bluuebunny/crossref_metadata_2025 huggingface dataset:
Reactivity of Dehydrometallophthalocyanines and ‐porphyrazines
Authors: Vagin Sergei I. , Frickenschmidt Antje , Kammerer Bernd , Hanack Michael
Published: October 2006

The zinc dehydrophthalocyanine 2 and zinc dehydrobenzoporhyrazine 8 a were generated from the 1N‐aminobenzotriazole‐annulated zinc phtalocyanine 1 and zinc benzoporhyrazine 8, respectively, by oxidation with Pb(OAc)4 in different solvents, for example, diethyl ether, tetrahydrofuran, acetic acid, and benzene. The reactivity of 2 and 8 a was studied in detail. These species not only easily undergo Diels–Alder additions with dienes, but also the used solvents can be added. Among the addition products with solvents ethoxy‐, acetoxy‐, acetoxybutyloxy‐substituted and barrelen‐fused phthalocyanines and benzoporpyrazines were isolated. No products resulting from the dimerization of two dehydro species were observed either for 2 or 8 a. Analysis of the reaction products in comparison with those obtained by oxidation of 1‐aminobenzotriazole 15 under similar conditions proves a higher reactivity (electrophilicity) of the dehydro‐PcZn 2 and dehydro‐PzZn 8 a in comparison with unsubstituted benzyne towards the solvents used, such as diethyl ether and benzene.
Synthesis and Reactivity of Rare Earth Metal Alkyl Complexes Stabilized by Anilido Phosphinimine and Amino Phosphine Ligands
Authors: Liu Bo , Cui Dongmei , Ma Jia , Chen Xuesi , Jing Xiabin
Published: October 2006

Anilido phosphinimino ancillary ligand H2L1 reacted with one equivalent of rare earth metal trialkyl [Ln{CH2Si(CH3)3}3(thf)2] (Ln=Y, Lu) to afford rare earth metal monoalkyl complexes [L1LnCH2Si(CH3)3(THF)] (1 a: Ln=Y; 1 b: Ln=Lu). In this process, deprotonation of H2L1 by one metal alkyl species was followed by intramolecular CH activation of the phenyl group of the phosphine moiety to generate dianionic species L1 with release of two equivalnts of tetramethylsilane. Ligand L1 coordinates to Ln3+ ions in a rare C,N,N tridentate mode. Complex l a reacted readily with two equivalents of 2,6‐diisopropylaniline to give the corresponding bis‐amido complex [(HL1)LnY(NHC6H3iPr2‐2,6)2] (2) selectively, that is, the CH activation of the phenyl group is reversible. When 1 a was exposed to moisture, the hydrolyzed dimeric complex [{(HL1)Y(OH)}2](OH)2 (3) was isolated. Treatment of [Ln{CH2Si(CH3)3}3(thf)2] with amino phosphine ligands HL2‐R gave stable rare earth metal bis‐alkyl complexes [(L2‐R)Ln{CH2Si(CH3)3}2(thf)] (4 a: Ln=Y, R=Me; 4 b: Ln=Lu, R=Me; 4 c: Ln=Y, R=iPr; 4 d: Ln=Y, R=iPr) in high yields. No proton abstraction from the ligand was observed. Amination of 4 a and 4 c with 2,6‐diisopropylaniline afforded the bis‐amido counterparts [(L2‐R)Y(NHC6H3iPr2‐2,6)2(thf)] (5 a: R=Me; 5 b: R=iPr). Complexes 1 a,b and 4 a–d initiated the ring‐opening polymerization of d,l‐lactide with high activity to give atactic polylactides.
Resonance Raman Studies of β‐Substituted Porphyrin Systems with Unusual Electronic Absorption Properties
Authors: Walsh Penny J. , Gordon Keith C. , Wagner Pawel , Officer David L.
Published: October 2006

ZnII and CuII porphyrins with β‐conjugated barbiturate functional groups have low‐energy electronic transitions which are unusual in that there are two strong bands in the Soret region. Resonance excitation of the two bands shows that each has features characteristic of both the porphyrin and barbiturate groups, with some perturbation to these features caused by the interaction of the two chromophores. The resonance Raman (RR) spectrum (λexc=413.1 nm) of the 412 nm band shows two bands at 1722 and 1743 cm−1 attributable to CO stretches in the substituent. Changes in frequency of porphyrin core modes due to the differing metal centres are reproduced by density functional theory calculations. The Q band RR spectra show modes with anomalous polarization which may be attributed to A2g modes, however no overtone or combination bands are observed.
Whole insect and mammalian embryo imaging with confocal microscopy: Morphology and apoptosis
Authors: Zucker Robert M.
Published: October 2006

