stained tissue sections from each study animal. Placebo control and PHI-346 dosed female CD-1 mice were mated with untreated males in order to evaluate if PHI-346 has any deleterious effects on the reproductive performance. There were no treatment-related mortalities. Mean body weight gain during the dosing period was not reduced by PHI-346 treatment. The hemogram or blood chemistry profiles revealed lack of systemic toxicity following daily intravaginal instillation of PHI-346 for 13 weeks. No clinically significant changes in absolute and relative organ weights were noted in PHI-346 dose groups. Extensive histopathological examination of tissues revealed no treatment-related abnormalities in any of the three PHI-346 dose groups. Repeated intravaginal exposure of CD-1 mice to increasing concentrations of PHI-346 for 13 weeks had no adverse effect on their subsequent reproductive capability, perinatal outcome, growth, and development of offspring. Collectively, these findings demonstrate that repetitive intravaginal administration of PHI-346 at concentrations as high as 1,400-times its spermicidal EC50 and 2 ×107 times its in vitro anti-HIV IC50 was not associated with local or systemic toxicity and did not adversely affect the reproductive performance in mice. PHI-346 may be useful as an active ingredient of a safe vaginal microbicide for prevention of the sexual transmission of multidrug-resistant HIV-1.
The Dilemmas of External Earnings in Public Sector Scientific Research
Authors: Gunasekara Chrys
Published: January 2003

Governments in a number of developed countries have repositioned their public research agencies to support economic objectives and to alleviate fiscal pressures. This has been accomplished through various strategies, including private-public partnerships and purchaser-provider arrangements. In the 1980s, the Australian Government introduced external earnings targets of up to 30% per annum for its public research agencies, as a means of strengthening their responsiveness to national economic needs. A case study of the Commonwealth Scientific and Industrial Research Organization is presented in this article to discuss the impacts of the external earnings policy. Impacts on the type of research being undertaken by public research agencies, the quality of public research-industry relationships, the nature of scientists' work and the role of stakeholders are considered. A number of dilemmas are raised relating to the role of public scientific research.
Early Electrical Communications Technology and Structural Change in the International Political Economy--The Cases of Telegraphy and Radio
Authors: Mcmahon Peter
Published: January 2003

It is increasingly apparent that the roots of current global transformation lie very much in the middle to late nineteenth century and the raft of basic political, economic, socio-cultural and technological changes that occurred at this time. This is mainly because of the development of a set of novel communications technologies that began the information technology-based transformation. This paper briefly reviews the period from 1845 to 1914 to highlight the role of the emergent information technologies of telegraphy and radio in the consolidation of liberal/international forces and then the rise of nationalist military-industrial tendencies. These technologies were primarily concerned with the control of processes associated with the particular forms of politico-economic development prevalent at the time, and as such were of fundamental importance in promoting structural change, including hegemonic transition as Britain was challenged by Germany and the US.
An imperfect match? The structure of the National Curriculum and education for sustainable development
Authors: Chatzifotiou Athanasia
Published: January 2003

The National Curriculum of primary schools in England and Wales sets out the statutory and non‐statutory requirements for teachers. It describes the aims and goals it aspires to achieve and the content of its subjects. In this article I suggest that the statutory and non‐statutory guidelines of the National Curriculum present conceptually different goals in relation to the holistic development of a student. The first section discusses the two basic aims of the curriculum, their recommended implementation and the implications they have upon the curriculum. I specifically emphasize that ‘statutory’ guidelines represent information‐based knowledge and that the ‘nonstatutory’ guidelines treat value‐based issues. This division of roles creates difficulties in implementing the non‐statutory part of the curriculum. In the second section I demonstrate this division by using education for sustainable development as a focal point. I conclude by highlighting important issues for education and, more particularly, for education for sustainable development.
Developing a global dimension in the curriculum
Authors: Graves Jaya
Published: January 2003

Jaya Graves trained as a Rudolph Steiner teacher. She also worked extensively with the peace movement through the 1970s and 80s. She is now co‐ordinator of Southern Voices, a small organization in Manchester committed to presenting views, perspectives and voices of people from Southern or ‘developing’ countries in different contexts where development education (DE) is practised and ‘developing’ issues are discussed. Implicit in this aim is the understanding that many ‘developing’ countries are ‘root cultures’ to children in an intercultural society. An understanding of ‘developing’ countries and issues will, therefore, help us understand our own society as well as the world in which we live.
Joined‐up thinking in theory and practice: the case of healthy schools?
Authors: Morrison Marlene , Harrison Jennifer , Kitson Neil , Wortley Angela
Published: January 2003

This article provides a critical analysis of recent policy initiatives associated with ‘Healthy Schools’. Its specific focus is upon evaluation research conducted by the authors at the School of Education, University of Leicester. The evaluation team investigated healthy school programmes in two Midlands authorities as they related to anticipated accreditation for the National Healthy School Standard (NHSS). Analysis is of findings that sought to identify the most effective approaches for supporting schools and colleges to achieve healthy school status. A specific focus is upon schools' capacity, and those of the health and educational professionals with whom they work, to meet the myriad expectations that are part of the standard. The ‘Healthy School’ is seen as a critical case where connections between health and education are expected to mirror government department interests in ‘joined‐up’ thinking, evaluation strategies and concerns about accountability and performance among schools and local authorities. This is illustrated in the language of the NHSS, where healthy schools are assumed to provide settings for promoting children's and young people's health through the curriculum, the school environment, and in partnership with parents, governors and local stakeholder communities drawn from education, health, social and community services. Curricular themes of citizenship and personal social and health education are being pulled together to provide a revised non‐statutory curriculum for Personal, Social and Health Education (PSHE) and, by September 2002, a new curriculum framework for citizenship. What are the prospects for such an ambitious linking of rhetoric and practice? The article identifies key aspects of the healthy schools challenge, namely the need to ascertain ways in which its development can be simultaneously extended yet sufficiently inclusive to meet the multiple interests of a healthy and educated citizenry, and of schools in which that challenge is supposed to be met. Yet the article also sounds warnings of a standard that, with limited resources and prioritization, might drown under the weight of its own inclusiveness. The concluding counterpoint is an illustrative case‐study of effective practice that pre‐dates yet anticipates proposed curriculum changes.