bluuebunny/crossref_metadata_2025 huggingface dataset:
Exopolysaccharide and Kestose Production by
Lactobacillus sanfranciscensis
LTH2590
Authors: Korakli Maher , Pavlovic Melanie , Gänzle Michael G. , Vogel Rudi F.
Published: April 2003

The effect was investigated of sucrose concentration on sucrose metabolism and on the formation of exopolysaccharide (EPS) by
Lactobacillus sanfranciscensis
LTH2590 in pH-controlled fermentations with sucrose concentrations ranging from 20 to 160 g liter
−1
. The EPS production increased and the relative sucrose hydrolysis activity decreased by increasing the sucrose concentration in the medium. The carbon recovery decreased from 95% at a sucrose concentration of 30 g liter
−1
to 58% at a sucrose concentration of 160 g liter
−1
because of the production of an unknown metabolite by
L. sanfranciscensis
. This metabolite was characterized as a fructo-oligosaccharide. The oligosaccharide produced by
L. sanfranciscensis
was purified and characterized as a trisaccharide with a glucose/fructose ratio of 1:2. The comparison of the retention time of this oligosaccharide and that of pure oligosaccharide standards using two different chromatography methods revealed that the oligosaccharide produced by
L. sanfranciscensis
LTH2590 is 1-kestose. Kestose production increased concomitantly with the initial sucrose concentration in the medium.
PCR-Restriction Fragment Length Polymorphism Analysis of the Phospholipase B (
PLB1
) Gene for Subtyping of
Cryptococcus neoformans
Isolates
Authors: Latouche G. Nicolas , Huynh Matthew , Sorrell Tania C. , Meyer Wieland
Published: April 2003

Cryptococcus neoformans
is a pathogenic yeast that is currently divided into three varieties, five serotypes, and eight molecular types. The following report describes the use of PCR-restriction fragment length polymorphism (RFLP) analysis of the phospholipase B gene (
PLB1
) as a simple tool to differentiate between
C. neoformans
subgroups. A
PLB1
fragment, 1,970 bp, was amplified and digested with either
Ava
I or
Hin
dIII. Both sets of profiles grouped the isolates into their respective varieties, but only the
Ava
I profiles allowed for the identification of the eight molecular types via the corresponding RFLP profiles A1 to A8. Digestion of the same fragments with
Hin
dIII resulted in RFLP profiles H1 to H5, which distinguished only between serotype A, AD, D, and B/C. Neither enzyme distinguished serotype B from serotype C. The serotype AD profile was a composite of the serotype A and D profiles. Further investigation showed that the serotype AD isolates used in this study are heterozygous, with one allele of
PLB1
originating from a serotype A parent and the other from a serotype D parent.
Secretory Antibodies Do Not Affect the Composition of the Bacterial Microbiota in the Terminal Ileum of 10-Week-Old Mice
Authors: Sait Leanne , Galic Maja , Strugnell Richard A. , Janssen Peter H.
Published: April 2003

Terminal restriction fragment length polymorphism (T-RFLP) analysis was conducted on the 16S rRNA genes of the bacterial communities colonizing the epithelial surfaces of the terminal ilea of open conventionally housed mice in an institutional small-animal facility. Polymeric-immunoglobulin-receptor-deficient (pIgR−/−) mice that were unable to secrete antibodies across mucosal surfaces were cohoused with normal and otherwise genetically identical wild-type (C57BL/6) mice for 4 weeks. If secretory antibodies played a role in modeling the gastrointestinal microbiota, C57BL/6 mice would have had a more distinct and uniform microbiota than their pIgR−/−cage mates. The T-RFLP profiles of the bacterial communities were compared by using Sorensen's pairwise similarity coefficient, a newly developed weighted pairwise similarity coefficient, and on the basis of Shannon's and Simpson's diversity indices. No systematic differences were observed between the dominant components of the mucosa-associated bacterial communities of the terminal ileal walls of the two types of mice, indicating that secretory antibodies do not control the composition of this microbiota. Similar analyses of experiments conducted at two different times, between which the bacterial community composition of the mouse colony in the small-animal facility appeared to have changed, showed that differences could have been detected, had they existed.
Cloning, Expression, and Purification of Choline Dehydrogenase from the Moderate Halophile
Halomonas elongata
Authors: Gadda Giovanni , McAllister-Wilkins Elien Elizabeth
Published: April 2003

Choline dehydrogenase (EC 1.1.99.1) catalyzes the four-electron oxidation of choline to glycine-betaine via a betaine-aldehyde intermediate. Such a reaction is of considerable interest for biotechnological applications in that transgenic plants engineered with bacterial glycine-betaine-synthesizing enzymes have been shown to have enhanced tolerance towards various environmental stresses, such as hypersalinity, freezing, and high temperatures. To date, choline dehydrogenase has been poorly characterized in its biochemical and kinetic properties, mainly because its purification has been hampered by instability of the enzyme in vitro. In the present report, we cloned and expressed in
Escherichia coli
the
betA
gene from the moderate halophile
Halomonas elongata
which codes for a hypothetical choline dehydrogenase. The recombinant enzyme was purified to more than 70% homogeneity as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and by treatment with 30 to 50% saturation of ammonium sulfate followed by column chromatography using DEAE-Sepharose. The purified enzyme showed similar substrate specificities with either choline or betaine-aldehyde as the substrate, as indicated by the apparent
V
/
K
values (where
V
is the maximal velocity and
K
is the Michaelis constant) of 0.9 and 0.6 μmol of O
2
min
−1
mg
−1
mM
−1
at pH 7 and 25°C, respectively. With 1 mM phenazine methosulfate as the primary electron acceptor, the apparent
V
max
values for choline and betaine-aldehyde were 10.9 and 5.7 μmol of O
2
min
−1
mg
−1
, respectively. These
V
max
values decreased four- to sevenfold when molecular oxygen was used as the electron acceptor. Altogether, the kinetic data are consistent with the conclusion that
H. elongata betA
codes for a choline dehydrogenase that can also act as an oxidase when electron acceptors other than molecular oxygen are not available.
Bacteriophages ofErwinia amylovora
Authors: Gill J. J. , Svircev A. M. , Smith R. , Castle A. J.
Published: April 2003

