bluuebunny/crossref_metadata_2025 huggingface dataset:
Biodegradation of the nitramine explosives hexahydro-1,3,5-trinitro-1,3,5-triazine and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine in cold marine sediment under anaerobic and oligotrophic conditions
Authors: Zhao Jian-Shen , Greer Charles W , Thiboutot Sonia , Ampleman Guy , Hawari Jalal
Published: April 2004

The in situ degradation of the two nitramine explosives, hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX), was evaluated using a mixture of RDX and HMX, incubated anaerobically at 10 °C with marine sediment from a previous military dumping site of unexploded ordnance (UXO) in Halifax Harbor, Nova Scotia, Canada. The RDX concentration (14.7 mg·L–1) in the aqueous phase was reduced by half in 4 days, while reduction of HMX concentration (1.2 mg·L–1) by half required 50 days. Supplementation with the carbon sources glucose, acetate, or citrate did not affect the removal rate of RDX but improved removal of HMX. Optimal mineralization of RDX and HMX was obtained in the presence of glucose. Using universally labeled (UL)-[14C]RDX, we obtained a carbon mass balance distributed as follows: CO2, 48%–58%; water soluble products, 27%–31%; acetonitrile extractable products, 2.0%–3.4%; and products covalently bound to the sediments and biomass, 8.9% (in the presence of glucose). The disappearance of RDX was accompanied by the formation of the mononitroso derivative hexahydro-1-nitroso-3,5-dinitro-1,3,5-triazine (MNX) and formaldehyde (HCHO) that subsequently disappeared. In the case of HMX, mineralization reached only 13%–27% after 115 days of incubation in the presence or absence of the carbon sources. The disappearance of HMX was also accompanied by the formation of the mononitroso derivative. The total population of psychrotrophic anaerobes that grew at 10 °C was 2.6 × 103 colony-forming units·(g sediment dry mass)–1, and some psychrotrophic sediment isolates were capable of degrading RDX under conditions similar to those used for sediments. Based on the distribution of products, we suggest that the sediment microorganisms degrade RDX and HMX via an initial reduction to the corresponding mononitroso derivative, followed by denitration and ring cleavage.Key words: biodegradation, nitramine explosives, marine sediment, psychrotrophic bacteria.
Over-expression ofrexAnullifies T4rIIexclusion inEscherichia coliK(λ) lysogens
Authors: Slavcev Roderick A , Hayes Sidney
Published: April 2004

Dosage and relative cellular levels of RexA and RexB proteins encoded by the rexA–rexB genes of a λ prophage are important for the Rex+phenotype, which was nullified when greater RexA or RexB was provided than was necessary for the complementation of a rexA–or a rexB–prophage.Key words: bacteriophage lambda (λ), T4rII exclusion (Rex) phenotype, lambda pM–cI–rexA–rexB–timmoperon.
Screening of a large panel of gastrointestinal peptide plasma levels is not adapted for the evaluation of digestive damage following irradiation
Authors: Dublineau I , Dudoignon N , Monti P , Combes O , Wysocki J , Grison S , Baudelin C , Griffiths N M , Scanff P
Published: April 2004

The aim of this study was to assess the potential of gastrointestinal peptide plasma levels as biomarkers of radiation-induced digestive tract damage. To this end, plasma levels of substance P, GRP, motilin, PYY, somatostatin-28, gastrin, and neurotensin were followed for up to 5 days in pigs after a 16-Gy whole-body X-irradiation, completed by a histopathological study performed at 5 days. Each peptide gave a specific response to irradiation. The plasma levels of GRP and substance P were not modified by irradiation exposure; neither were those of motilin and PYY. Concerning gastrin, a 23-fold increase of plasma concentration was observed in pig, which presented the most important histological alterations of the stomach. The plasma levels of somatostatin, unchanged from 1 to 4 days after irradiation, was also increased by 130% at 5 days. In contrast, a diminution of neurotensin plasma levels was noted, firstly at 1 day (88%), and from 3 days after exposure (50%). The present study suggested that changes in gastrin and neuro tensin plasma levels were associated with structural alterations of the stomach and ileum, respectively, indicating that they may be relevant biological indicators of radiation-induced digestive damage to these segments.Key words: gastrin, neurotensin, somatostatin, PYY, substance P, irradiation.
Alteration of the enterohepatic recirculation of bile acids in rats after exposure to ionizing radiation
Authors: Scanff P , Souidi M , Grison S , Griffiths N M , Gourmelon P
Published: April 2004

The aim of this work was to study acute alterations of the enterohepatic recirculation (EHR) of bile acids 3 days after an 8-Gy radiation exposure in vivo in the rat by a washout technique. Using this technique in association with HPLC analysis, the EHR of the major individual bile acids was determined in control and irradiated animals. Ex vivo ileal taurocholate absorption was also studied in Ussing chambers. Major hepatic enzyme activities involved in bile acid synthesis were also measured. Measurements of bile acid intestinal content and intestinal absorption efficiency calculation from washout showed reduced intestinal absorption with significant differences from one bile acid to another: absorption of taurocholate and tauromuricholate was decreased, whereas absorption of the more hydrophobic taurochenodeoxycholate was increased, suggesting that intestinal passive diffusion was enhanced, whereas ileal active transport might be reduced. Basal hepatic secretion was increased only for taurocholate, in accordance with the marked increase of CYP8B1 activity in the liver. The results are clearly demonstrate that concomitantly with radiation-induced intestinal bile acid malabsorption, hepatic bile acid synthesis and secretion are also changed. A current working model for pathophysiological changes in enterohepatic recycling after irradiation is thus proposed.Key words: irradiation, bile acids, intestine, liver, enterohepatic recycling.
Alterations in vascular endothelial function in the aorta and mesenteric artery in type II diabetic rats
Authors: Matsumoto Takayuki , Wakabayashi Kentaro , Kobayashi Tsuneo , Kamata Katsuo
Published: April 2004

