bluuebunny/crossref_metadata_2025 huggingface dataset:
Simulated effects of low atmospheric CO2 on structure and composition of North American vegetation at the Last Glacial Maximum
Authors: Sharon A. Cowling
Published: March 2003

1. Physiological experiments have indicated that the lower CO2 levels of the last glaciation (200 μmol mol−1) probably reduced plant water‐use efficiency (WUE) and that they combined with increased aridity and colder temperatures to alter vegetation structure and composition at the Last Glacial Maximum (LGM).2. The effects of low CO2 on vegetation structure were investigated using BIOME3 simulations of leaf area index (LAI), and a two‐by‐two factorial experimental design (modern/LGM CO2, modern/LGM climate).3. Using BIOME3, and a combination of lowered CO2 and simulated LGM climate (from the NCAR‐CCM1 model), results in the introduction of additional xeric vegetation types between open woodland and closed‐canopy forest along a latitudinal gradient in eastern North America.4. The simulated LAI of LGM vegetation was 25–60% lower in many regions of central and eastern United States relative to modern climate, indicating that glacial vegetation was much more open than today.5. Comparison of factorial simulations show that low atmospheric CO2 has the potential to alter vegetation structure (LAI) to a greater extent than LGM climate.6. If the magnitude of LAI reductions simulated for glacial North America were global, then low atmospheric CO2 may have promoted atmospheric warming and increased aridity, through alteration of rates of water and heat exchange with the atmosphere.
The fifteenth‐ and twentieth‐century colonization of the Basin of Mexico by the Great‐tailed Grackle (Quiscalus mexicanus)
Authors: Christensen Alexander F.
Published: March 2003

Historical evidence indicates that Great‐tailed Grackles colonized the Basin of Mexico from the Gulf Coast lowlands in the fifteenth century. They were probably assisted by an intentional introduction, but colonization succeeded because of anthropogenic habitat alterations over the previous two centuries. During the Colonial period, grackles withdrew from the Basin, only to recolonize it in recent decades. This withdrawal was also due probably to changes in land use, including drainage of much of the water from the Basin's lakes.
Climatic and biotic controls on annual carbon storage in Amazonian ecosystems
Authors: Tian H. , Melillo J. M. , Kicklighter D. W. , McGuire A. D. , Helfrich Iii J. , Moore Iii B. , Vörösmarty C. J.
Published: March 2003

The role of undisturbed tropical land ecosystems in the global carbon budget is not well understood. It has been suggested that interannual climate variability can affect the capacity of these ecosystems to store carbon in the short term. In this paper, we use a transient version of the Terrestrial Ecosystem Model (TEM) to estimate annual carbon storage in undisturbed Amazonian ecosystems during the period 1980–94, and to understand the underlying causes of the year‐to‐year variations in net carbon storage for this region.
Effect of global atmospheric carbon dioxide on glacial–interglacial vegetation change
Authors: Bennett K. D. , Willis K. J.
Published: March 2003

Global vegetation changes at the time‐scale of the Earth's orbital variations (104–105 years) have been interpreted as a direct effect of consequential climatic changes, especially temperature. At mid‐ and high latitudes, the evidence from fossil data and general circulation models (GCMs) supporting this hypothesis is strong, but at low latitudes there is a major discrepancy. GCMs predict temperature changes that are less than those inferred from palaeoclimatic data, including the plant fossil record. However, changes in atmospheric CO2 concentrations can account for a high proportion of the low‐latitude vegetation change hitherto attributed to temperature change, and may thus explain the discrepancy. The implications of this finding are considerable for understanding patterns of macroevolution and ecosystem development throughout the geological record.
Characterization of chemokine receptor expression and cytokine production in circulating CD4+ T cells from patients with atopic dermatitis: up‐regulation of C‐C chemokine receptor 4 in atopic dermatitis
Authors: Okazaki H. , Kakurai M. , Hirata D. , Sato H. , Kamimura T. , Onai N. , Matsushima K. , Nakagawa H. , Kano S. , Minota S.
Published: March 2003

Background Th2 and Th1 cells have been suggested to express CCR3/CCR4 and CCR5/CXCR3, respectively.

Objective We examined CCR3, CCR4, CCR5 and CXCR3 expression and cytokine production in peripheral blood CD4+ T cells from patients with atopic dermatitis (AD), which has been postulated to be a Th2‐type cell‐mediated disease, and then analysed the possible correlation between these values and the levels of several clinical parameters.

Methods Intracellular cytokine production and chemokine receptor expression in peripheral blood CD4+ T cells from 40 AD patients and 20 sex‐ and age‐matched healthy control subjects were studied by flow cytometry.

Results The frequencies of IL‐4‐ and IL‐13‐producing CD4+ T cells from patients with AD were significantly higher than those from healthy control subjects (IL‐4:3.9 ± 2.1% vs. 1.6 ± 0.7%, P = 0.0005, IL‐13:4.0 ± 2.1% vs. 1.8 ± 0.8%, P = 0.0023), whereas the frequencies of IL‐2‐ and IFN‐γ‐producing CD4+ T cells were significantly decreased in AD patients (IL‐2:38.1 ± 10.3% vs. 51.3 ± 6.3%, P = 0.0003, IFN‐γ: 9.9 ± 3.5% vs. 26.4 ± 4.6%, P < 0.0001). The percentage of CCR4+ cells in CD4+ CD45RO+ T cells in AD patients was significantly higher than that in healthy control subjects (24.4 ± 8.0% vs. 10.9 ± 2.3%, P < 0.0001) and was correlated positively with the total serum IgE, serum lactic dehydrogenase (LDH) level, eosinophil number, eruption score, and IL‐4 and IL‐13 secretion in CD4+ T cells, and inversely with IL‐2 and IFN‐γ secretion in CD4+ T cells. In contrast, CCR3 was not detected on circulating CD4+ T cells even in AD patients. On the other hand, the percentage of CCR5+ or CXCR3+ cells in CD4+ CD45RO+ T cells in AD patients was significantly decreased (CCR5:23.2 ± 7.0% vs. 28.4 ± 5.4%, P = 0.023, CXCR3:29.9 ± 11.4% vs. 38.5 ± 6.7%, P = 0.028) and was positively correlated with eruption score (P < 0.05). Multiple regression analyses showed that the percentage of CCR4 expression highly correlated with serum IgE, LDH, eosinophil number and eruption in AD patients.