Background:

After fluorochromes are incorporated into cells, tissues, and organisms, confocal microscopy can be used to observe three‐dimensional structures. LysoTracker Red (LT) is a paraformaldehyde fixable probe that concentrates into acidic compartments of cells and indicates regions of high lysosomal activity and phagocytosis, which both correlate to apoptosis activity. LT has been shown to be an indicator of apoptotic cell death which is correlated to other standard apoptotic assays.

Methods:

The mammalian samples were stained with LT, fixed with paraformaldehyde/glutaraldehyde, dehydrated with methanol (MEOH), and cleared with benzyl alcohol/benzyl benzoate (BABB). Following this treatment, the tissues were nearly transparent. Mosquitoes were fixed with MEOH and stained with propidium iodide. Next the tissues were dehydrated with MEOH and cleared with BABB.

Results:

Tissues as thick as 500 μm can be visualized after clearing with BABB. LT staining revealed apoptotic regions in mammalian limbs, fetuses, and embryos. Morphological observation of insect tissue consisted of combining autofluorescence with either nucleic acid staining (either propidium iodide or ethidium bromide).

Conclusions:

The use of BABB matches the RI of the tissue within the suspending medium. It helps in increasing the penetration of laser light in a confocal microscope by reducing the amount of light scattering artifacts and allows for the visualization of morphology in thick tissues. LT is a probe that stains the acid regions of tissues and cells and has been correlated to apoptosis. Morphological features of a tissue or organism (embryo, mosquito larvae) can be elucidated by fixation aldehydes, autofluorescence, and red‐emitting probes. This sample preparation procedure with optimization of confocal laser scanning microscopy allowed for the detection and visualization of apoptosis in fetal limbs and embryos which were ∼500‐μm thick. © 2006 International Society for Analytical Cytology
Muscle‐specific expression of the smyd1 gene is controlled by its 5.3‐kb promoter and 5′‐flanking sequence in zebrafish embryos
Authors: Du Shao Jun , Rotllant Josep , Tan Xungang
Published: October 2006

Zebrafish SmyD1 is a SET and MYND domain‐containing protein that plays an important role in myofiber maturation and muscle contraction. SmyD1 is required for myofibril organization and sarcomere assembly during myofiber maturation. Whole‐mount in situ hybridization revealed that smyd1 mRNAs are specifically expressed in skeletal and cardiac muscles in zebrafish embryos. However, it is unknown if smyd1 is expressed in other striated muscles, such as cranial and fin muscles, and moreover, the regulatory elements required for its muscle‐specific expression. We report here the analyses of smyd1 expression using smyd1‐gfp transgenic zebrafish. smyd1‐gfp transgenic zebrafish were generated using the 5.3‐kb smyd1 promoter and its 5′‐flanking sequence. GFP expression was found in the skeletal and cardiac muscles of smyd1‐gfp transgenic embryos. GFP expression appeared stronger in slow muscles than fast muscles in transgenic zebrafish larvae. In addition, GFP expression was also detected in cranial and fin muscles of smyd1‐gfp transgenic zebrafish larvae. In situ hybridization confirmed smyd1 mRNA expression in these tissues, suggesting that the expression of the smyd1‐gfp transgene recapitulated that of the endogenous smyd1 gene. Deletion analysis revealed that the 0.5‐kb sequence in the proximal promoter of smyd1 was essential for its muscle specificity. Together, these data indicate that smyd1 is specifically expressed in most, if not all, striated muscles, and the muscle specificity is controlled by the 5.3‐kb promoter and flanking sequences. Developmental Dynamics 235:3306–3315, 2006. © 2006 Wiley‐Liss, Inc.
Development of an anti‐IL‐17A auto‐vaccine that prevents experimental auto‐immune encephalomyelitis
Authors: Uyttenhove Catherine , Van Snick Jacques
Published: October 2006