We used the partial protection exerted by suitable dosages of nicotinamide against the β-cytotoxic effect of streptozotocin (STZ) to create an experimental diabetic syndrome in adult rats that appears closer to type II diabetes mellitus than other available animal models. The dosage of 230 mg/kg of nicotinamide given intraperitoneally 15 min before STZ administration (65 mg/kg i.v.) yielded animals with hyperglycemia (187.8 ± 17.8 vs. 103.8 ± 2.8 mg/dL in controls; P < 0.001) and preservation of plasma insulin levels. This study assessed the relationship between endothelial dysfunction and agonist-induced contractile responses in such rats. In the thoracic aorta, the acetylcholine (ACh) induced relaxation was significantly reduced and the noradrenaline (NA) induced contractile response was significantly increased in diabetic rats compared with age-matched control rats. In the superior mesenteric artery, the ACh-induced relaxation was similar in magnitude between diabetic and age-matched control rats; however, the ACh-induced endothelium-derived hyperpolarizing factor (EDHF) type relaxation was significantly weaker in diabetic rats than in the controls. The phenylephrine (PE) induced contractile response was not different between the two groups. The plasma concentration of NOx (NO2–+ NO3–) was significantly lower in diabetic rats than in control rats. We conclude that vasomotor activities in conduit arteries are impaired in this type II diabetes model.Key words: aorta, contraction, endothelium-derived hyperpolarizing factor, endothelium-mediated relaxation, mesenteric artery, type II diabetes.
Effet radiosensibilisateur de l'acide linoléique conjugué chez les cellules cancéreuses du sein MCF-7 et MDA-MB-231
Authors: Drouin Geneviève , Douillette Annie , Lacasse Pierre , Paquette Benoit
Published: April 2004

Apoptotic pathways in breast cancer cells are frequently altered, reducing the efficiency of radiotherapy. Conjugated linoleic acid (CLA), known to trigger apoptosis, was tested as radiosensitizer in breast cancer cells MCF-7 and MDA-MB-231. The CLA-mix, made up of the isomers CLA-9cis 11trans and CLA-10trans 12cis, was compared to three purified isomers, i.e., the CLA-9cis 11cis, CLA-9cis 11trans, and CLA-10trans 12cis. Using the apoptotic marker YO-PRO®-1, the CLA-9cis 11cis at 50 µmol/L turned out to be the best apoptotic inducer leading to a 10-fold increase in MCF-7 cells and a 2,5-fold increase in MDA-MB-231 cells, comparatively to the CLA-mix. Contrary to previous studies on colorectal and prostate cancer cells, CLA-10trans 12cis does not lead to an apoptotic response on breast cancer cell lines MCF-7 and MDA-MB-231. Our results also suggest that the main components of the CLA-mix (CLA-9cis 11trans and CLA-10trans 12cis) are not involved in the induction of apoptosis in the breast cancer cells studied. A dose of 5 Gy did not induce apoptosis in MCF-7 and MDA-MB-231 cells. The addition of CLA-9cis 11cis or CLA-mix has allowed us to observe a radiation-induced apoptosis, with the CLA-9cis 11cis being about 8-fold better than the CLA-mix. CLA-9cis 11cis turned out to be the best radiosensitizer, although the isomers CLA-9cis 11trans and CLA-10trans 12cis have also reduced the cell survival following irradiation, but using a mechanism not related to apoptosis. In conclusion, the radiosensitizing property of CLA-9cis 11cis supports its potential as an agent to improve radiotherapy against breast carcinoma.Key words: breast cancer, conjugated linoleic acid (CLA), radiotherapy, apoptosis.
In vitro evaluation of percutaneous diffusion of uranyl nitrate through intact or excoriated skin of rat and pig
Authors: Petitot F , Moreels A M , Paquet F
Published: April 2004

At the present time, the International Commission on Radiological Protection (ICRP) has not published any model concerning internal radioactive contamination by uptake from wounds. The aims of our work were to determine the time available to treat contamination of intact or wounded skin before a significant uptake of uranium occurred and to evaluate the consequences of incomplete decontamination on uranium uptake. The kinetics of percutaneous diffusion of uranium through intact or excoriated skin and its distribution in skin layers were evaluated using an in vitro technique. Our data demonstrated a dramatic increase of uranium percutaneous diffusion through excoriated skin compared with intact skin. Significant uptake of uranium through excoriated skin occurred in only 30 min, indicating that there is only a short interval available to treat a contaminated wound effectively. Moreover, in the case of an incompletely decontaminated superficial wound, viable epidermis behaved as a reservoir for uranium that remained bioavailable. At the present time, potential uptake of uranium and perhaps other radionuclides through intact or wounded skin is not adequately taken into account by radiological protection agencies. Our results emphasize the need for further study and modeling of uptake of radionuclides through intact or wounded skin.Key words: wound, skin, uranium, rat, pig.
Screening for supra-additive effects of cytotoxic drugs and gamma irradiation in an in vitro model for hepatocellular carcinoma
Authors: Lambert B , De Ridder L , Slegers G , de Gelder V , Dierckx R A , Thierens H
Published: April 2004