Conclusion CCR4+ cells might be involved in the aetiopathogenesis of AD.
Prevalence of symptoms, sensitization to rats, and airborne exposure to major rat allergen (Rat n 1) and to endotoxin in rat‐exposed workers: a cross‐sectional study
Authors: Lieutier‐Colas F. , Meyer P. , Pons F. , Hedelin G. , Larsson P. , Malmberg P. , Pauli G. , De Blay F.
Published: March 2003

Objective To analyse the relation between airborne exposure to major rat allergen and to endotoxins in exclusively rat‐exposed workers and the prevalence of rat‐related symptoms and sensitization.

Methods A total of 113 workers answered a standardized questionnaire on their atopy status, occupational exposure to rats, and possible work‐related symptoms. Specific IgE against rat urinary proteins (RUP) was measured for 73 subjects. Individual airborne exposure to Rat n 1 and endotoxin were determined with static (n = 256) samplings. Rat n 1 was measured with enzyme‐linked immunosorbent assay (ELISA) and endotoxin by the Limulus method.

Results Forty‐four of 113 subjects (38.9%) reported at least one rat‐related symptom: asthma (4.4%), rhinitis (34%) and conjunctivitis (16%). Twelve per cent were sensitized to RUP (specific IgE > 0.35 KU/L). But only 30.8% of all symptomatic subjects were sensitized to rat allergens. Airborne Rat n 1 levels were not related to symptoms in workers. Symptomatic patients not sensitized to rats were exposed to higher endotoxin levels, but airborne exposure to endotoxins did not significantly protect against or increase sensitization to RUP or rat‐related symptoms.

Conclusion Most symptomatic workers were not sensitized to rat allergen; but no significant relation between rat‐related symptoms and endotoxin levels was found. This suggests that more studies are needed to determine causes other than rat allergens or endotoxins that may be responsible for symptoms in rat‐exposed workers.
Atrophoderma of Moulin with preceding inflammation
Authors: Browne Charmaine ,   FRCPC, Fisher Benjamin K. ,   FRCPC
Published: March 2003

A 16‐year‐old Vietnamese man presented to the Dermatology Clinic with a 10‐year history of bizarre brown patches, which initially started as red asymptomatic ‘‘bumps’' on the trunk, upper and lower extremities, and face. His past medical history was significant for hypothyroidism and idiopathic urticaria. He was on Eltroxin for hypothyroidism. The family history was noncontributory.

Physical examination revealed two types of lesion: erythematous, well‐circumscribed papules in a linear configuration along with linear hyperpigmented atrophic patches following Blaschko's lines were noted on the lower extremities (Fig. 1), right upper extremity, right flank (Fig. 2), and right jawline.

Hyperpigmented atrophic macules involving the lower extremities

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Hyperpigmented atrophic macules involving the trunk and upper extremity

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Initial biopsies taken from the papular lesions on the right thigh and right elbow revealed the following changes. The first biopsy showed a slightly thinned epidermis with prominent dilated blood vessels in the superficial dermis. There also appeared to be a slight increase in the amount of collagen in the deep dermis. The findings were reported as in keeping with ‘‘epithelial atrophy.’' The second biopsy from the lesion on the right elbow revealed an acanthotic epidermis. The granular layer was absent in several areas and there was marked overlying parakeratosis. In the dermis, there was a heavy perivascular lymphocytic infiltrate. The appearances were consistent with a psoriasiform dermatitis (Fig. 3). A biopsy taken from the left thigh approximately 18 months later showed slight irregular acanthosis with dermal edema, dilated blood vessels, and a patchy lymphocytic infiltrate. The appearances were compatible with mild inflammation.

Psoriasiform dermatitis

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Reflections on health care consumerism: insights from feminism
Authors: Williamson Charlotte
Published: March 2003

Health care consumerism is a movement concerned with patients’ interests in health care, crucially those that are repressed or partly repressed by dominant interest‐holders. Like feminism, health care consumerism attracts dislike and confusion as well as enthusiasm. But just as the voicing of women’s repressed interests leads to their gradual acceptance by dominant interest‐holders, so does the voicing of patients’ repressed interests.
Public participation and marginalized groups: the community development model
Authors: O’Keefe Eileen , Hogg Christine
Published: March 2003

Objectives To develop ways of reaching house‐bound people and enabling them to give their views in planning and monitoring health and social care.

Strategy HealthLINK – a project based in a community health council – explored ways of involving older house‐bound people in the London Borough of Camden, in planning and monitoring health and social care using community development techniques.

Results HealthLINK set up an infrastructure to enable house‐bound people to have access to information and to enable them to give their views. This resulted in access for health and local authorities to the views of house‐bound older people and increased the self esteem and quality of life of those who became involved.