IL‐17 has been associated with multiple inflammatory disorders such as rheumatoid arthritis, asthma and multiple sclerosis. As these diseases require long‐term treatment we turned to an auto‐vaccine strategy for IL‐17 neutralization in vivo. Mouse IL‐17A was covalently linked to ovalbumin and used to immunize C57BL/6 mice. This vaccine induced the production of antibodies that blocked IL‐17A bioactivity in vitro but did not react with the other IL‐17 isoforms, including IL‐17F. As the half‐life of the Ab titers after the last immunogen administration was approximately 4 months, the vaccine provides for long lasting and selective inhibition of IL‐17A activity in vivo. A monoclonal Ab (mAb) derived from these mice showed the same specificity for IL‐17A. To test the ability of the vaccine to confer protection against an IL‐17‐dependent disorder, SJL mice were vaccinated with IL‐17‐OVA and encephalomyelitis (EAE) was induced by proteolipid protein (PLP) peptide 139–151. Vaccinated mice were completely protected against the disease. The above‐mentioned anti‐IL‐17A mAb also prevented EAE development. The absence of clinical symptoms contrasted with unaltered PLP‐induced cytokine production in vitro and unmodified anti‐PLP IgG titers and isotypes. These results suggest that an anti‐IL‐17A auto‐vaccine offers new perspectives for therapy of autoimmune diseases.

See accompanying commentary: http://dx.doi.org/10.1002/eji.200636760
Hopeanol: A Potent Cytotoxin with A Novel Skeleton from Hopea exalata
Authors: Ge Hui Ming , Xu Chen , Wang Xiao Ting , Huang Bo , Tan Ren Xiang
Published: October 2006

Hopeanol, a rearranged resveratrol dimer ester with an unprecedent carbon skeleton, was isolated from the bark ofHopea exalata. Its structure was determined by comprehensive spectroscopic analysis. Hopeanol exhibited potent cytotoxicity against six human cancer cell lines with its IC50 = 0.52–19.36 μM. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2006)
Hydrological processes and streamflow in a lake dominated watercourse
Authors: Spence Christopher
Published: October 2006

This paper discusses results of a nested basin study investigating run‐off responses along the watercourse in a 137 km2 subarctic Canadian Shield watershed. Catchment areas contributing surface run‐off to the basin outlet shrink after a maximum during spring snowmelt because of storage demands in headwater wetlands and lakes. This decrease is significant as summer contributing areas to the basin outlet are only 4% the size of spring melt contributing areas. This reduction in contributing area and decrease in antecedent lake levels lead to a two order of magnitude decrease in run‐off ratios from spring snowmelt to midsummer. Furthermore, the attenuating influence of lakes on the run‐off hydrograph increases the opportunity for evaporative losses, which prevents small run‐off events from maintaining a measurable flood wave through the basin. The location at which a run‐off event is interrupted in the watercourse is a function of lake size relative to upstream run‐off inputs. Run‐off is more susceptible to evaporative losses in large receiving lakes where inflow per unit area is less. These processes cause intermittent streamflow through the basin. These results have repercussions for coupled atmospheric/hydrological models whose resolution is approaching the basin scale. The incorporation of lateral transfer schemes that encompass these processes and physiographic influences on streamflow response is encouraged. Copyright © 2006 Crown in the right of Canada. Published by John Wiley & Sons, Ltd.
Energy and water vapour exchanges over a mixedwood boreal forest in Ontario, Canada
Authors: Pejam M. R. , Arain M. A. , McCaughey J. H.
Published: October 2006