Hepatocellular carcinoma (HCC) is one of the most common malignancies in the world. A wide variety of treatment modalities is available for palliative therapy of HCC, although there is no strong evidence that these treatments can have a significant impact on survival. The aim of this work was to screen cytotoxic drugs relevant in the treatment of HCC for enhancement of the effect of irradiation in an in vitro model. As the majority of patients presenting with HCC suffer reduced liver function, attention was paid to low-dose effects of the cytotoxic drugs tested. To reflect this situation in vivo, multicellular tumor aggregates or "spheroids" of HepG2 cells were cultured and exposed to gamma irradiation alone or in combination with cisplatin for 4 h, gemcitabin for 4 or 24 h, or 5-fluorouracil for 4 h. In one experiment, the spheroids were cultured for 4 weeks in multiwell plates that allowed adhesion. Measurement of two-dimensional spheroid outgrowth was made every week for each spheroid. This kind of growth depends on the proliferation and motility of the cells that form the spheroid. In a second experiment, toxicity was evaluated by comparative growth curves by means of a three-dimensional growth assay and by histology. Supra-additive effects lasting for 4 weeks were observed for all drugs tested in combination with a gamma irradiation of 10 Gy.Key words: hepatocellular carcinoma, cisplatin, gemcitabine, 5-fluorouracil, gamma irradiation.
Radiation-induced peroxidation of small unilamellar vesicles of phosphatidylcholine generated by sonication
Authors: Vitrac H , Courrègelongue M , Couturier M , Collin F , Thérond P , Rémita S , Peretti P , Jore D , Gardès-Albert M
Published: April 2004

The present study was aimed at determining the peroxidation of model membranes constituted of liposomes of 1-palmitoyl-2-linoleoyl-sn-glycero-3-phosphocholine (PLPC) submitted to hydroxyl free radicals (generated by γ-radiolysis) attack. Liposomes of PLPC were prepared using the sonication technique, and dynamic light-scattering (DLS) measurements allowed characterization of the liposomal dispersions. Irradiation damages in sonication-generated liposomes were assessed by monitoring several oxidation products, such as conjugated dienes (by means of UV–visible spectrophotometry) and hydroperoxides (using reverse phase high-performance liquid chromatography (HPLC) associated with chemiluminescence detection). It has been shown that three different families of hydroperoxides are formed: the first one (at low radiation doses) results from HO· attack on the linoleyl chain of PLPC, giving phosphatidylcholine hydroperoxides possessing a conjugated dienic structure; the two others (at high radiation doses) are obtained by the secondary HO· attack on the primary hydroperoxide family. The quantification of these products associated with the comparison of their radiation-dose-dependent formation has provided valuable information concerning the mechanisms of their formation. Analysis by HPLC – mass spectrometry has confirmed the presence of hydroperoxides and underlined various other products, like chain-shortened fragments and oxygenated derivatives of polyunsaturated sn-2 fatty acyl chain residues. Structural assignment proposals of some oxidation products have been proposed.Key words: radiolysis, phospholipids, peroxidation, hydroperoxides, liposomes.
Effect of U and 137Cs chronic contamination on dopamine and serotonin metabolism in the central nervous system of the rat
Authors: Houpert P , Lestaevel P , Amourette C , Dhieux B , Bussy C , Paquet F
Published: April 2004

Following the Chernobyl accident, the most significant problem for the population of the former Soviet Union for the next 50-70 years will be chronic internal contamination by radionuclides. One of the few experiments carried out in this field reported that neurotransmitter metabolism in the central nervous system of the rat was disturbed after feeding with oats contaminated by 137Cs for 1 month. The present study assessed the effect of chronic contamination by depleted U or 137Cs on the metabolism of two neurotransmitters in cerebral areas of rats. Dopamine and serotonin were chosen because their metabolism has been shown to be disturbed after external irradiation, even at moderate doses. Dopamine, serotonin, and some of their catabolites were measured by high-pressure liquid chromatography coupled with an electrochemical detector in five cerebral structures of rats contaminated over a 1-month period by drinking water (40 mg U·L–1 or 6500 Bq 137Cs·L–1). In the striatum, hippocampus, cerebral cortex, thalamus, and cerebellum, the dopamine, serotonin, and catabolite levels were not significantly different between the control rats and rats contaminated by U or 137Cs. These results are not in accordance with those previously described.Key words: uranium, caesium, dopamine, serotonin, HPLC.
Morphological variation in the gastral sterna of female Apoidea (Insecta: Hymenoptera)
Authors: Packer Laurence
Published: April 2004

A large amount of morphological variation is described for gastral sterna 2 through 6 of female bees and wasps belonging to the Apoidea. This variation includes differences among taxa in the degree of development of the antecosta, acrosternum, gradulus, sternal apodemes, and posterolateral areas and the size and shape of the postgradular region. There is considerable variation among sterna 2 through 6 within individual bees. Some of this variation is mostly consistent among taxa, such as the increasing anterior divergence of the sternal apodemes and the decreasing ratio of posterior to anterior width of the more posterior sterna. Other characteristics vary inconsistently among sterna within or among taxa. Examples include the presence or absence of graduli or premarginal lines on different sterna. Variation is most extreme for the apical external sternum, the sixth, which exhibits a large number of apomorphic traits, particularly in the Andrenidae and cleptoparasitic Apidae. The potential utility of sternal characters for phylogenetic studies of the Apoidea is discussed. There would seem to be particularly useful variation in the form of sternum 6 among different lineages of cleptoparasitic Apidae, presumably associated with different means of gaining entry to and (or) ovipositing within cells of the host bee. Sternal characters might help resolve relationships among these morphologically otherwise often strongly convergent taxa.
Separation of foraging habitat among breeding sites of a colonial marine predator, the northern fur seal (Callorhinus ursinus)
Authors: Robson Bruce W , Goebel Michael E , Baker Jason D , Ream Rolf R , Loughlin Thomas R , Francis Robert C , Antonelis George A , Costa Daniel P
Published: April 2004