Fifty bacteriophage isolates ofErwinia amylovora, the causal agent of fire blight, were collected from sites in and around the Niagara region of southern Ontario and the Royal Botanical Gardens, Hamilton, Ontario. Forty-two phages survived the isolation, purification, and storage processes. The majority of the phages in the collection were isolated from the soil surrounding trees exhibiting fire blight symptoms. Only five phages were isolated from infected aerial tissue in pear and apple orchards. To avoid any single-host selection bias, six bacterial host strains were used in the initial isolation and enrichment processes. Molecular characterization of the phages with a combination of PCR and restriction endonuclease digestions showed that six distinct phage types, described as groups 1 to 6, were recovered. Ten phage isolates were related to the previously characterizedE. amylovoraPEa1, with some divergence of molecular markers between phages isolated from different sites. A study of the host ranges of the phages revealed that certain types were unable to efficiently lyse someE. amylovorastrains and that some isolates were able to lyse the epiphytic bacteriumPantoea agglomerans. Representatives from the six molecular groups were studied by electron microscopy to determine their morphology. The phages exhibited distinct morphologies when examined by an electron microscope. Group 1 and 2 phages were tailed and contractile, and phages belonging to groups 3 to 6 had short tails or openings with thin appendages. Based on morphotypes, the bacteriophages ofE. amylovorawere placed in the orderCaudovirales, in the familiesMyoviridaeandPodoviridae.
Karyotype Rearrangements in a Wine Yeast Strain by
rad52-
Dependent and
rad52
-Independent Mechanisms
Authors: Carro David , Bartra Enric , Piña Benjamin
Published: April 2003

Yeast strains isolated from the wild may undergo karyotype changes during vegetative growth, a characteristic that compromises their utility in genetic improvement projects for industrial purposes. Karyotype instability is a dominant trait, segregating among meiotic derivatives as if it depended upon only a few genetic elements. We show that disrupting the
RAD52
gene in a hypervariable strain partially stabilizes its karyotype. Specifically,
RAD52
disruption eliminated recombination at telomeric and subtelomeric sequences, had no influence on ribosomal DNA rearrangement rates, and reduced to 30% the rate of changes in chromosomal size. Thus, there are at least three mechanisms related to karyotype instability in wild yeast strains, two of them not requiring
RAD52
-mediated homologous recombination. When utilized for a standard sparkling-wine second fermentation,
Δrad52
strains retained the enological properties of the parental strain, specifically its vigorous fermentation capability. These data increase our understanding of the mechanisms of karyotype instability in yeast strains isolated from the wild and illustrate the feasibility and limitations of genetic remediation to increase the suitability of natural strains for industrial processes.
Survival of
Yersinia pestis
on Environmental Surfaces
Authors: Rose Laura J. , Donlan Rodney , Banerjee Shailen N. , Arduino Matthew J.
Published: April 2003

The survival of two strains of
Yersinia pestis
(avirulent A1122 and virulent Harbin) on the surfaces of four materials was investigated. Viability was evaluated with epifluorescence microscopy by using the metabolic stain cyanoditolyl tetrazolium chloride and plate counts. Small numbers of cells suspended in phosphate buffer survived 2 to 4 h after visible drying on stainless steel, polyethylene, or glass and beyond 48 h on paper. Cells suspended in brain heart infusion broth (BHI) persisted more than 72 h on stainless steel, polyethylene, and glass. Small numbers of cells suspended in BHI were still viable at 120 h on paper. These data suggest that
Y. pestis
maintains viability for extended periods (last measured at 5 days) under controlled conditions.
Conservation of Ornamental Stone byMyxococcus xanthus-Induced Carbonate Biomineralization
Authors: Rodriguez-Navarro Carlos , Rodriguez-Gallego Manuel , Ben Chekroun Koutar , Gonzalez-Muñoz Maria Teresa
Published: April 2003

Increasing environmental pollution in urban areas has been endangering the survival of carbonate stones in monuments and statuary for many decades. Numerous conservation treatments have been applied for the protection and consolidation of these works of art. Most of them, however, either release dangerous gases during curing or show very little efficacy. Bacterially induced carbonate mineralization has been proposed as a novel and environmentally friendly strategy for the conservation of deteriorated ornamental stone. However, the method appeared to display insufficient consolidation and plugging of pores. Here we report thatMyxococcus xanthus-induced calcium carbonate precipitation efficiently protects and consolidates porous ornamental limestone. The newly formed carbonate cements calcite grains by depositing on the walls of the pores without plugging them. Sonication tests demonstrate that these new carbonate crystals are strongly attached to the substratum, mostly due to epitaxial growth on preexisting calcite grains. The new crystals are more stress resistant than the calcite grains of the original stone because they are organic-inorganic composites. Variations in the phosphate concentrations of the culture medium lead to changes in local pH and bacterial productivity. These affect the structure of the new cement and the type of precipitated CaCO3polymorph (vaterite or calcite). The manipulation of culture medium composition creates new ways of controlling bacterial biomineralization that in the future could be applied to the conservation of ornamental stone.
Killing of
Bacillus
Spores by Aqueous Dissolved Oxygen, Ascorbic Acid, and Copper Ions
Authors: Cross J. B. , Currier R. P. , Torraco D. J. , Vanderberg L. A. , Wagner G. L. , Gladen P. D.
Published: April 2003