Conclusions Community development approaches that enable an infrastructure to be established may be an effective way of reaching marginalized communities. However, there are tensions in this approach between the different requirements for public involvement of statutory bodies and of users, and between representation of groups and listening to individual voices.
Public involvement in health care priority setting: an economic perspective
Authors: Roberts Tracy , Bryan Stirling , Heginbotham Chris , McCallum Alison
Published: March 2003

Background Public involvement in health care decision making and priority setting in the UK is being promoted by recent policy initiatives. In 1993, the British Medical Association called for public consultation where rationing of services was to be undertaken. The approach to priority setting advocated by many health economists is the maximization of quality adjusted life years (QALYs). Typically, for a particular health care programme, the QALY calculation takes account of four features: (1) the number of patients receiving the programme, (2) the survival gain, (3) the gain in quality of life and, (4) the probability of treatment success. Only one feature, that relating to quality of life, is based upon public preferences. If the QALY is to be used as a tool for health care resource allocation at a societal level then it should incorporate broader societal preferences.

Methods This study used an interview‐based survey of 91 members of the general public to explore whether the traditional QALY maximization model is a good predictor of public responses to health care priority setting choices.

Results and conclusions Many respondents did not choose consistently in line with a QALY maximization objective and were most influenced by quality of life concerns. There was little support for health care programmes that provided a prognostic improvement but left patients in relatively poor states of health. The level of respondent engagement in the survey exercise was not sensitive to the provision of supporting clinical information.
The use of patients’ stories by self‐help groups: a survey of voluntary organizations in the UK on the register of the College of Health
Authors: Yaphe John , Rigge Marianne , Herxheimer Andrew , McPherson Ann , Miller Rachel , Shepperd Sasha , Ziebland Sue
Published: March 2003

Objective

First‐hand accounts of illness experience are believed to provide important insights for other patients and their carers. We report the results of a survey that explored how patients’ stories are being collected and used by self‐help and voluntary groups.

Methods

The annual College of Health survey contacts 2 458 addresses, which includes many self‐help groups and voluntary associations. A brief questionnaire for the self‐help groups on the register was attached to the summer 1999 survey on behalf of the DIPEx (database of individual patient experience) project.

Results

DIPEx received replies from 309 organizations representing a wide range of interests and conditions. These indicated that 202 (65%) of the groups currently use patients’ stories in various ways. A further 59 (19%) of the groups reported that although they are not currently using them, they would like to in the future. Organizations that use patients’ stories were invited to describe how they use them and provide examples, if applicable. Content analysis of the free text descriptions revealed 22 distinct uses among the 202 organizations using patient stories. The most frequent uses are the inclusion of patient stories in interviews or articles for the group newsletter (74 or 37%) and the use of stories for inclusion in newspaper articles or media broadcasts (31 or 15%). Some form of database of patients’ stories was maintained by 23 groups (12%).

Conclusions

These findings suggest that patients’ stories are widely collected and used to support a wide range of the recognized functions of self‐help and voluntary groups. This is encouraging to the DIPEx project’s efforts to collect and analyse accounts of illness experience, which will be presented with evidence‐based information about the effects of treatments.
Measurement of consumer preference for treatments used to induce labour: a willingness‐to‐pay approach
Authors: Taylor Susan J. , Armour Carol L.
Published: March 2003

Aim

The purpose of the study was to assess the acceptability to consumers of two methods of induction of labour using a willingness‐to‐pay (WTP) approach. The methods compared were amniotomy plus oxytocin and prostaglandin E2 vaginal gel, followed by oxytocin if necessary.

Methods

A description of each method was presented, in questionnaire format, to pregnant women attending a public hospital ante‐natal clinic. Women were asked to choose one of the two treatments, then give a valuation in dollar terms for both their preferred treatment and the alternative.

Results

It was found that 73.7% of patients preferred gel. The mean maximum WTP for amniotomy plus oxytocin was Aus$133 while that for gel was Aus$178 (P=0.0001). Those who chose amniotomy plus oxytocin were WTP 90% more for their preferred treatment compared with the alternative (Aus$180 vs. Aus$95). Similarly, those who preferred gel were WTP 90% more for their preferred treatment compared with the alternative (Aus$222 vs. Aus$119).

Conclusion

Consumers were able to assess drug information provided on the two therapies, make an informed choice and to value that choice. Information obtained in this way, combined with information on costs, could be used in policy decision‐making.
Patients' opinions of the use of psychiatric case‐finding questionnaires in general practice
Authors: Wood Fiona , Pill Roisin , Prior Lindsay , Lewis Glyn
Published: March 2003

Background  The common mental disorders of depression and anxiety often remain undetected in general practice. Psychiatric screening instruments have been recommended to assist detection.

Aim  To assess patients' attitudes towards the use of psychiatric screening questionnaires for common mental disorders within general practice.

Design  Twenty focus groups comprising 127 patients were conducted in Wales, UK. The groups were distinguished by age, sex and locality.

Results  The majority of respondents felt that the use of a screening questionnaire was acceptable and would be willing to complete one within a general practice setting. However, a number of individuals expressed concern about the validity of the kinds of questions asked and the usefulness of the instrument in practice. Respondents also discussed their concerns relating to issues of confidentiality and the doctor's response to the assessment.