This study examines whether lactating northern fur seals (Callorhinus ursinus) from different breeding sites on the Pribilof Islands in the eastern Bering Sea forage in separate areas. Satellite transmitters were attached to 97 northern fur seal females from nine breeding areas for 119 complete foraging trips during the 1995 and 1996 breeding seasons. Females from St. Paul and St. George islands tended to travel in different directions relative to their breeding site in both years of the study. St. Paul Island females dispersed in all directions except to the southeast, where St. George Island females foraged. Habitat separation was also observed among breeding areas on northeastern and southwestern St. Paul Island and to a lesser degree on northern and southern St. George Island. Although foraging direction led to geographical separation among sites, the maximum distance traveled and the duration of foraging trips did not differ significantly among islands in either year. The results of this study document that lactating fur seals from the same site share a common foraging area and that females from different breeding sites tend to forage in separate areas and hydrographic domains.
Influence of egg aggregation and soil moisture on incubation of flexible-shelled lacertid lizard eggs
Authors: Marco Adolfo , Díaz-Paniagua Carmen , Hidalgo-Vila Judit
Published: April 2004

Many oviparous terrestrial species deposit flexible-shelled eggs into the soil. These eggs are sensitive to the hydration level of the nest environment. Among other factors, water exchange of eggs during incubation may be affected by the soil water potential. To evaluate whether egg aggregation influences embryonic development, we incubated flexible-shelled Schreiber's green lizard (Lacerta schreiberi) eggs under three levels of soil water potential (wet: –150 kPa; intermediate: –650 kPa; dry: –1150 kPa) and under two levels of aggregation (aggregated: in groups of six eggs with physical contact among them; isolated: groups of six eggs each 1 cm apart). The availability of water during egg incubation influenced egg mass and hatchling size. Eggs incubated in dry soils absorbed less water and produced smaller hatchlings. The selected levels of soil water potential did not influence incubation duration or hatching success. When soil was wet or dry, we did not find any effect of egg aggregation in embryonic development. However, when soil water potential was intermediate, aggregated eggs absorbed less water and their embryos hatched at smaller sizes compared with isolated eggs. Moreover, variability and range of egg water absorption and hatchling size were higher among aggregated eggs than among solitary ones when access to water was restricted. In these cases, eggs competed with different success for water, a limited resource in the nest environment.
Inversion polymorphism in a Connecticut RiverAxarusspecies (Diptera: Chironomidae): biometric effects of a triple inversion heterozygote
Authors: Werle Sean F , Klekowski Ed , Smith Douglas G
Published: April 2004

The authors sampled three spatially isolated populations of a chironomid midge in the genus Axarus living in the Connecticut River both early and late in the larval life cycle of one generation. Larvae were scored for both length and inversion frequency using the polytene chromosomes from salivary gland cells. We found polymorphism for four paracentric inversions. Inversion C1–6exhibits a geographic cline, increasing in frequency with increasing latitude but remaining stable over time. Also stable over time were two other paracentric inversions designated A1–5and F13–20, which were present at similar frequencies in all populations. None of these inversions was associated with larval length. A complex triple inversion designated G2–7was significantly correlated with decreased larval length and also exhibited a significant increase in frequency (within one cohort) in the two more northerly populations. We propose that this increase is due to size-selective predation eliminating larger larvae.
Lipoteichoic acid-stimulated p42/p44 MAPK activation via Toll-like receptor 2 in tracheal smooth muscle cells
Authors: Lee Chiang-Wen , Chien Chin-Sung , Yang Chuen-Mao
Published: April 2004

Lipoteichoic acid (LTA), the principal component of the cell wall of gram-positive bacteria, triggers several inflammatory responses. However, the mechanisms underlying its action on human tracheal smooth muscle cells (HTSMCs) were largely unknown. This study was to investigate the mechanisms underlying LTA-stimulated p42/p44 mitogen-activated protein kinase (MAPK) using Western blotting assay. LTA stimulated phosphorylation of p42/p44 MAPK via a Toll-like receptor 2 (TLR2). Pretreatment with pertussis toxin attenuated the LTA-induced responses. LTA-stimulated phosphorylation of p42/p44 MAPK was attenuated by inhibitors of tyrosine kinase (genistein), phosphatidylcholine-phospholipase C (PLC; D609), phosphatidylinositol (PI)-PLC (U-73122), PKC (staurosporine, Gö-6976, rottlerin, or Ro-318220), MEK1/2 (U-0126), PI 3-kinase (LY-294002 and wortmannin), and an intracellular Ca2+chelator (BAPTA-AM). LTA directly evoked initial transient peak of [Ca2+]i, supporting the involvement of Ca2+mobilization in LTA-induced responses. These results suggest that in HTSMCs, LTA-stimulated p42/p44 MAPK phosphorylation is mediated through a TLR2 receptor and involves tyrosine kinase, PLC, PKC, Ca2+, MEK, and PI 3-kinase.
Anti-monocyte chemoattractant protein-1 gene therapy attenuates pulmonary fibrosis in mice
Authors: Inoshima Ichiro , Kuwano Kazuyoshi , Hamada Naoki , Hagimoto Naoki , Yoshimi Michihiro , Maeyama Takashige , Takeshita Akira , Kitamoto Shiro , Egashira Kensuke , Hara Nobuyuki
Published: April 2004