An approach to decontamination of biological endospores is discussed. Specifically, the performance of an aqueous modified Fenton reagent is examined. A modified Fenton reagent formulation of cupric chloride, ascorbic acid, and sodium chloride is shown to be an effective sporicide under aerobic conditions. The traditional Fenton reaction involves the conversion of hydrogen peroxide to hydroxyl radical by aqueous ionic catalysts such as the transition metal ions. Our modified Fenton reaction involves the conversion of aqueous dissolved oxygen to hydrogen peroxide by an ionic catalyst (Cu
2+
) and then subsequent conversion to hydroxyl radicals. Results are given for the modified Fenton reagent deactivating spores of
Bacillus globigii
. A biocidal mechanism is proposed that is consistent with our experimental results and independently derived information found in the literature. This mechanism requires diffusion of relatively benign species into the interior of the spore, where dissolved O
2
is then converted through a series of reactions which ultimately produce hydroxyl radicals that perform the killing action.
Rope-Producing Strains of
Bacillus
spp. from Wheat Bread and Strategy for Their Control by Lactic Acid Bacteria
Authors: Pepe Olimpia , Blaiotta Giuseppe , Moschetti Giancarlo , Greco Teresa , Villani Francesco
Published: April 2003

Two types of white wheat bread (high- and low-type loaves) were investigated for rope spoilage. Thirty of the 56 breads tested developed rope spoilage within 5 days; the high-type loaves were affected by rope spoilage more than the low-type loaves. Sixty-one
Bacillus
strains were isolated from ropy breads and were characterized on the basis of their phenotypic and genotypic traits. All of the isolates were identified as
Bacillus subtilis
by biochemical tests, but molecular assays (randomly amplified polymorphic DNA PCR assay, denaturing gradient gel electrophoresis analysis, and sequencing of the V3 region of 16S ribosomal DNA) revealed greater
Bacillus
species variety in ropy breads. In fact, besides strains of
B. subtilis, Bacillus
licheniformis
,
Bacillus
cereus
, and isolates of
Bacillus
clausii
and
Bacillus
firmus
were also identified. All of the ropy
Bacillus
isolates exhibited amylase activity, whereas only 32.4% of these isolates were able to produce ropiness in bread slices after treatment at 96°C for 10 min. Strains of lactic acid bacteria previously isolated from sourdough were first selected for antirope activity on bread slices and then used as starters for bread-making experiments. Prevention of growth of approximately 10
4
rope-producing
B. subtilis
G1 spores per cm
2
on bread slices for more than 15 days was observed when heat-treated cultures of
Lactobacillus plantarum
E5 and
Leuconostoc mesenteroides
A27 were added. Growth of
B. subtilis
G1 occurred after 7 days in breads started with
Saccharomyces cerevisiae
T22,
L. plantarum
E5, and
L. mesenteroides
A27.
Reconstruction of Mycobacterial Dehalogenase Rv2579 by Cumulative Mutagenesis of Haloalkane Dehalogenase LinB
Authors: Nagata Yuji , Prokop Zbyněk , Marvanová Soňa , Sýkorová Jana , Monincová Marta , Tsuda Masataka , Damborský Jiří
Published: April 2003

The homology model of protein Rv2579 fromMycobacterium tuberculosisH37Rv was compared with the crystal structure of haloalkane dehalogenase LinB fromSphingomonas paucimobilisUT26, and this analysis revealed that 6 of 19 amino acid residues which form an active site and entrance tunnel are different in LinB and Rv2579. To characterize the effect of replacement of these six amino acid residues, mutations were introduced cumulatively into the six amino acid residues of LinB. The sixfold mutant, which was supposed to have the active site of Rv2579, exhibited haloalkane dehalogenase activity with the haloalkanes tested, confirming that Rv2579 is a member of the haloalkane dehalogenase protein family.
Release of Extracellular Transformable Plasmid DNA from
Escherichia coli
Cocultivated with Algae
Authors: Matsui Kazuaki , Ishii Nobuyoshi , Kawabata Zen'ichiro
Published: April 2003

We studied the effects of cocultivation with either
Euglena gracilis
(Euglenophyta),
Microcystis aeruginosa
(Cyanophyta),
Chlamydomonas neglecta
(Chlorophyta), or
Carteria inversa
(Chlorophyta) on the production of extracellular plasmid DNA by
Escherichia coli
LE392(pKZ105). Dot blot hybridization analysis showed a significant release of plasmid DNA by cocultivation with all the algae tested. Further analysis by electrotransformation confirmed the release of transformable plasmid DNA by cocultivation with either
E
.
gracilis
,
M
.
aeruginosa
, or
C
.
inversa
. These results suggest algal involvement in bacterial horizontal gene transfer by stimulating the release of transformable DNA into aquatic environments.
Cyanobacteria Associated with Coral Black Band Disease in Caribbean and Indo-Pacific Reefs
Authors: Frias-Lopez Jorge , Bonheyo George T. , Jin Qusheng , Fouke Bruce W.
Published: April 2003