Conclusion  The study indicates that lay beliefs towards mental illness, particularly relating to perceived stigma, will continue to pose problems for the routine use of psychiatric case‐finding questionnaires within general practice.
Inactivation of the Na+‐translocating NADH:ubiquinone oxidoreductase from Vibrio alginolyticus by reactive oxygen species
Authors: Steuber Julia , Rufibach Michèle , Fritz Günter , Neese Frank , Dimroth Peter
Published: March 2003

The Na+‐translocating NADH:quinone oxidoreductase (Na+‐NQR) from Vibrio alginolyticus was inactivated by reactive oxygen species. Highest Na+‐NQR activity was observed in anaerobically prepared membranes that exhibited 1 : 1 coupling of NADH oxidation and Q reduction activities (1.6 U·mg−1). Optical and EPR spectroscopy documented the presence of b‐type cytochromes, a [2Fe−2S] cluster and an organic radical signal in anaerobically prepared membranes from V. alginolyticus. It is shown that the [2Fe−2S] cluster previously assigned to the Na+‐NQR originates from the succinate dehydrogenase or the related enzyme fumarate reductase.
Tissue factor pathway inhibitor
Authors: Panteleev Mikhail A. , Zarnitsina Veronica I. , Ataullakhanov Fazoil I.
Published: March 2003

We have analyzed several mathematical models that describe inhibition of the factor VIIa–tissue factor complex (VIIa–TF) by tissue factor pathway inhibitor (TFPI). At the core of these models is a common mechanism of TFPI action suggesting that only the Xa–TFPI complex is the inhibitor of the extrinsic tenase activity. However, the model based on this hypothesis could not explain well all the available experimental data. Here, we show that a good quantitative description of all experimental data could be achieved in a model that contains two more assumptions. The first assumption is based on the hypothesis originally proposed by Baugh et al. [Baugh, R.J., Broze, G.J. Jr & Krishnaswamy, S. (1998) J. Biol. Chem.273, 4378–4386], which suggests that TFPI could inhibit the enzyme–product complex Xa–VIIa–TF. The second assumption proposes an interaction between the X–VIIa–TF complex and the factor Xa–TFPI complex. Experiments to test these hypotheses are suggested.
The role of zinc in the methylation of the coenzyme M thiol group in methanol:coenzyme M methyltransferase from Methanosarcina barkeri
Authors: Krüer Markus , Haumann Michael , Meyer‐Klaucke Wolfram , Thauer Rudolf K. , Dau Holger
Published: March 2003

Methanol:coenzyme M methyltransferase from methanogenic archaea is a cobalamin‐dependent enzyme composed of three different subunits: MtaA, MtaB and MtaC. MtaA is a zinc protein that catalyzes the methylation of coenzyme M (HS‐CoM) with methylcob(III)alamin. We report zinc XAFS (X‐ray absorption fine structure) results indicating that, in the absence of coenzyme M, zinc is probably coordinated by a single sulfur ligand and three oxygen or nitrogen ligands. In the presence of coenzyme M, one (N/O)‐ligand was replaced by sulfur, most likely due to ligation of the thiol group of coenzyme M. Mutations in His237 or Cys239, which are proposed to be involved in ligating zinc, resulted in an over 90% loss in enzyme activity and in distinct changes in the zinc ligands. In the His237→Ala and Cys239→Ala mutants, coenzyme M also seemed to bind efficiently by ligation to zinc indicating that some aspects of the zinc ligand environment are surprisingly uncritical for coenzyme M binding.
Nuclear proteins that bind to metal response element a (MREa) in the Wilson disease gene promoter are Ku autoantigens and the Ku‐80 subunit is necessary for basal transcription of the WD gene
Authors: Oh Won Jun , Kim Eun Kyung , Ko Jung Ho , Yoo Seung Hee , Hahn Si Houn , Yoo Ook‐Joon
Published: March 2003

Wilson disease (WD), an inherited disorder affecting copper metabolism, is characterized by hepatic cirrhosis and neuronal degeneration, which result from toxic levels of copper that accumulate in the liver and brain, respectively. We reported previously that the ≈ 1.3‐kb promoter of the WD gene contains four metal response elements (MREs). Among the four MREs, MREa plays the most important role in the transcriptional activation of the WD promoter. Electrophoretic mobility shift assays (EMSAs) using synthetic MREa and an oligonucleotide containing the binding site for transcription factor Sp1 revealed the presence of nuclear factors that bind specifically to MREa. Two MREa‐binding proteins of 70 and 82 kDa were purified using avidin–biotin affinity chromatography. Amino acid sequences of peptides from each protein were found to be highly homologous to the Ku proteins. Immunoblot analysis and EMSAs showed that the MREa‐binding proteins are immunologically related to the Ku proteins. To study further the functional significance of these Ku‐related proteins in transcriptional regulation of the WD gene, we performed RNA interference (RNAi) assays using a Ku‐80 inverted‐repeat gene to inhibit expression of the Ku‐80 gene in vivo. Results of the RNAi assays showed that expression of the Ku‐80 protein was suppressed in transfected cells, which in turn led to the suppression of the WD gene. In addition, a truncated Ku‐80 (ΔKu‐80) mutant inhibited WD promoter activity in HepG2 cells in a dominant‐negative manner. We also found that WD promoter activity was decreased in Xrs5 cells, which, unlike the CHO‐K1 cells, are defective in the Ku‐80 protein. When Ku‐80 cDNA was transfected into Xrs5 and CHO cells, WD promoter activity was recovered only in Xrs5 cells. Taken together, our findings suggest that the Ku‐80 subunit is required for constitutive expression of the WD gene.
The reductive hotspot hypothesis of mammalian aging
Authors: de Grey Aubrey D. N. J.
Published: March 2003