Monocyte chemoattractant protein-1 (MCP-1) is a proinflammatory chemokine and may play an important role in the development of pulmonary fibrosis. We examined a new therapeutic strategy that comprises the transfection of the mutant MCP-1 gene into skeletal muscles as a biofactory for anti-MCP-1 therapy against bleomycin-induced pulmonary fibrosis in mice. Overexpression of the mutant MCP-1 gene at 10–14 days after intratracheal instillation of bleomycin resulted in decreased DNA damage, apoptosis, and pulmonary fibrosis at 14 days. However, overexpression of the mutant MCP-1 at 0–4 days after bleomycin instillation did not result in decreased pathological grade, DNA damage, or apoptosis at 7 and 14 days. Because, in this model, inflammatory cell infiltration begins at 3 days and is followed by interstitial fibrosis, it is likely that MCP-1 has an important role to play in the development of fibrogenesis but not in the development of early lung inflammation. This method does not require the use of viral vector or neutralizing antibody, and, as such, it is possible to avoid problems regarding the pathogenicity of the viral vector or immunocomplex. This new strategy may be a beneficial method of treating pulmonary fibrosis from the viewpoint of clinical application.
Pertussis toxin activatesl-arginine uptake in pulmonary endothelial cells through downregulation of PKC-α activity
Authors: Zharikov Sergey I. , Krotova Karina Y. , Belayev Leonid , Block Edward R.
Published: April 2004

Pertussis toxin (PTX) induces activation of l-arginine transport in pulmonary artery endothelial cells (PAEC). The effects of PTX on l-arginine transport appeared after 6 h of treatment and reached maximal values after treatment for 12 h. PTX-induced changes in l-arginine transport were not accompanied by changes in expression of cationic amino acid transporter (CAT)-1 protein, the main l-arginine transporter in PAEC. Unlike holotoxin, the β-oligomer-binding subunit of PTX did not affect l-arginine transport in PAEC, suggesting that Gαiribosylation is an important step in the activation of l-arginine transport by PTX. An activator of adenylate cyclase, forskolin, and an activator of protein kinase A (PKA), Sp-cAMPS, did not affect l-arginine transport in PAEC. In addition, inhibitors of PKA or adenylate cyclase did not change the activating effect of PTX on l-arginine uptake. Long-term treatment with PTX (18 h) induced a 40% decrease in protein kinase C (PKC)-α but did not affect the activities of PKC-ϵ and PKC-ζ in PAEC. An activator of PKC-α, phorbol 12-myristate 13-acetate, abrogated the activation of l-arginine transport in PAEC treated with PTX. Incubation of PTX-treated PAEC with phorbol 12-myristate 13-acetate in combination with an inhibitor of PKC-α (Go 6976) restored the activating effects of PTX on l-arginine uptake, suggesting PTX-induced activation of l-arginine transport is mediated through downregulation of PKC-α. Measurements of nitric oxide (NO) production by PAEC revealed that long-term treatment with PTX induced twofold increases in the amount of NO in PAEC. PTX also increased l-[3H]citrulline production from extracellular l-[3H]arginine without affecting endothelial NO synthase activity. These results demonstrate that PTX increased NO production through activation of l-arginine transport in PAEC.
Nitric oxide modulates caveolin-1 and matrix metalloproteinase-9 expression and distribution at the endothelial cell/tumor cell interface
Authors: Phillips Patricia G. , Birnby Linda M.
Published: April 2004

We used a two-compartment coculture model comprising human endothelial cells (EC) and non-small cell lung carcinoma (CA) cells to study capillary formation. Elevated NO concentrations, contributed in part by CA cells, lead to inhibited capillary formation (Phillips PG, Birnby LM, Narendran A, and Milonovich WL. Am J Physiol Lung Cell Mol Physiol 281: L278–L290, 2001). Here we demonstrate using gelatin substrate zymography that high NO concentrations, whether produced endogenously or by NO donor spermine-NONOate or peroxynitrite-generating compound SIN-1, significantly inhibit MMP-9 expression and activation. Furthermore, high NO concentrations decrease Cav-1 abundance and alter its cellular distribution in EC. Cav-1 is essential for capillary formation in this model because Cav-1 antisense treatments targeted to EC significantly inhibit capillary formation. Laser confocal microscopy demonstrated extensive colocalization of MMP-9 with Cav-1 in sprouting EC, primarily at the basolateral surfaces of EC in focal structures associated with directed migration. This codistribution was NO concentration dependent, and elevated NO concentrations lead to marked dissociation of these two proteins. We propose that compartmentalization of MMP-9 within caveolar structures does occur, and that this could facilitate directed proteolysis essential for early migratory and invasive processes. Our data suggest elevated NO concentrations could impact on capillary formation via a combination of direct effects on MMP activation and by altering the distribution or abundance of Cav-1. Consequences of Cav-1 alterations may include impaired activation of proteolytic enzymes that utilize caveolar structure for stabilization and/or compartmentalization of MMP-9 as well as other putative members of an ECM proteolytic cascade.
Thioredoxin liquefies and decreases the viscoelasticity of cystic fibrosis sputum
Authors: Rancourt Raymond C. , Tai Shusheng , King Malcolm , Heltshe Sonya L. , Penvari Churee , Accurso Frank J. , White Carl W.
Published: April 2004

The persistent and viscous nature of airway secretions in cystic fibrosis (CF) disease leads to airway obstruction, opportunistic infection, and deterioration of lung function. Thioredoxin (Trx) is a protein disulfide reductase that catalyzes numerous thiol-dependent cellular reductive processes. To determine whether Trx can alter the rheological properties of mucus, sputum obtained from CF patients was treated with TRX and its reducing system (0.1 μM thioredoxin reductase + 2 mM NADPH), and liquid phase-gel phase ratio (percent liquid phase) was assessed by compaction assay. Exposure to low Trx concentrations (1 μM) caused significant increases in the percentage of liquid phase of sputum. Maximal increases in percent liquid phase occurred with 30 μM Trx. Additional measurements revealed that sputum liquefaction by the Trx reducing system is dependent on NADPH concentration. The relative potency of the Trx reducing system also was compared with other disulfide-reducing agents. In contrast with Trx, glutathione and N-acetylcysteine were ineffective in liquefying sputum when used at concentrations <1 mM. Sputum viscoelasticity, measured by magnetic microrheometry, also was diminished significantly following 20-min treatment with 3, 10, or 30 μM Trx. Similarly, this reduction in viscoelasticty also was dependent on NADPH concentration. Further investigation has indicated that Trx treatment increases the solubility of high-molecular-weight glycoproteins and causes redistribution of extracellular DNA into the liquid phase of sputum. Recognizing that mucins are the major gel-forming glycoproteins in mucus, we suggest that Trx alters sputum rheology by enzymatic reduction of glycoprotein polymers present in sputum.
Albumin endocytosis in endothelial cells induces TGF-β receptor II signaling
Authors: Siddiqui Shahid S. , Siddiqui Zeba K. , Malik Asrar B.
Published: April 2004