Measurements of energy, carbon dioxide and water vapour fluxes and supporting meteorological variables were made from 1 September 2003 to 31 August 2004 in a 74‐year‐old mixedwood forest in northern Ontario, Canada, using the eddy covariance technique. Land surface cover data analysis showed that although this forest is comprised of deciduous and coniferous species, their proportion throughout the forest was almost equal, resulting in a homogeneously mixedwood stand. The seasonal pattern of albedo showed a distinct pattern as a result of mixed deciduous and coniferous species with the first low radiation reflection period in early spring (between snowmelt and deciduous leaf‐out), and the second low reflection period in late autumn (between senescence and snowfall). Root zone soil water content remained above 0·32 throughout the growing season, which is characteristic of wet soil conditions. The evaporative flux exceeded the sensible heat flux during most of the growing season. Daily mean evapotranspiration values during the peak growing season were about 2–2·5 mm day−1 with maximum values reaching up to 4–5 mm day−1 on sunny days. The total water loss over the 12‐month measurement period was 480 ± 30 mm, while the total precipitation was 835 mm. Despite high soil water content, bulk surface conductance calculated using the inverted Penman–Monteith formulation, showed a strong correlation with vapour pressure deficit (VPD), indicating stomatal control on water loss during the afternoons with high atmospheric demand and during occasional dry periods. Bulk surface conductance values in this mixedwood forest were relatively higher than those in other boreal coniferous forests and were more similar to values observed in the southern boreal deciduous forests. The Priestley–Taylor α values showed a wide range but during most of the growing season its values were close to unity, indicating that water stress does not play a major role in the overall water loss from this boreal forest ecosystem, while energy supply has a strong control on evapotranspiration. These results will help parameterize accurately energy and water exchanges for the moist mixedwood forests in land surface‐atmosphere interaction and hydrologic models. Copyright © 2006 John Wiley & Sons, Ltd.
Lead‐induced experimental lesions of the testis and their treatment
Authors: Ghelberg Naum W. , Bordas Emeric
Published: October 2006

Thirty‐six healthy albino rats were divided into three groups and the histologic structure of the testes was examined after treatment: group ‘C’–10 healthy control rats were untreated; group ‘Pb’ – eight rats received intraperitoneal injections of 8.5 mg per kg body weight lead acetate for 154 days (15 doses in all); group ‘PbA’ – eight rats intoxicated with lead, like group ‘Pb’ but which, during the last 21 days before killing, were treated with 5 ml per kg body weight per day of a 2% injectable solution of sodium aspartate. As compared with the controls, group ‘Pb’ showed alteration of the histopathologic pattern and reduction in the size of seminiferous tubules, basal membrane separation and a decrease in the cell content of the germinal layer, lesions of the spermatocytes and spermatids, and slight oedematous dissociation in the interstice. In 75% of the group ‘PbA’ animals, normal aspects similar to those of group ‘C’ were found.
Mutagenicity study on mice given mercuric chloride
Authors: Poma Kathleen , Kirsch‐Volders Micheline , Susanne Charles
Published: October 2006

The mutagenic activity of different roncentrations of mercuric chloride (0, 2, 4 and 6 mg HgCl2 per kg body wt) has been analysed in Swiss Albino mice at several periods (12, 24, 36 and 48 h) after intraperitoneal injection. No increased frequency of chromosomal aberrations was observed in bone marrow cells or in spermatogonia.
The metabolism and distribution of 4,4′‐Methylene‐bis(2‐chloroaniline) (MBOCA) in rats
Authors: Farmer P. B. , Rickard J. , Robertson S.
Published: October 2006

4,4′‐Methylene‐bis(2‐chloroaniline) (MBOCA), a known carcinogen in several animal species, is frequently found in the urine of humans exposed to the compound in the plastics industry. A knowledge of the metabolism and tissue distribution of MBOCA would be of value in monitoring exposure to this potential human carcinogen. Following intraperitoneal (i.p.) or oral administration of [14C]MBOCA to rats, approximately one third of the radioactivity was excreted in the urine. Only 1–2% of the radioactivity was identified as MBOCA by high performance liquid chromatography (HPLC) and gas chromatography–mass spectrometry. At least nine other metabolites were separated by reversed‐phase HPLC. Treatment of the urine with sulphatase–glucuronidase liberated two major deconjugated metabolites. No evidence was obtained for any major long‐term retention of MBOCA in the rat, although 48h after dosing approximately 2% of the administered radioactivity was associated with the liver. Despite the extensive metabolism of MBOCA in the rat, analysis of the urine of exposed workers showed only MBOCA at concentrations up to 1500 nmol 1−1. No evidence was found for the major metabolites identified in the rat.
Porous polylactide/chitosan scaffolds for tissue engineering
Authors: Wan Ying , Fang Ya , Wu Hua , Cao Xiaoying
Published: October 2006