A severe challenge to the idea that mitochondrial DNA mutations play a major role in the aging process in mammals is that clear loss‐of‐function mutations accumulate only to very low levels (under 1% of total) in almost any tissue, even by very old age. Their accumulation is punctate: some cells become nearly devoid of wild‐type mitochondrial DNA and exhibit no activity for the partly mitochondrially encoded enzyme cytochrome c oxidase. Such cells accumulate in number with aging, suggesting that they survive indefinitely, which is itself paradoxical. The reductive hotspot hypothesis suggests that these cells adjust their metabolism to use plasma membrane electron transport as a substitute for the mitochondrial electron transport chain in the reoxidation of reduced dinucleotides, and that, like mitochondrial electron transport, this process is imperfect and generates superoxide as a side‐effect. This superoxide, generated on the outside of the cell, can potentially initiate classical free radical chemistry including lipid peroxidation chain reactions in circulating material such as lipoproteins. These, in turn, can be toxic to mitochondrially nonmutant cells that import them to satisfy their cholesterol requirements. Thus, the relatively few cells that have lost oxidative phosphorylation capacity may be toxic to the rest of the body. In this minireview, recent results relevant to this hypothesis are surveyed and approaches to intervening in the proposed process are discussed.
Bromoperoxidase activity of vanadate‐substituted acid phosphatases from Shigella flexneri and Salmonella enterica ser. typhimurium
Authors: Tanaka Naoko , Dumay Valérie , Liao Qianning , Lange Alex J. , Wever Ron
Published: March 2003

Vanadium haloperoxidases and the bacterial class A nonspecific acid phosphatases have a conserved active site. It is shown that vanadate‐substituted recombinant acid phosphatase from Shigella flexneri (PhoN‐Sf) and Salmonella enterica ser. typhimurium (PhoN‐Se) in the presence of H2O2 are able to oxidize bromide to hypobromous acid. Vanadate is essential for this activity. The kinetic parameters for the artificial bromoperoxidases have been determined. The Km value for H2O2 is about the same as that for the vanadium bromoperoxidases from the seaweed Ascophyllum nodosum. However, the Km value for Br– is about 10–20 times higher, and the turnover values of about 3.4 min−1 and 33 min−1 for PhoN‐Sf and PhoN‐Se, respectively, are much slower, than those of the native bromoperoxidase. Thus, despite the striking similarity in the active‐site structures of the vanadium haloperoxidases and the acid phophatase, the turnover frequency is low, and clearly the active site of acid phosphatases is not optimized for haloperoxidase activity. Like the native vanadium bromoperoxidase, the vanadate‐substituted PhoN‐Sf and PhoN‐Se catalyse the enantioselective sulfoxidation of thioanisole.
Bass hepcidin is a novel antimicrobial peptide induced by bacterial challenge
Authors: Shike Hiroko , Lauth Xavier , Westerman Mark E. , Ostland Vaughn E. , Carlberg James M. , Van Olst Jon C. , Shimizu Chisato , Bulet Philippe , Burns Jane C.
Published: March 2003

We report the isolation of a novel antimicrobial peptide, bass hepcidin, from the gill of hybrid striped bass, white bass (Morone chrysops) × striped bass (M. saxatilis). After the intraperitoneal injection of Micrococcus luteus and Escherichia coli, the peptide was purified from HPLC fractions with antimicrobial activity against Escherichia coli. Sequencing by Edman degradation revealed a 21‐residue peptide (GCRFCCNCCPNMSGCGVCCRF) with eight putative cysteines. Molecular mass measurements of the native peptide and the reduced and alkylated peptide confirmed the sequence with four intramolecular disulfide bridges. Peptide sequence homology to human hepcidin and other predicted hepcidins, indicated that the peptide is a new member of the hepcidin family. Nucleotide sequences for cDNA and genomic DNA were determined for white bass. A predicted prepropeptide (85 amino acids) consists of three domains: a signal peptide (24 amino acids), prodomain (40 amino acids) and a mature peptide (21 amino acids). The gene has two introns and three exons. A TATA box and several consensus‐binding motifs for transcription factors including C/EBP, nuclear factor‐κB, and hepatocyte nuclear factor were found in the region upstream of the transcriptional start site. In white bass liver, hepcidin gene expression was induced 4500‐fold following challenge with the fish pathogen, Streptococcus iniae, while expression levels remained low in all other tissues tested. A novel antimicrobial peptide from the gill, bass hepcidin, is predominantly expressed in the liver and highly inducible by bacterial exposure.
Chromatin remodeling in nuclear cloning
Authors: Wade Paul A. , Kikyo Nobuaki
Published: March 2003

Nuclear cloning is a procedure to create new animals by injecting somatic nuclei into unfertilized oocytes. Recent successes in mammalian cloning with differentiated adult nuclei strongly indicate that oocyte cytoplasm contains unidentified remarkable reprogramming activities with the capacity to erase the previous memory of cell differentiation. At the heart of this nuclear reprogramming lies chromatin remodeling as chromatin structure and function define cell differentiation through regulation of the transcriptional activities of the cells. Studies involving the modification of chromatin elements such as selective uptake or release of binding proteins, covalent histone modifications including acetylation and methylation, and DNA methylation should provide significant insight into the molecular mechanisms of nuclear dedifferentiation and redifferentiation in oocyte cytoplasm.
Nucleotide excision repair and chromatin remodeling
Authors: Ura Kiyoe , Hayes Jeffrey J.
Published: March 2003

The organization of DNA within eukaryotic cell nuclei poses special problems and opportunities for the cell. For example, assembly of DNA into chromatin is thought to be a principle mechanism by which adventitious general transcription is repressed. However, access to genomic DNA for events such as DNA repair must be facilitated by energy‐intensive processes that either directly alter chromatin structure or impart post‐translational modifications, leading to increased DNA accessibility. The assembly of DNA into chromatin affects both the incidence of damage to DNA and repair of that damage. Correction of most damage to DNA caused by UV irradiation occurs via the nucleotide excision repair (NER) process. NER requires extensive involvement of large multiprotein complexes with relatively large stretches of DNA. Here, we review recent evidence suggesting that at least some steps of NER require ATP‐dependent chromatin remodeling activities while perhaps others do not.
Endogenous cardiac glycosides, a new class of steroid hormones
Authors: Schoner Wilhelm
Published: March 2003