For 30 years it has been assumed that a single species of cyanobacteria,
Phormidium corallyticum
, is the volumetrically dominant component of all cases of black band disease (BBD) in coral. Cyanobacterium-specific 16S rRNA gene primers and terminal restriction fragment length polymorphism analyses were used to determine the phylogenetic diversity of these BBD cyanobacteria on coral reefs in the Caribbean and Indo-Pacific Seas. These analyses indicate that the cyanobacteria that inhabit BBD bacterial mats collected from the Caribbean and Indo-Pacific Seas belong to at least three different taxa, despite the fact that the corals in each case exhibit similar signs and patterns of BBD mat development.
Effect of natural antioxidants and apocynin on LPS-induced endotoxemia in rabbit
Authors: Ben-Shaul V , Lomnitskil L , Nyska A , Carbonatto M , Peano S , Zurovskyl Y , Bergman M , Eldridge S R , Grossman S
Published: April 2003

The objective of this study was to compare the prophylactic effects of the natural antioxidant from spinach (NAO) and apocynin, on the hepatic oxidative stress and liver damage induced by lipopolysaccharide (LPS). Male New Zealand rabbits were challenged with LPS with or without 8 days of antioxidant pretreatment. Pretreatment with NAO, but not apocynin, significantly (p<0.05) decreased the levels of hydroperoxides and malondialdehyde (MDA) in the liver cytosolic fraction and the activity of NADPH oxidase-generated superoxide in the microsomal fraction, compared to LPS alone. The activity of glutathione peroxidase (G-POX) was significantly (p<0.05) increased in the LPS-treated group, whereas treatment with NAO, but not apocynin, significantly (p< 0.05) decreased G-POX activity. Pretreatment with the same antioxidants had no significant effects on superoxide dismutase (SOD) activity, whereas an increased level of catalase (CAT) was obtained in all LPS-treated groups. TUNEL immunohistochemical staining in the LPS-treated animals indicated that there was no increase in apoptosis outside of necrotic foci. However, apoptotic hepatocytes were observed within areas of focal necrosis in animals exposed to LPS alone or LPS plus apocynin. Hepatocyte cell proliferation was tested by the proliferating-cell nuclear antigen (PCNA) tool, which indicated a proliferative effect in the LPS group, whereas the effect disappeared in the antioxidant-treated groups. The prophylactic effect of NAO on liver pathology and the significant decreases in lipid peroxidation products and NADPH oxidase activity suggest the use of NAO as an efficient strategy for treatanent of endotoxemia.
Radiotoxicity of h-R3 monoclonal antibody labeled with 188Re administered intracerebrally in rats
Authors: Gonzalez B , Casaco A , Alvarez P , Leon M , Arteaga M , Leon A , Santana E , Bada A , Figueredo R , Hernandez R , Iznaga-Escobar N , Gonzailez F , Perez R
Published: April 2003

Brain tumors are often incurable despite current aggressive treatmentmodalities. Regional intracerebral administration of labeled monoclonal antibodies (Mabs) can maximize the radioisotope and Mab concentration to tumor sites while reducing systemic toxicity. h 3 is a humanized antiepidemal growth factor receptor Mab that successfully targets the epidermal growth factor receptor, which is overexpressed in glioblastomas. We studied the acute local and systemic toxicity effects of intraventricular 188Re 3 in rats. Forty rats were distributed into four groups with five animals of each sex in each group. A single 5-II dose (2.5 pl into the left and 2.5 MI into the right lateral ventricles) of neutral solution containing50pgofh-R31abeledwith49.5 t 1.7,284±13.7or 579±23.7 p Ci of 188Re were stereotactically administered to each animal. Control animals received vehicle alone. Each animal was observed twice daily for detection of toxicity signs. Bodyweights were recorded on days 0, 7 and 14. Blood samples for analysis of hematological and clinical chemistry parameters were taken on days 0 and 14. Necropsy and histopathological studies were carried out after completion of the study. All animals, but one, remained clinically stable. Toxicities included local radionecrosis, discrete increase in ALAT and creatinine blood values at higher dose level. We concluded that a single intraventricular administration of relatively large doses of 188Re 3 is tolerable and causes minimal local and systemic toxicity effects in rats. Nevetheless, further studies are necessary to discard learning and behavioral problems.
Antidotes — inappropriate timely availability
Authors: Higgins M A , Evans R
Published: April 2003

The aim of this study was to determine the availability of antidotes to poisons in Wales and the South West of England. A stocklist of antidotes that are available to accident and emergency departments was requested and was compared with recommendations from the International Programme on Chemical Safety (IPCS). Chief pharmacists were invited to complete a short questionnaire regarding knowledge of existing guidelines. Thirty-four of 43 centres replied (response rate 77%). No department held all 36 antidotes (mean 13, range 7-33). All departments held antidotes that were frequently used. Ninety-one percent of departments held one cyanide antidote. Eighty-eight percent held one heavy metal chelating agent. The remaining antidotes were variably stocked. New agents such as 4-methylpyrazole, hydroxocobalamin and the heavy metal chelating agents DMSA and DMPS were infrequently held. Twenty of 34 chief pharmacists were unfamiliar with existing UK guidelines. A trend exists whereby larger departments stocked more antidotes. Some antidotes to poisons are not available in a timely fashion in Wales and the South West of England. There is a lack of awareness of existing guidelines. New recommendations relevant to clinical need and local practice should ideally be developed.
Radiation hormesis: the demise of a legitimate hypothesis
Authors: Calabrese E J , Baldwin L A
Published: April 2003