Novel porous scaffolds were fabricated using biodegradable polylactide/chitosan blends. A combinational technique involving solvent‐extracting, liquid‐solid separation, and freeze‐drying paths were employed. The processing parameters were optimized in order to produce desired porous scaffolds and thus obtained scaffolds showed well distributed and interconnected porous structures with controllable porosities varying from around 50–85% and regulative pore sizes being distributed within a region between 2 and 190 μm. These scaffolds exhibited remarkably improved hydrophilicity based on the measurements for their swelling index. The results obtained from in vitro incubation of scaffolds in phosphate buffered saline solutions at 37°C during various periods up to 10 weeks indicated that chitosan component inside the scaffolds, on the one hand, effectively buffered the acidic degradation products of polylactide/chitosan scaffolds and on the other hand, the degradation of the scaffolds was also conspicuously delayed. These porous scaffolds maintained well‐defined compressive mechanical properties and by well‐blending polylactide with chitosan component, improved toughness on the resultant porous scaffolds was also observed. © 2006 Wiley Periodicals, Inc. J Biomed Mater Res, 2006
Changes in collagen cross‐linking in bleomycin‐induced pulmonary fibrosis
Authors: Reiser Karen M. , Tryka A. Francine , Lindenschmidt Robert C. , Last Jerold A. , Witschi Hanspeter R.
Published: October 2006

Collagen cross‐linking was analyzed in lungs of rats two, four, and ten weeks after intratracheal instillation of 1.5 units of bleomycin. Similar analyses were performed on lungs of mice 18 months after intratracheal instillation of bleomycin with or without subsequent exposure to 70% oxygen (O2) for 72 hours. Lungs were analyzed to determine the content of the reduced difunctional cross‐links dihydroxylysinonorleucine (DHLNL) and hydroxylysinonorleucine (HLNL) and of the nonreducible trifunctional cross‐link hydroxypyridinium (OHP). Ratios of DHLNL:HLNL were elevated in the rat lungs at two and four weeks, due to increased levels of DHLNL. There were no changes in the difunctional cross‐links in any of the mouse lungs. Hydroxypyridinium content was elevated in the rat lungs at ten weeks and in the mouse lungs exposed to bleomycin and oxygen. We conclude that increases in DHLNL may serve as an early indicator that potentially “fibrotic collagen” is being synthesized in lungs acutely exposed to fibrogenic stimuli, while increases in OHP may serve as a permanent marker of a fibrogenic event that could have occurred months to years earlier.
The binding of reactive metabolites of the carcinogen N‐hydroxy‐2‐acetylaminofluorene to DNA and protein in isolated rat liver nuclei: Effects of glutathione and methionine
Authors: Meerman John H. N. , Tijdens Roeline B. , Mulder Gerard J.
Published: October 2006

The covalent binding of reactive metabolites of the carcinogen N‐hydroxy‐2‐acetylaminofluorene to DNA and protein in isolated, intact rat liver nuclei was studied. The chemically synthesized 2‐acetylaminofluorene‐N‐sulfate became covalently bound to DNA and protein to form adducts, 50% to 60% of which retained the N‐acetyl group. Glutathione decreased the covalent binding of acetylated adducts to DNA by 18% and to protein by 50%. Methionine was more effective; it decreased DNA binding by 52% and protein binding by 79%.

N‐Hydroxy‐2‐acetylaminofluorene was deacetylated by the nuclear preparation. Almost exclusively, deacetylated 2‐aminofluorene adducts to DNA and protein were formed. Glutathione decreased the covalent binding of deacetylated adducts to DNA by only 14%. Protein binding, however, was decreased by 57%. Methionine had no effect on the formation of these adducts to DNA and protein. Formation of 2‐aminofluorene‐glutathione conjugates was reduced by ascorbic acid by 65%. Covalent binding of deacetylated adducts to DNA and protein, however, was not decreased by ascorbic acid.