Vascular endothelial cells undergo albumin endocytosis using a set of albumin binding proteins. This process is important for maintaining cellular homeostasis. We showed by several criteria that the previously described 73-kDa endothelial cell surface albumin binding protein is the 75-kDa transforming growth factor (TGF)-β receptor type II (TβRII). Albumin coimmunoprecipitated with TβRII from a membrane fraction from rat lung microvascular endothelial cells. Albumin endocytosis-negative COS-7 cells became albumin endocytosis competent when transfected with wild-type TβRII but not when transfected with a domain-negative kinase mutant of TβRII. An antibody specific for TβRII inhibited albumin endocytosis. A mink lung epithelial cell line, which expresses both the TGF-β receptor type I (TβRI) and the TβRII receptor, exhibited albumin binding to the cell surface and endocytosis. In contrast, mutant L-17 and DR-26 cells lacking TβRI or TβRII, respectively, each showed a dramatic reduction in binding and endocytosis. Albumin endocytosis induced Smad2 phosphorylation and Smad4 translocation as well as increased protein expression of the inhibitory Smad, Smad7. We identified regions of significant homology between amino acid sequences of albumin and TGF-β, suggesting a structural basis for the interaction of albumin with the TGF-β receptors and subsequent activation of TβRII signaling. The observed albumin-induced internalization of TβRII signaling may be an important mechanism in the vessel wall for controlling TGF-β responses in endothelial cells.
In vivo exposure to hyperoxia induces DNA damage in a population of alveolar type II epithelial cells
Authors: Roper Jason M. , Mazzatti Dawn J. , Watkins Richard H. , Maniscalco William M. , Keng Peter C. , O'Reilly Michael A.
Published: April 2004

It is well established that hyperoxia injures and kills alveolar endothelial and type I epithelial cells of the lung. Although type II epithelial cells remain morphologically intact, it remains unclear whether they are also damaged. DNA integrity was investigated in adult mice whose type II cells were identified by their endogenous expression of pro-surfactant protein C or transgenic expression of enhanced green fluorescent protein. In mice exposed to room air, punctate perinuclear 8-oxoguanine staining was detected in ∼4% of all alveolar cells and in 30% of type II cells. After 48 or 72 h of hyperoxia, 8-oxoguanine was detected in 11% of all alveolar cells and in >60% of type II cells. 8-Oxoguanine colocalized by confocal microscopy with the mitochondrial transmembrane protein cytochrome oxidase subunit 1. Type II cells isolated from hyperoxic lungs exhibited nuclear DNA strand breaks by comet assay even though they were viable and morphologically indistinguishable from cells isolated from lungs exposed to room air. These data reveal that type II cells exposed to in vivo hyperoxia have oxidized and fragmented DNA. Because type II cells are essential for lung remodeling, our findings raise the possibility that they are proficient in DNA repair.
Diabetogenic Effect of STZ Diminishes with the Loss of Nitric Oxide: Role of Ultraviolet Light and Carboxy-PTIO
Authors: Durán-Reyes Genoveva , Pascoe-Lira Dalila , Vilar-Rojas Cecilia , Medina-Navarro Rafael , Díaz-Flores Margarita , Ortega-Camarillo Clara , García-Macedo Rebeca , Cruz Miguel , Rodríguez Jesús Kumate
Published: April 2004

Nitric oxide has been demonstrated to participate in β-cell damage during streptozotocin (STZ)-induced diabetes. STZ consists of 2-deoxy-D-glucose substituted by N-methyl-N-nitrosourea at C-2 and therefore can liberate ·NO. However, it has not been proven whether ·NO generation from STZ is responsible for the disease. We found that STZ treated in vitro with ultraviolet (UV) light liberated significantly more ·NO than non-irradiated STZ (1,134.4 ± 104 vs. 256.9 ± 240 nmol). Moreover, the diabetogenic effect of STZ was abolished by UV irradiation before its administration to experimental animals. In these animals the glucose and insulin values were significantly different from those of the diabetic group (151.3 ± 16.6 vs. 364.6 ± 63.4 mg/dl and 36.3 ± 17.9 vs. 0.08 ± 5.5 µIU/ml, respectively) and similar to those of the non-diabetic group (127.2 ± 34.1 mg/dl and 41.7 ± 13.9 µIU/ml, respectively). Carboxy-PTIO treatment returned glycemia to nearly normal levels in 60% of STZ-induced diabetic rats (157.5 ± 11.8 vs. 364.6 ± 63.6 mg/dl of the diabetic group). L-NAME and dexamethasone cannot return either glucose or insulin to normal levels. In conclusion, UV light increased ·NO liberation from STZ and suppressed its diabetogenic activity. It is possible that the diabetogenic activity of STZ is related to the liberation of nitric oxide from STZ, since carboxy-PTIO scavenger had a protective effect, while L-NAME and dexamethasone did not. It is possible that an increase in ·NO concentration into cell, independently of its endogenous or exogenous origin, can induce β-cell damage and diabetes.
Liver-Type Fatty Acid-Binding Protein Is Highly Expressed in Intestinal Metaplasia and in a Subset of Carcinomas of the Stomach without Association with the Fatty Acid Synthase Status in the Carcinoma
Authors: Hashimoto Takeaki , Kusakabe Takashi , Watanabe Kazuo , Sugino Takashi , Fukuda Takeaki , Nashimoto Atsushi , Honma Kei-ichi , Sato Yukio , Kimura Hideki , Fujii Hiroshi , Suzuki Toshimitsu
Published: April 2004