This paper examines the underlying factors that contributed to the marginalization of radiation hormesis in the early and middle decades of the 20th century. The most critical factor affecting the demise of radiation hormesis was a lack of agreement over how to define the concept of hormesis and quantitatively describe its dose-response features. If radiation hormesis had been defined as a modest overcompensation to a disruption in homeostasis as would have been consistent with the prevailing notion in the area of chemical hormesis, this would have provided the theoretical and practical means to blunt subsequent legitimate criticism of this hypothesis. A second critical factor undermining the radiation hormesis hypothesis was the generally total lack of recognition by radiation scientists of the concept of chemical hormesis which was markedly more advanced, substantiated and generalized than in the radiation domain. The third factor was that major scientific criticism of low dose stimulatory responses was galvanized at the time that the National Research Council (NRC) was organizing a national research agenda on radiation and the hormetic hypothesis was generally excluded from the future planned research opportunities. Furthermore, the criticisms of the leading scientists of the 1930s which undermined the concept of radiation hormesis were limited in scope and highly flawed and then perpetuated over the decades by other ‘prestigious’ experts who appeared to simply accept the earlier reports. This setting was then linked to a growing fear of radiation as a cause of birth defects, mutation and cancer, factors all reinforced by later concerns over the atomic bomb. Strongly supportive findings on hormetic effects in the 1940s by Soviet scientists were either generally not available to US scientists or disregarded as part of the Cold War mindset without adequate analysis. Finally, a massive, but poorly designed, US Department of Agriculture experiment in the late 1940s to assess the capacity for low dose plant stimulation by radionuclides failed to support the hormetic hypothesis thereby markedly lessening enthusiasm for research and funding in this area. Thus, the combination of a failed understanding of the hormetic hypothesis and its linkage with a strong chemical hormesis database, flawed analyses by prestigious scientists at the critical stage of scientific research development, reinforced by a Cold War mentality led to marginalization of an hypothesis (i.e., radiation hormesis) that had substantial scientific foundations and generalizability.
Tales of two similar hypotheses: the rise and fall of chemical and radiation hormesis
Authors: Calabrese E J , Baldwin L A
Published: April 2003

This paper compares the historical developments of chemical and radiation hormesis from their respective inceptions in the late 1880‘s for chemical hormesis and early 1900‘s for radiation hormesis to the mid 1930‘s to 1940 during which both hypotheses rose to some prominence but then became marginalized within the scientific community. This analysis documents that there were marked differences in their respective temporal developments, and the direction and maturity of research. In general, the formulation of the chemical hormesis hypothesis displayed an earlier, more-extensive and more sophisticated development than the radiation hormesis hypothesis. It was able to attract prestigious researchers with international reputations from leading institutions, to be the subject of numerous dissertations, to have its findings published in leading journals, and to have its concepts incorporated into leading microbiological texts. While both areas became the object of criticism from leading scientists, the intensity of the challenge was greatest for chemical hormesis due to its more visible association with the medical practice of homeopathy. Despite the presence of legitimate and flawed criticism, the most significant limitations ofboth chemical and radiation hormesis and their respective ultimate undoing were due to their: (1) lack of development of a coherent doseresponse theory using data of low dose stimulation from both the chemical and radiation domains; (2) difficulty in replication of low dose stimulatory responses without an adequate study design especially with respect to an appropriate number and properly spaced doses below the toxic threshold; (3) modest degree of stimulation even under optimal conditions which was difficult to distinguish from normal variation; and (4) lack of appreciation of the practical and/or commercial applications of the concepts of low dose stimulation.
Sodium monochloroacetate causes cytotoxic effects, an increased lactate and pyruvate level and induces ultra structural and cytoskeletal alterations in cultured kidney and liver epithelial cells
Authors: Dartsch P C , Wolburg H , Makdessi S Al , Schiek D , Sweidan H , Kimmel R , Schmahl F W
Published: April 2003