These data suggest that “harder” nucleophiles like methionine can be used to protect macromolecules in vivo from damage by “hard” electrophiles such as those generated from the reactive 2‐acetylaminofluorene‐N‐sulfate. However, such nucleophiles seem not to be effective with N‐hydroxylamines, such as N‐hydroxy‐2‐aminofluorene, formed by deacetylation of N‐hydroxy‐2‐acetylaminofluorene.
Stereoselective formation of a cholesterol ester conjugate from fenvalerate by mouse microsomal carboxyesterase(s)
Authors: Miyamoto Junshi , Kaneko Hideo , Takamatsu Yasuko
Published: October 2006

In accordance with in vivo findings, of the four chiral isomers of fenvalerate (S‐5602 SumicidinR, PydrinR, [RS]‐α‐cyano‐3‐phenoxybenzyl [RS]‐2‐(4‐chlorophenyl)isovalerate), only the [2R,αS]‐isomer (B‐isomer) yielded cholesteryl [2R]‐2‐(4‐chlorophenyl)isovalerate (CPIA‐cholesterol ester) in the in vitro study using several tissue homogenates of mice, rats, dogs, and monkeys. There were species differences in the extent of CPIA‐cholesterol‐ester formation, with mouse tissues showing relatively higher activity than those of other animals. The kidney, brain, and spleen of mice showed relatively higher capacities to form this ester compared to other tissues, and the enzyme activity was mainly localized in microsomal fractions. The CPIA‐cholesterol ester did not seem to be produced by three known biosynthetic pathways of endogenous cholesterol esters–acyl‐CoA:cholesterol O‐acyltransferase (ACAT),2 lecithin:cholesterol O‐acyltransferase (LCAT)3 and cholesterol esterase.4 Carboxyesterase(s) of mouse kidney microsomes solubilized by digitonin hydrolyzed only the Bα‐isomer of fenvalerate, yielding CPIA, whereas they yielded the corresponding cholesterol ester in the presence of artificial liposomes containing cholesterol. Thus, it appears that the stereoselective formation of the CPIA‐cholesterol ester results from the stereoselective formation of the CPIA‐carboxyesterase complex only from the Bα‐isomer, which subsequently undergoes cleavage by cholesterol to yield the CPIA‐cholesterol ester.
Effects of cadmium in vitro on microsomal steroid metabolism in the inner and outer zones of the guinea pig adrenal cortex
Authors: Colby H. D. , Pope M. R. , Johnson P. B. , Sherry J. H.
Published: October 2006

Studies were carried out to evaluate the effects of cadmium in vitro on microsomal steroid metabolism in the inner (zona reticularis) and outer (zona fasciculata and zona glomerulosa) zones of the guinea pig adrenal cortex. Microsomes from the inner zone have greater 21‐hydroxylase than 17α‐hydroxylase activity, resulting in the conversion of progesterone primarily to 11‐deoxycorticosterone and of 17α‐hydroxy progesterone principally to its 21‐hydroxylated metabolite, 11‐deoxycortisol. Microsomes from the outer zones, by contrast, have far greater 17α‐hydroxylase and C17,20‐lyase activities than 21‐hydroxylase activity. As a result, progesterone is converted primarily to its 17‐hydroxylated metabolite, 17α‐hydroxyprogesterone; and 17α‐hydroxyprogesterone is converted principally to δ4‐androstenedione, with only small amounts of 21‐hydroxylated metabolites being produced. Addition of cadmium to incubations with inner zone microsomes causes concentration‐dependent decreases in 21‐hydroxylation and increases in 17α‐hydroxylase and C17,20‐lyase activities, resulting in a pattern of steroid metabolism similar to that in normal outer zone microsomes. Cadmium similarly decreases 21‐hydroxylation by outer zone microsomes but has no effect on the formation of 17‐hydroxylated metabolites or on androgen (Δ4‐androstenedione) production. In neither inner nor outer zone microsomes did cadmium affect cytochrome P‐450 concentrations, steroid interactions with cytochrome(s) P‐450, or NADPH–cytochrome P‐450 reductase activities. The results indicate that cadmium produces both quantitative and qualitative changes in adrenal microsomal steroid metabolism and that the nature of the changes differs in the inner and outer adrenocortical zones. In inner zone microsomes, there appears to be a reciprocal relationship between 21‐hydroxylase and 17α‐hydroxylase/C17,20‐lyase activities which may influence the physiological function(s) of that zone.
Inhibition of prostaglandin H synthase–catalyzed cooxidation of diethylstilbestrol by α‐naphthoflavone and β‐naphthoflavone
Authors: Degen Gisela H.
Published: October 2006

Prostaglandin H synthase (PHS) has gained interest as a drugmetabolizing enzyme and has been shown to cooxidize and metabolically activate diethylstilbestrol (DES) in vitro. Both 7,8‐benzoflavone (α‐naphthoflavone, ANF) and 5,6‐benzoflavone (β‐naphthoflavone, BNF) have now been studied for their effects on PHS from ram seminal vesicle microsomes by means of several in vitro assays.