Objective: To investigate the relation of liver-type fatty-acid-binding protein (L-FABP) expression to the clinicopathological characteristics or the fatty acid synthase status of gastric cancers. Methods: L-FABP expression was examined immunohistochemically in 667 gastric cancers, 60 gastric adenomas, and non-neoplastic epithelium contiguous with cancer tissue including normal foveolae, intestinal metaplasia, regenerative epithelium, and gastric glands. Results: L-FABP was positive in 38% (high in 9% and low in 29%) of gastric cancers. It occurred preferentially in papillary carcinomas, female cases, and in patients under 50 years. In gastric cancers, L-FABP expression had no intimate correlation with the FAS status, and it showed no relationship with prognosis and cancer progression as indicated by venous and lymphatic permeation, and nodal or hepatic metastasis. Gastric tubular adenomas mainly revealed low (22%) expression of L-FABP while intestinal metaplasia showed the most frequent (>95%) and intense L-FABP expression. Normal foveolae and gastric glands showed no or less L-FABP expression. Conclusions: L-FABP is highly and intensely expressed in metaplasia and in a subset of gastric adenocarcinomas, without association with progression, prognosis and fatty acid synthase status of the carcinoma.
Depletion of Tumor-Infiltrating Macrophages Is Associated with Amphoterin Expression in Colon Cancer
Authors: Kuniyasu Hiroki , Sasaki Takamitsu , Sasahira Tomonori , Ohmori Hitoshi , Takahashi Tadateru
Published: April 2004

Macrophage infiltration into colon cancer and amphoterin expression in cancer cells was examined in 42 human colon cancers invading the subserosa. The mean number of infiltrating macrophages was significantly higher in Dukes’ B cases than in Dukes’ C cases (p = 0.0065). Tumors with few infiltrating macrophages (macrophage depletion) were significantly more frequent in Dukes’ C cases than in Dukes’ B cases (p = 0.0014). No Dukes’ C cases with relevant macrophage infiltration showed macrophage-cancer cell contact, whereas 5 Dukes’ B cases showed such contact (p < 0.0001). In human colon cancer cells implanted in the cecum of nude mice, KM12SM (highly metastatic) tumors yielded less macrophage infiltration and more liver metastases than KM12C (low risk of metastasis) tumors (14 ± 3 vs. 78 ± 32 and 24 ± 6 vs. 5 ± 3 per liver, respectively). Amphoterin expression was detected at high frequency in both Dukes’ B and C cases (p = 0.0684). In macrophage-depleted cases, amphoterin expression was significantly higher than that in non-depleted cases (p = 0.0015). To confirm biological effects of amphoterin on macrophages, an infiltration assay using the cell-layered Boyden chamber was done. Infiltration of PMA-treated U937 monocytes through the KM12SM cell layer was increased by pretreatment of KM12SM cells with amphoterin antisense S-oligodeoxynucleotide exposure. Moreover, extracted amphoterin inhibited PMA-U937 monocyte infiltration in a dose-dependent manner. Thus, amphoterin may play an important role in the inhibition of macrophage infiltration into colon cancer.
Lung Pathology in Response to Repeated Exposure to Staphylococcus aureus in Congenic Residual Function Cystic Fibrosis Mice Does Not Increase in Response to Decreased CFTR Levels or Increased Bacterial Load
Authors: Davidson Donald J. , Webb Sheila , Teague Peter , Govan John R. W. , Dorin Julia R.
Published: April 2004

Objective: To establish the role of defects in murine Cftr in the susceptibility to Staphylococcus aureus lung disease using mouse models of cystic fibrosis (CF), congenic or inbred strains. Methods: We describe the histopathological analyses of CF mice repeatedly exposed by aerosolisation to a CF isolate of S. aureus, using residual function Cftr mice and compound heterozygotes generated by intercrossing these with Cftr ‘null’ mice, all congenic on the C57Bl6/N background. Results: We demonstrate that mice congenic on the C57Bl/6 background develop significantly more severe lung pathology than non-CF littermates in response to repeated exposure to the most frequent early CF lung pathogen S. aureus. Furthermore, reducing the level of Cftr by half in compound heterozygote mice does not impact upon disease severity, even in response to an increased bacterial dose. Conclusions: These results are consistent with an airway clearance defect, or abnormal inflammatory response secondary to Cftr mutation. These studies confirm the primary role for Cftr mutation in the development of this lung phenotype. In addition, these results demonstrate that a further 50% decrease in residual wild-type Cftr mRNA levels in this model does not impact the severity of the histopathological response to S. aureus, suggesting a critical threshold level for functional CFTR.
Characteristics and Trends of Clarithromycin-Resistant Helicobacter pylori Isolates in Japan over a Decade
Authors: Masuda H. , Hiyama T. , Yoshihara M. , Tanaka S. , Haruma K. , Chayama K.
Published: April 2004