1 Monochloroacetic acid (MCAA) and its sodium salt, sodium monochloroacetate (SMCA) are widely used in chemical industries as intermediates in the synthesis of carboxymethylcellulose, phenoxyacetic acid, thio-glycolic acid, glycine, indigoid dyes and others. More-over, MCAA has been found as a by-product of the chlorination disinfection of drinking water and as an environmental contaminant of the atmosphere from the photodechlorination reactions of chlorinated hydrocarbons. Little is known about the mode of action of both compounds on the cellular level. From cases of accidental poisoning of man it is known that MCAA accumulates in liver and kidney. 2 In this study, the cytotoxicity of SMCA on cultured liver (Chang liver cells) and kidney epithelial cells of the proximal tubule (Opossum kidney cells) was investigated and its effect on metabolism, ultrastruc-ture and organization of cytoskeleton was examined. 3 Independent from the growth state of the cells (proliferating or quiescent), the results clearly show that SMCA causes a dose-dependent decrease in cell viability after an exposure period of 24 h. In all experiments, proliferating cells were more sensitive than quiescent and confluent cells. Liver cells were less sensitive against SMCA treatment than kidney epithe-lial cells. In contrast to liver cells, kidney cells exhibited a dose-dependent decrease in cell volume. The decrease in cell viability was accompanied by an increase of lactate and pyruvate concentrations released into the culture medium. In the case of Opossum kidney cells, lactate and pyruvate levels increased 5-6-fold, whereas in the case of Chang liver cells the increase was approximately twofold. While the ultrastructure of liver cells remained unaltered after drug treatment, kidney cells exhibited cytoplas-mic vacuolization, membraneous disruption and especially mitochondrial alterations. In accordance with the changes in the ultrastructure of Opossum cells, was the reorganization of cytoskeletal elements with an increased stress fiber network at the basolat-eral surface as well as a partial depolymerization of microtubules and vimentin filaments. A cytoskeletal reorganization was not observed for Chang liver cells after SMCA treatment. 4 The results demonstrate that SMCA causes a dose-dependent cytotoxicity which is accompanied by metabolic, mitochondrial and cytoskeletal alterations in the cells.
Antidotal effects of 2,3–dimercaptoprop ane– 1–sulfonic acid (DMPS) and meso–2,3– dimercaptosuccinic acid (DMSA) on the organotoxicity of dibutyltin dichioride (DBTC) in rats
Authors: Merkord J , Weber H , Krdning G , Hennighausen G
Published: April 2003

1 Dialkyltin compounds have been widely used in industry and agriculture, mainly as biocides, catalysts and plast stabilizer. In dependence on the length of the alkyl chains these organotins exert toxic effects on the immune system, the bile duct, liver and pancreas. It has been supposed that similar to organoarsenic the toxicity of the dialkyltin compounds is related to reactions with biological dithiol groups. Therefore, in the present study, the antidotal effects of 2,3-dimer-capto-propane-l-sulfonic acid (DMPS) and meso-2,3-dimercaptosuccinic acid (DMSA) on the organotoxic effects of dibutyltin dichloride (DBTC, single adminis-tration of 27,umol kg-' b.w. i.v.) in rats were studied using different doses (100 and 500 limol kg-1 b.w.) and routes of administration (i.p. and p.o.) of both chelators. Several parameters of organotoxicity (thy-mus weight and cellularity, bile duct diameter, histological lesions of pancreas and liver, activities of amylase, lipase and alkaline phosphatase, bilirubin and hyaluronic acid in serum) were measured from 6 h to 8 weeks. 2 DMPS and DMSA diminished the DBTC induced bile duct, pancreas and liver lesions stronger than the thymus atrophy. Moreover, the development of a fibrosis of the pancreas and a cirrhosis of liver several weeks after single administration of DBTC to rats was inhibited by DMPS and DMSA. The antidotal effects on serum parameter were observed after both adminis-tration routes of the chelators. DMPS was more effective than DMSA in most measured parameters. The decrease in the biliary excretion of organotin by DMPS and DMSA seems to be the reason for the pronounced protective effects of DMPS and DMSA on bile duct, pancreas and liver. 3 For the treatment of poisonings with dibutyltin compounds, the administration of DMPS or DMSA can be recommended.
Assessment of immunosuppressant drug carcinogenicity: standard and alternative animal models
Authors: Hastings K L
Published: April 2003

Drugs intended for use in preventing allograft rejection in transplant patients are likely to be administered chroni-cally; thus, it is normally expected that sponsors would conduct nonclinical studies to determine the carcinogenic potential of candidate compounds. For pharmaceuticals other than biologic agents, this would mean that rodent carcinogenicity bioassays would be performed under most circumstances. Immunosuppressant drugs have presented unique challenges with respect to the issue of carcino-genicity bioassays. The pharmacological activity of ther-apeutic immunosuppressants is thought to make them highly likely to act as promoters/cocarcinogens, even in the absence of genotoxic activity. Thus, it is assumed that this class of drug would represent a carcinogenic hazard in the absence of confirmatory standard rodent bioassay data. In addition, rodents typically have been sensitive to the pharmacological/toxicological effects of immunosup-pressants. It has proven to be difficult, therefore, to conduct life-time bioassays at doses reasonably equiva-lent to those that would be used clinically. For this and other reasons, alternative models might be more appro-priate for risk assessment with this class of drugs.
Disruption of the reproductive system and reproductive performance by administration of nonyiphenol to newborn rats
Authors: Nagao T , Saito Y , Usumi K , Nakagomi M , Yoshimura S , Ono H
Published: April 2003

A number of alkylphenolic compounds are used in a variety of commercial products and have been shown in in vitro studies to be weakly estrogenic, but few in vivo data are available addressing this issue in mammals. Human ex-posure to alkylphenols may occur not only from these environmental contaminants but also through contact with manufactured and metabolic breakdown products. The reproductive function of rats treated subcutaneously with nonylphenol (NP, 500 mg/kg/day) or 17Q3-estradiol (E2, 2 mg/kg/day) as a positive control, from postnatal days 1 to 5 was examined after puberty. In addition, masculine sexual behavior, sperm motion, plasma testosterone concentration and histopathological changes in the reproductive organs of the rats were examined. Furthermore, male rats were subjected to an open field test and wheel cage test to evaluate locomotor activity, and the estrous cycle was examined in female rats. All male and female rats exposed neonatally to NP or E2 showed macroscopic and/or microscopic altera-tions of the gonads. Females treated with NP or E2 showed an altered estrous cycle and abnormal reproductive function, while males treated with NP or E2 showed normal reproduc-tion. In males exposed neonatally to NP or E2, no abnorm-alities were observed in locomotor activity, sperm motion or plasma testosterone concentration. The results of this study indicate that early neonatal exposure to NP causes dysfunc-tion of postpubertal reproductive function in female rats, as well as disrupted development of gonads in male and female rats. More detailed studies are warranted to assess the possible risks to human and wildlife reproduction from exposure to NP and other environmental chemicals with estrogenic activity.
Biochemical and morphological changes in lung tissue and isolated lung cells of rats induced by short-term nitrogen dioxide exposure
Authors: van Bree L , Rietjens I MCM , Alink G M , Dormans J AMA , Marra M , Rombout P JA
Published: April 2003