The PHS‐catalyzed cooxidation of DES, as measured by high‐performance liquid chromatography (HPLC) analysis, is inhibited by BNF and ANF at micromolar concentrations, with median inhibitory concentrations (IC‐50) of<20 and 40 μM, respectively. The oxidation of DES is inhibited whether it is initiated by arachidonic acid or by hydrogen peroxide, indicating that the benzoflavones inhibit PHS by a mechanism different from that of indomethacin. Monitoring of cyclooxygenase activity in an oxygraph also reveals an inhibition of PHS by BNF which depends only weakly on arachidonic acid concentration; inhibition by ANF is less pronounced under these conditions. Since PHS‐catalyzed conversion of the benzoflavone compounds was detected under conditions permitting cooxidation, the inhibition of PHS by benzoflavones in vitro could either be a direct effect or possibly mediated via metabolites.

Our data imply that ANF and BNF, in addition to their well‐known role as modifiers of mixed‐function oxidases, can affect the PHS‐catalyzed metabolism of xenobiotics. This is discussed in the context of adverse effects caused by DES in vivo and in cell culture and must be taken into account when interpreting the modifying effect of benzoflavones on these endpoints.
Evaluation of the calcium mobilizing action of acetaminophen and bromobenzene in rat hepatocyte cultures
Authors: Long R. M. , Moore L.
Published: October 2006

Acetaminophen (APAP) and bromobenzene (BrB) are reported to selectively inhibit plasma membrane (PM) but not endoplasmic reticulum (ER) Ca2+ transport in rat liver (1). The ability of these hepatotoxicants to increase cytoplasmic Ca2+ levels as a result of disrupted Ca2+ pumping was determined in cultured rat hepatocytes by monitoring the activity of glycogen phosphorylase a, a Ca2+ ‐sensitive (via phosphorylase kinase) enzyme. Following exposure to 2.5 to 10 mM APAP for five minutes, dose‐dependent increases in phosphorylase a activity were observed (58 to 190 U/g protein). Endoplasmic reticulum Ca2+ pump activity was not inhibited after any dose of APAP (56 nmol Ca2+ per milligram protein per 30 minutes). Phosphorylase a activity remained elevated for 60 minutes after exposure to APAP (124 μl/g protein). Following exposure to 0.5 to 2 mM BrB for five minutes, phosphorylase a activity also increased (58 to 229 U/g protein) in a dose‐related manner. Endoplasmic reticulum Ca2+ pump activity was inhibited after BrB exposure (from 58 to 16 nmol Ca2+ per milligram protein per 30 minutes). Phosphorylase a activity remained elevated for 60 minutes after exposure to BrB (147 U/g protein). Evidence of elevated cytoplasmic Ca2+ is consistent with the inhibition of Ca2+ ‐extruding/sequestering mechanisms at hepatocyte PM and/or ER. Prolonged elevation of cytosolic Ca2+ levels could overstimulate Ca2+ ‐sensitive processes within liver cells and thus initiate or contribute to hepatotoxic injury.
The inactivation of rhodanese by nitrite and inhibition by other anions in vitro
Authors: Alexander Keith , Procell Lawrence R. , Kirby Stephen D. , Baskin Steven I.
Published: October 2006

Cyanide detoxification in mammals occurs, in part, by sulfur transfer by rhodanese to form the less toxic thiocyanate. Thiosulfate and nitrite are often used in combination for the treatment of cyanide intoxication. This report shows that nitrite can inhibit the rate of sulfur transfer by rhodanese in vitro. Nitrate, chloride, sulfate, and acetate were also examined as inhibitors. Inhibition by nitrite appeared to be more complex than for the other anions tested. Closer examination showed that nitrite can inactivate the sulfur‐free rhodanese. Our observation leads to the suggestion that, in vivo, either rhodanese is maintained in its more stable sulfur‐substituted form or cellular compartmentalization prevents inactivation by nitrite.
Glucose and sulfate conjugation of phenolic compounds by the spiny lobster (panulirus argus)
Authors: Schell John D. , James Margaret O.
Published: October 2006