The search for endogenous digitalis has led to the isolation of ouabain as well as several additional cardiotonic steroids of the cardenolide and bufadienolide type from blood, adrenals, and hypothalamus. The concentration of endogenous ouabain is elevated in blood upon increased Na+ uptake, hypoxia, and physical exercise. Changes in blood levels of ouabain upon physical exercise occur rapidly. Adrenal cortical cells in tissue culture release ouabain upon addition of angiotensin II and epinephrine, and it is thought that ouabain is released from adrenal cortex in vivo. Ouabain levels in blood are elevated in 50% of Caucasians with low‐renin hypertension. Infusion over several weeks of low concentrations of ouabain, but not of digoxin, induces hypertension in rats. A digoxin‐like compound, which has been isolated from human urine and adrenals, as well various other endogenous cardiac glycosides may counterbalance their actions within a regulatory framework of water and salt metabolism. Marinobufagenin, for instance, whose concentration is increased after cardiac infarction, may show natriuretic properties because it inhibits the α1 isoform of Na+/K+‐ATPase, the main sodium pump isoform of the kidney, much better than other sodium pump isoforms. In analogy to other steroid hormones, cardiotonic steroid hormones in blood are bound to a specific cardiac glycoside binding globulin. The discovery of ouabain as a new adrenal hormone affecting Na+ metabolism and the development of the new ouabain antagonist PST 2238 allows for new possibilities for the therapy of hypertension and congestive heart failure. This will lead in turn to a better understanding of the disease on a physiological and endocrinological level and of the action of ouabain on the cellular level as a signal that is transduced to the plasma membrane as well as to the cell nucleus.
Role of critical charged residues in reduction potential modulation of ferredoxin‐NADP+ reductase
Authors: Faro Merche , Gómez‐Moreno Carlos , Stankovich Marian , Medina Milagros
Published: March 2003

Reduction potential determinations of K75E, E139K and E301A ferredoxin‐NADP+ reductases provide valuable information concerning the factors that contribute to tune the flavin reduction potential. Thus, while E139 is not involved in such modulation, the K75 side‐chain tunes the flavin potential by creating a defined environment that modulates the FAD conformation. Finally, the E301 side‐chain influences not only the flavin reduction potential, but also the electron transfer mechanism, as suggested from the values determined for the E301A mutant, where Eox/rd and Esq/rd shifted +41 and +102 mV, respectively, with regard to wild‐type. Reduction potentials allowed estimation of binding energies differences of the FAD cofactor upon reduction.
Purification and characterization of novel kininogens from spotted wolffish and Atlantic cod
Authors: Ylönen Anne , Helin Jari , Bøgwald Jarl , Jaakola Anu , Rinne Ari , Kalkkinen Nisse
Published: March 2003

Kininogens are multifunctional proteins found so far mainly in mammals. They carry vasoactive kinins as well as participate in defense, blood coagulation and the acute phase response. In this study, novel kininogens were isolated from Atlantic cod (Gadus morhua L.) and spotted wolffish(Anarhichas minor) by papain‐affinity chromatography. The molecular mass of cod kininogen determined by MALDI‐TOF mass spectrometry to be 51.0 kDa and it had pI values of 3.6, 3.9 and 4.4. The molecular mass of wolffish kininogen was 45.8 kDa and it had pI values of 4.1, 4.3, 4.35 and 4.4. Partial amino‐acid sequences determined from both kininogens showed clear homology with previously determined kininogen sequences. Both kininogens were found to inhibit cysteine proteinases like papain and ficin but they had no effect on trypsin, a serine proteinase. Wolffish kininogen carried α2,3‐sialylated biantennary and triantennary N‐glycans with extensive sialic acid O‐acetylation. Cod kininogen carried similar glycan structures but about 1/3 of its glycans carried sulfate at their N‐acetylglucosamine units.
Thermal stability of peroxidase from the african oil palm tree Elaeis guineensis
Authors: Rodríguez Anabel , Pina David G. , Yélamos Belén , León John J. Castillo , Zhadan Galina G. , Villar Enrique , Gavilanes Francisco , Roig Manuel G. , Sakharov Ivan Yu. , Shnyrov Valery L.
Published: March 2003

The thermal stability of peroxidase from leaves of the African oil palm tree Elaeis guineensis (AOPTP) at pH 3.0 was studied by differential scanning calorimetry (DSC), intrinsic fluorescence, CD and enzymatic assays. The spectral parameters as monitored by ellipticity changes in the far‐UV CD spectrum of the enzyme as well as the increase in tryptophan intensity emission upon heating, together with changes in enzymatic activity with temperature were seen to be good complements to the highly sensitive but integral method of DSC. The data obtained in this investigation show that thermal denaturation of palm peroxidase is an irreversible process, under kinetic control, that can be satisfactorily described by the two‐state kinetic scheme, N  D, where k␣is a first‐order kinetic constant that changes with temperature, as given by the Arrhenius equation; N is the native state, and D is the denatured state. On the basis of this model, the parameters of the Arrhenius equation were calculated.
Mistletoe viscotoxins increase natural killer cell‐mediated cytotoxicity
Authors: Tabiasco Julie , Pont Frédéric , Fournié Jean‐Jacques , Vercellone Alain
Published: March 2003