Clarithromycin has been administered to patients in Japan since 1991. Clarithromycin-resistant Helicobacter pylori strains have been on the rise in Japan. We obtained H. pylori isolates between 1989 and 2000 and examined mutations of the 23S rRNA gene, which are closely associated with clarithromycin resistance. Isolates were obtained from 356 patients with H. pylori infection treated at the Hiroshima University. Sixty-one of the patients received clarithromycin-based H. pylori eradication therapy. Mutations of the 23S rRNA gene were examined by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) followed by sequencing analysis. Mutant strains were found in 42 of the 356 patients (11.8%). The prevalence of mutant strains increased from 0 to 20.4% during the 12-year study period. The prevalence increased to more than 10% by 1995 and then to more 20% after 1999. The H. pylori eradication rate was significantly higher in patients with wild-type strains than in patients with mutant strains (72.0 vs. 36.4%, p = 0.024). Our data indicate that clarithromycin-resistant H. pylori strains have increased rapidly since 1995 and that the effectiveness of clarithromycin-based H. pylori eradication therapies may soon be compromised. Other new therapies may be necessary as first-line treatments in Japan.
Expression of the Components of the Cip/Kip Family in Malignant Lymphoma of the Thyroid
Authors: Ito Yasuhiro , Yoshida Hiroshi , Matsuzuka Fumio , Matsuura Nariaki , Nakamura Yasushi , Nakamine Hirokazu , Kakudo Kennichi , Kuma Kanji , Miyauchi Akira
Published: April 2004

Objective: For evaluating the characteristics of human neoplasms, investigation of regulators of cell cycle progression is important. We studied three components of the Cip/Kip family, p57, p27 and p21, in malignant lymphomas of the thyroid. Methods: We immunohistochemically investigated the expression of the three proteins for 49 cases of thyroid lymphomas, as well as 10 cases of chronic thyroiditis. Results: In infiltrating lymphocytes in chronic thyroiditis, p57 and p27 were diffusely expressed, whereas p21 immunoreactivity was only occasionally observed. In lymphoma, decreased expression of p57 (less than 50% of lymphoma cells were positive) was observed in 24 cases (49.0%). p57 expression was inversely linked to the MIB-1 labeling index, grade of malignancy and aberrant p53 expression. Decreased expression of p27 was observed in 25 cases (51.0%), which was also related to the MIB-1 labeling index and aberrant p53 expression. On the other hand, p21 expression was occasional, and when the cut-off value was set at 3%, 38 cases (77.6%) were regarded as negative. P21 expression was not linked to any clinicopathological parameters. Conclusion: These findings suggest that, among the 3 components of the Cip/Kip family, p57 and p27 play a role in negatively regulating the cell proliferation of thyroid lymphoma cells and decreased expression of them contributes to the progression of this disease.
Detection of Treponema pallidum in Skin Lesions of Secondary Syphilis and Characterization of the Inflammatory Infiltrate
Authors: Wenhai Li , Jianzhong Zhang , Cao Ying
Published: April 2004

Background: Syphilis is an ancient sexually transmitted disease. However, the pathogenesis of mucocutaneous lesions of secondary syphilis is not completely understood. Methods: We analyzed the presence of Treponema pallidum in formalin-fixed, paraffin-embedded biopsy specimens from mucocutaneous lesions of secondary syphilis using highly sensitive nested polymerase chain reaction (PCR). The inflammatory infiltrates from the same specimens are also characterized using immunohistochemical methods. Results and Conclusions: Ten out of 24 (41.7%) specimens are T. pallidum positive using nested PCR, whereas none of them is T. pallidum positive using traditional silver staining. The presence of T. pallidum in the mucocutaneous lesions indicates that mucocutaneous lesions of secondary syphilis might be caused by direct T. pallidum invasion rather than by an allergic reaction. Furthermore, the majority of inflammatory infiltrating cells are CD45RO-positive T cells and CD68-positive macrophages, suggesting that cellular immunity plays an important role in the host reaction against T. pallidum infection in secondary syphilis.
Association between Chronic Urticaria and Thyroid Autoimmunity: A Prospective Study Involving 99 Patients
Authors: Verneuil L. , Leconte C. , Ballet J.J. , Coffin C. , Laroche D. , Izard J.P. , Reznik Y. , Leroy D.
Published: April 2004

Background: The association between chronic urticaria and thyroid autoimmunity has been a subject of debate. However, this link was suggested in studies searching thyroid microsomal antibodies (TMA), which are less sensitive and less specific than anti-thyroperoxidase antibodies, moreover these studies did not measure anti-TSH receptor antibodies, nor did they use a control group. As a consequence, the results of these studies are difficult to interpret. Objective: The aim of this study was to determine whether chronic urticaria is statistically associated with thyroid autoimmunity. Methods: In a prospective case-control study, we compared the frequency of thyroid autoantibodies in 45 patients with chronic urticaria and in 30 healthy adult volunteers; we also compared the frequency of chronic urticaria in 32 patients with thyroid diseases with thyroid autoantibodies and in 22 patients with thyroid diseases without thyroid autoantibodies. Thyroid autoantibodies and thyroid hormones were measured in all the subjects; antinuclear antibodies, rheumatoid factors, complement, IgE were assessed and routine laboratory tests were done in patients with chronic urticaria. Fisher’s exact statistics were used to test our hypothesis. Results: The frequency of thyroid autoantibodies was significantly higher in patients with chronic urticaria than in healthy controls (26.7%/3.3%; p < 0.01). All the patients with thyroid autoantibodies had thyroid hormone concentrations within the normal limits. The frequency of chronic urticaria was not significantly different (12.5%/9.1%; p = 0.7) in patients with thyroid diseases with or without thyroid antibodies. The rest of the biological investigations revealed only 1 patient with connective tissue disease. Conclusion: This study shows a significant association between chronic urticaria and thyroid autoimmunity, and that tests to detect thyroid autoantibodies are relevant in patients with chronic urticaria, whereas extensive laboratory tests are not.