To investigate the effects of repeated exposure to nitrogen dioxide (NO2) on antioxidant enzymes in lung tissue and isolated lung cells, rats were continuously exposed to 20 mg/m3 NO2 (10.6 ppm) for 4 days. The activities of glucose-6-phosphate dehydrogenase (G6PDH), glutathione reductase (GR), and glutathione peroxidase (GSHPx) were measured in the cytosolic fraction of lung tissue of both control and NO2-exposed rats as well as in isolated alveolar macrophages (AMs) and type II cells. Qualitative and quantitative changes in AM and type II cells were studied by electron microscopy and by morphometric analyses using enzyme and immunohistochemistry. NO2 exposure resulted in significantly increased pulmonary activities of G6PDH, GR, and GSHPx, both expressed per lung and per gram of lung weight. Morphometric data show that NO2 exposure significantly increased the number of type II cells, predominantly in the centriacinar region, indicating proliferation of epithelium following cellular injury. Type II cells in lungs of NO2-exposed rats had a squamous, less cuboidal appearance with more lamellar bodies compared to type II cells in lungs of control rats. Compared to control lungs, a higher number of macrophages could be isolated from NO2-exposed lungs, while numbers of type II cells isolated from lungs of control and NO2-exposed rats were the same. Isolated type II cells from control and NO2-exposed rats were polymorphic, with a small number of lamellar bodies and without polarity. Isolated macro-phages were rounded and contained many filopodia. NO2 exposure caused increases in the activities of G6PDH and GSHPx in isolated type II cells and of GSHPx in isolated macrophages, when expressed per number of cells. Macrophages and type II cells isolated from control and NO2-exposed rats and re-exposed in vitro to NO2, showed no differences in phagocytosis and viability features. Our results indicate that NO2-induced increases in pulmonary antioxidant enzymes are also reflected in isolated AM and type II cells. Since these lung cells do not display a decreased sensitivities toward an in vitro NO2 exposure, overall increase in antioxidant enzyme activities do not seem to play the most pivotal role in controlling cellular NO2 sensitivity and oxidant defence. Combined data from biochemical, morphological, and morphometric analyses of lungs and lung cells suggest that lung cell and tissue oxidant sensitivity and defence largely depends on the cell and tissue organisation, i.e., cell numbers and morphology as well as the ratio of surface area to cytoplasmic volume.
Prospects for reducing and refining the use of dogs in the regulatory toxicity testing of pharmaceuticals
Authors: Broadhead C L , Betton G , Combes R , Damment S , Everett D , Garner C , Godsafe Z , Healing G , Heywood R , Jennings M , Lumley C , Oliver G , Smith D , Straughan D , Topham J , Wallis R , Wilson S , Buckley P
Published: April 2003

A workshop was held to critically discuss the need for a non-rodent species and the role of the dog in regulatory toxicity testing of pharmaceuticals; to discuss opportunities to reduce and refine the use of dogs in preclinical toxicology; and to identify a number of specific recommendations which could be feasibly achieved to move the process forward. To facilitate a preliminary evaluation of the contribution of dog studies to the risk assessment process, anonymised, unpublished data were provided from fully evaluated, repeat dose toxicity studies in the rat and dog. Results ofthe International Life Sciences Institute (ILSI) Human Toxicity Project were also presented and discussed. Analysis of the data demonstrated that the dog can provide additional toxicity information, which, in some cases, was shown to be predictive for humans. Discussions indicated that there is potential for achieving a reduction in dog use and several possible approaches were identified. To further the progress of this initiative, there is a need to collate the results of pharmacology, toxicology, and clinical studies to address some of the proposed approaches. One of the outcomes of the workshop will be the establishment of a steering group to co-ordinate data collation for further analysis.
A true threshold dose in chemical carcinogenesis cannot be defined for a population, irrespective of the mode of action
Authors: Lutz W K
Published: April 2003

Strongly sigmoidal (S-shaped) dose-cancer incidence relationships are often observed in animal bioassays for carcinogenicity. If a genotoxic contribution is not plausible, an epigenetic mode of carcinogen action is proposed and a thresholded low dose-response suggested. In a strictsense, a threshold implies a no-yes situation, i.e., no effect up to the threshold dose and an effect above the threshold dose. A convincing explanation ofthe discontinuity ofthe gradient of the dose-response curve at the threshold dose is not available to me. However, the existence of a threshold is accepted for an individual. The threshold dose is the dose required for the manifestation of the tumor in an individual exactly at the end of a defined period of observation (for instance, 2 years in an animal bioassay, 75 years in humans). Because of genetic and lifestyle-dependent susceptibility differences, each animal or human has his individual threshold dose. For a group, no single threshold dose can be defined, irrespective of the mode of carcinogen action. Furthermore, in view of the stochastic elements in the process of carcinogenesis, the exact threshold dose can only be defined after tumor incidence and cannot be predicted.
The composition of cigarette smoke: a retrospective, with emphasis on polycyclic components
Authors: Rodgman A , Smith C J , Perfetti T A
Published: April 2003