Mistletoe extracts have immunomodulatory activity. We show that nontoxic concentrations of Viscum album extracts increase natural killer (NK) cell‐mediated killing of tumor cells but spare nontarget cells from NK lysis. The compounds responsible for this bioactivity were isolated from mistletoe and characterized. They have low molecular mass and are thermostable and protease‐resistant. After complete purification by HPLC, they were identified by tandem MS as viscotoxins A1, A2 and A3 (VTA1, VTA2 and VTA3, respectively). Whereas micromolar concentrations of these viscotoxins are cytotoxic to the targets, the bioactivity with respect to NK lysis is within the nanomolar range and differs between viscotoxin isoforms: VTA1 (85 nm), VTA2 (18 nm) and VTA3 (8 nm). Microphysiometry and assays of cell killing indicate that, within such nontoxic concentrations, viscotoxins do not activate NK cells, but act on cell conjugates to increase the resulting lysis.
Thermodynamics and kinetics of the cleavage of DNA catalyzed by bleomycin A5
Authors: Liang Yi , Du Fen , Zhou Bing‐Rui , Zhou Hui , Zou Guo‐Lin , Wang Cun‐Xin , Qu Song‐Sheng
Published: March 2003

Microcalorimetry and UV‐vis spectroscopy were used to conduct thermodynamic and kinetic investigations of the scission of calf thymus DNA catalyzed by bleomycin A5 (BLM‐A5) in the presence of ferrous ion and oxygen. The molar reaction enthalpy for the cleavage, the Michaelis–Menten constant for calf thymus DNA and the turnover number of BLM‐A5 were calculated by a novel thermokinetic method for an enzyme‐catalyzed reaction to be −577 ± 19 kJ·mol−1, 20.4 ± 3.8 µm and 2.28 ± 0.49 × 10−2 s−1, respectively, at 37.0 °C. This DNA cleavage was a largely exothermic reaction. The catalytic efficiency of BLM‐A5 is of the same order of magnitude as that of lysozyme but several orders of magnitude lower than those of TaqI restriction endonuclease, NaeI endonuclease and BamHI endonuclease. By comparing the molar enthalpy change for the cleavage of calf thymus DNA induced by BLM‐A5 with those for the scission of calf thymus DNA mediated by adriamycin and by (1,10‐phenanthroline)‐copper, it was found that BLM‐A5 possessed the highest DNA cleavage efficiency among these DNA‐damaging agents. These results suggest that BLM‐A5 is not as efficient as a DNA‐cleaving enzyme although the cleavage of DNA by BLM‐A5 follows Michaelis–Menten kinetics. Binding of BLM‐A5 to calf thymus DNA is driven by a favorable entropy increase with a less favorable enthalpy decrease, in line with a partial intercalation mode involved in BLM‐catalyzed breakage of DNA.
Escherichia coli small heat shock proteins, IbpA and IbpB, protect enzymes from inactivation by heat and oxidants
Authors: Kitagawa Masanobu , Miyakawa Mizuho , Matsumura Yoshinobu , Tsuchido Tetsuaki
Published: March 2003

To examine functions of two small heat shock proteins of Escherichia coli, IbpA and IbpB, we constructed His–IbpA and His–IbpB, in which a polyhistidine tag was fused to the N‐terminals. Both purified His–IbpA and His–IbpB formed multimers, which have molecular masses of about 2.0–3.0 MDa and consist of about 100–150 subunits. They suppressed the inactivation of several enzymes including citrate synthase and 6‐phosphogluconate dehydrogenase by heat, potassium superoxide, hydrogen peroxide and freeze‐thawing, but not the inactivation of glyceraldehyde‐3‐phosphate dehydrogenase by hydrogen peroxide. Both His–IbpA and His–IbpB suppressed enzyme inactivation by various treatments and were also found to be associated with their non‐native forms. However, both His–IbpA and His–IbpB were not able to reactivate enzymes inactivated by heat, oxidants or guanidine hydrochloride. When heated to 50 °C, each multimeric form of His–IbpA or His–IbpB was dissociated to form a monomer for His–IbpA, and an oligomer of about one‐quarter size for His–IbpB. These structural changes were reversible, as both heated proteins regained the multimeric structures after incubation at 25 °C. However, when exposed to hydrogen peroxide or potassium superoxide, the large multimeric forms of His–IbpA and His–IbpB were maintained. The results suggest that His–IbpA and His–IbpB suppress the inactivation of enzymes and bind non‐native proteins to protect their structures from heat and oxidants.
The trans‐sialidase from the african trypanosome Trypanosoma brucei
Authors: Montagna Georgina , Cremona M. Laura , Paris Gastón , Amaya M. Fernanda , Buschiazzo Alejandro , Alzari Pedro M. , Frasch Alberto C. C.
Published: March 2003

Trypanosoma brucei is the cause of the diseases known as sleeping sickness in humans (T. brucei ssp. gambiense and ssp. rhodesiense) and ngana in domestic animals (T. brucei brucei) in Africa. Procyclic trypomastigotes, the tsetse vector stage, express a surface‐bound trans‐sialidase that transfers sialic acid to the glycosylphosphatidylinositol anchor of procyclin, a surface glycoprotein covering the parasite surface. Trans‐sialidase is a unique enzyme expressed by a few trypanosomatids that allows them to scavenge sialic acid from sialylated compounds present in the infected host. The only enzyme extensively characterized is that of the American trypanosome T. cruzi (TcTS). In this work we identified and characterized the gene encoding the trans‐sialidase from T. brucei brucei (TbTS). TbTS genes are present at a small copy number, at variance with American trypanosomes where a large gene family is present. The recombinant TbTS protein has both sialidase and trans‐sialidase activity, but it is about 10 times more efficient in transferring than in hydrolysing sialic acid. Its N‐terminus contains a region of 372 amino acids that is 45% identical to the catalytic domain of TcTS and contains the relevant residues required for catalysis. The enzymatic activity of mutants at key positions involved in the transfer reaction revealed that the catalytic sites of TcTS and TbTS are likely to be similar, but are not identical. As in the case of TcTS and TrSA, the substitution of a conserved tryptophanyl residue changed the substrate specificity rendering a mutant protein capable of hydrolysing both α‐(2,3) and α‐(2,6)‐linked sialoconjugates.
Toxicity of substrate‐bound amyloid peptides on vascular smooth muscle cells is enhanced by homocysteine
Authors: Mok Su San , Turner Bradley J. , Beyreuther Konrad , Masters Colin L. , Barrow Colin J. , Small David H.
Published: March 2003