The difficulties encountered in extrapolating biological activity from cigarette smoke composition provide generally applicable lessons as they are representative of the problems encountered with other complex mixtures. Researchers attempting to assess risk are faced with attempting to interpret data from a number of areas including: tobacco science; smoke/aerosol chemistry specific to tobacco; sophisticated analytical chemistry applications and techniques for trapping, collecting, separating, and quantifying very specific compounds at nanogram to picogram levels; numerous biological testing methodologies; and animal models of tumors and carcinogenesis. Numerous hypotheses have been developed over the past five decades and tested with the technology of the day in attempts to interpret the biological activity of cigarette smoke in relation to the chemistry of this complex mixture. These hypotheses fall into several categories discussed in this review: mechanisms of pyrogenesis of polycyclic aromatic hydrocarbons (PAHs) in tobacco smoke; levels of PAHs in cigarette mainstream smoke (MS) and its tumorigenicity in mouse skin-painting experiments; control of PAH levels in MS; chemical indicators of cigarette smoke condensate (CSC) tumorigenicity; control of levels of MS components partitioned between the vapor phase and particulate phase of MS; tumorigenic threshold limits of CSC and many of its components; tumorigenic aza-arenes in tobacco smoke; MS components reported to be ciliastatic to smokers' respiratory tract cilia; anticarcinogenic tobacco-smoke components. Of 52 hypotheses reviewed in this paper, 15 have excellent data supporting the hypothesis based on today's technology. The remaining 37 hypotheses, although originally plausible, have since become insupportable in light of new and contradictory data generated over the years. Such data were generated sometimes by the original authors of the hypotheses and sometimes by other investigators. The hypotheses presented today are less likely to be supplanted because they are well conceived and have a strong mechanistic basis. The challenge for the future is the generation and interpretation of data relating the chemistry and biological activity associated with the dynamic and complex mixture of tobacco smoke.
Aspirin impairs antioxidant system and causes peroxidation in human erythrocytes and guinea pig myocardial tissue
Authors: Durak I , Karaayvaz M , Çimen M Y B , Avci A , Çimen O B , Büyukkogak S , Oztfirk H S , Ozbek Hanefi , Kagmaz M
Published: April 2003

This study aims to investigate possible effects of aspirin treatment on cellular oxidant/antioxidant system. In the first part of the study, 15 guinea pigs were given aspirin at three different doses (2200, 440 and 10 mg/kg/day) for 30 days and five were fed on the same diet without aspirin. After a month, animals were killed and their hearts were removed for use in analyses. In the other part, after fasting blood samples were obtained from 11 volunteer subjects, they were given aspirin (approximately 10 mg/kg/day) for 30 days and second blood samples were obtained after 1 month. Five volunteer subjects also participated as placebo control. Oxidant/antioxidant parameters, namely superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), catalase (CAT), malondialdehyde (MDA), nonenzymatic superoxide scavenger activity (NSSA), susceptibility to oxidation (SO) and antioxidant potential (AOP) values, were assayed in the samples. Antioxidant system was found to be impaired in the heart tissue from guinea pigs and in the erythrocytes from volunteer subjects. AOP and NSSA values were lower and MDA higher after aspirin treatment in both heart tissues and erythrocytes. In guinea pig heart tissue, SO was lower, but GSH-Px and CAT were unchanged after aspirin treatment. In human erythrocytes, SO was unchanged, but GSH-Px and CAT activities were increased after aspirin treatment. Changes in guinea pig heart tissues from animals treated with higher aspirin doses were more drastic relative to those of human erythrocytes, but no meaningful differences were observed between analysis parameters of control and lower-dose (10 mg/kg/day) aspirin-treated animals. Our results suggest that high-dose aspirin exerts significant toxicity to guinea pig myocardium and normal dose aspirin may cause peroxidation in the human erythrocytes due to its oxidant potential. We suppose that antioxidant supplementation may be beneficial for the people using aspirin for longer periods in order to prevent peroxidation damages.
Silica-induced cytokine release from A549 cells: importance of surface area versus size
Authors: Hefland R B , Schwarzel P E , Johansen B V , Myran T , Uthus N , Refsnes M
Published: April 2003

Physical and chemical properties such as structure, composition and surface reactivity determine the biological activity of mineral particles. Long-term exposureto crystalline silica is known to cause persistent pulmonary inflammation leading to adverse health effects. There is less information about the potential health effects of amorphous (noncrystal-line) silica. In this study, the inflammatory and cytotoxic potency of crystalline and amorphous silica in relation to particle size and surface area was assessed. Human epithelial lung cells (A549) were exposed to different size fractions of quartz (aerodynamic diameter 0.5,2 and 10 sm) and amorphous silica (diameter 0.3 pm). All particles induced increased release ofthe proinflammatory cytokines interleukin (IL)-6 and IL-8. When cells were exposed to equal masses of quartz, the smallest size fraction was the most potent. These differences, however, disappeared when cytokine release was related to equal surface areas. When amorphous silica and quartz were compared, the amorphoussilicawas mostpotentto induce IL-6 regardless of how exposure was expressed, whereas the smallest size fraction of quartz was the most potent inducer of IL-8. Thus, the surface area seems to be the critical determinant when potency of different sizes of quartz is compared.