Τhe main component of cerebral amyloid angiopathy (CAA) in Alzheimer's disease is the amyloid‐β protein (Aβ), a 4‐kDa polypeptide derived from the β‐amyloid protein precursor (APP). The accumulation of Aβ in the basement membrane has been implicated in the degeneration of adjacent vascular smooth muscle cells (VSMC). However, the mechanism of Aβ toxicity is still unclear. In this study, we examined the effect of substrate‐bound Aβ on VSMC in culture. The use of substrate‐bound proteins in cell culture mimics presentation of the proteins to cells as if bound to the basement membrane. Substrate‐bound Aβ peptides were found to be toxic to the cells and to increase the rate of cell death. This toxicity was dependent on the length of time the peptide was allowed to ‘age’, a process by which Aβ is induced to aggregate over several hours to days. Oxidative stress via hydrogen peroxide (H2O2) release was not involved in the toxic effect, as no decrease in toxicity was observed in the presence of catalase. However, substrate‐bound Aβ significantly reduced cell adhesion compared to cells grown on plastic alone, indicating that cell–substrate adhesion may be important in maintaining cell viability. Aβ also caused an increase in the number of apoptotic cells. This increase in apoptosis was accompanied by activation of caspase‐3. Homocysteine, a known risk factor for cerebrovascular disease, increased Aβ‐induced toxicity and caspase‐3 activation in a dose‐dependent manner. These studies suggest that Aβ may activate apoptotic pathways to cause loss of VSMC in CAA by inhibiting cell–substrate interactions. Our studies also suggest that homocysteine, a known risk factor for other cardiovascular diseases, could also be a risk factor for hemorrhagic stroke associated with CAA.
Differential response of neuronal cells to a fusion protein of ciliary neurotrophic factor/soluble CNTF‐receptor and leukemia inhibitory factor
Authors: März Pia , Özbek Suat , Fischer Martina , Voltz Nicole , Otten Uwe , Rose‐John Stefan
Published: March 2003

Ciliary neurotrophic factor (CNTF) displays neurotrophic activities on motor neurons and neural cell populations both in vivo and in vitro. On target cells lacking intrinsic expression of specific receptor α subunits cytokines of the IL‐6 family only act in the presence of their specific agonistic soluble receptors. Here, we report the construction and expression of a CNTF/soluble CNTF‐receptor (sCNTF‐R) fusion protein (Hyper‐CNTF) with enhanced biological activity on cells expressing gp130 and leukemia inhibitory factor receptor (LIF‐R), but not membrane‐bound CNTF‐R. At the cDNA level, the C‐terminus of the extracellular domain of human CNTF‐R (amino acids 1–346) was linked via a single glycine residue to the N‐terminus of human CNTF (amino acids 1–186). Recombinant Hyper‐CNTF protein was expressed in COS‐7 cells. Hyper‐CNTF efficiently induced dose‐dependent STAT3 phosphorylation and proliferation of BAF‐3 cells stably transfected with gp130 and LIF‐R cDNAs. While on BAF3/gp130/LIF‐R cells, Hyper‐CNTF and LIF exhibited similar biological responses, the activity of Hyper‐CNTF on pheochromocytoma cells (PC12 cells) was quite distinct from that of LIF. In contrast to LIF, Hyper‐CNTF stimulated neurite outgrowth of PC12 cells in a time‐ and dose‐dependent manner correlating with the ability to phosphorylate MAP kinases. These data indicate that although LIF and Hyper‐CNTF use the same heterodimeric receptor complex of gp130 and LIFR, only Hyper‐CNTF induces neuronal differentiation. The therapeutic potential of Hyper‐CNTF as a superagonistic neurotrophin is discussed.
Substrates modulate the rate‐determining step for CO binding in cytochrome P450cam (CYP101)
Authors: Jung Christiane , Bec Nicole , Lange Reinhard
Published: March 2003

The high‐pressure stopped‐flow technique is applied to study the CO binding in cytochrome P450cam (P450cam) bound with homologous substrates (1R‐camphor, camphane, norcamphor and norbornane) and in the substrate‐free protein. The activation volume ΔV # of the CO on‐rate is positive for P450cam bound with substrates that do not contain methyl groups. The kon rate constant for these substrate complexes is in the order of 3 × 106 m−1·s−1. In contrast, P450cam complexed with substrates carrying methyl groups show a negative activation volume and a low kon rate constant of ≈ 3 × 104 m−1·s−1. By relating kon and ΔV # with values for the compressibility and the influx rate of water for the heme pocket of the substrate complexes it is concluded that the positive activation volume is indicative for a loosely bound substrate that guarantees a high solvent accessibility for the heme pocket and a very compressible active site. In addition, subconformers have been found for the substrate‐free and camphane‐bound protein which show different CO binding kinetics.