Previous work has shown that the hepatopancreas of the spiny lobster (Panulirus argus) contains a mixed‐function oxidase system capable of catalyzing the monooxygenation of polycyclic aromatic hydrocarbons to highly toxic products similar to those formed by mammalian tissues. Studies were designed to determine the ability of the spiny lobster to conjugate the phenolic compounds 4‐methylumbelliferone, p‐nitrophenol, β‐naphthol, and 3‐hydroxy‐benzo[a]pyrene with endogenous molecules. The hepatopancreas contained UDP‐glucose (UDPG) dependent glucosyltransferase, while no activity was detected when UDP‐glucuronic acid was used as the cosubstrate. Atypical Michaelis‐Menten kinetics result with varying concentrations of UDPG, indicating that multiple forms of glucosyltransferase may exist in this organ. The activity was localized in the microsomal fraction, exhibited a pH optimum at 8.0–8.5, and a temperature optimum of 30°C. Sulfate conjugation was found only in the cytosolic fraction of the antennal gland and used adenosine 3′‐phosphate 5′‐phosphosulfate (PAPS) as the sulfate donor (Km (apparent) =9.0 ± 4.9 μM). Hepatopancreas cytosol inhibited sulfotransferase activity. The pH optimum of antennal gland sulfotransferase was a function of the substrate and ranged from 5.5 to 7.4. Analysis of spiny lobster urine 24 hr following exposure to 3‐hydroxybenzo[a]pyrene demonstrated the ability of the lobster to form both the sulfate and glucoside conjugate in vivo.
Oxidative desulfuration of chlorpyrifos, chlorpyrifos‐methyl, and leptophos by rat brain and liver
Authors: Chambers Janice E. , Chambers Howard W.
Published: October 2006

The oxidative desulfuration of the three phosphorothionate insecticides—chlorpyrifos, chlorpyrifos‐methyl, and leptophos—was studied in rat brain and liver. Hepatic microsomes demonstrated activities of 4–28 nmol/g/min, with male activity 2‐ to 4‐fold higher than female activity. Very low desulfuration activity of all three compounds was observed in both microsomal and crude mitochondrial fractions from brain (3–27 pmol/g/min). There were no sex differences in the brain. Although the liver displayed 140‐ to 2100‐fold greater activity than brain on a wet‐weight basis, the brain desulfuration activities of these three compounds as well as those of some previously reported phosphorothionates generally correlate well with the toxicity and may be important in determining the overall acute toxicity levels of phosphorothionate insecticides.
Induction of peroxisomal enzymes in cultured porcine endothelial cells by the hypolipidemic drug ciprofibrate
Authors: Glauert Howard P. , Hennig Bernhard , Chow Hannah S.
Published: October 2006

The purpose of this study was to determine if the hypolipidemic peroxisome proliferator ciprofibrate, which induces peroxisomes in the liver, can induce peroxisomes in cultured porcine pulmonary endothelial cells. Ciprofibrate was added at three concentrations to cell cultures for a 6‐day period. The induction of peroxisomes in the cells was detected by determining total peroxisomal β‐oxidation and peroxisomal catalase activity. The addition of ciprofibrate was found to increase peroxisomal enzyme activities in a dose‐dependent manner, with the highest activity being reached at 1000 μM ciprofibrate. Ciprofibrate also caused an increased transfer of albumin across endothelial cells cultured on micropore filters. This study shows that peroxisomal enzyme activities can be induced by ciprofibrate in endothelial cells, which may have implications in diseases mediated by vascular injury.
Mirex induces ornithine decarboxylase in female rat liver
Authors: Mitra Ashoke , Richards Ira , Kitchin Kirk , Conolly Rory , Kulkarni Arun P.
Published: October 2006

Ornithine decarboxylase, the rate‐limiting enzyme in polyamine synthesis, was significantly induced in female rat liver following oral administration of the pesticide mirex. After dual oral exposure (120 mg /kg of mirex; 21 and 4 hr prior to sacrifice), ornithine decarboxylae activity in rat liver cytosol was 70‐fold higher than control values. A single oral dose of mirex (180 mg /kg) induced hepatic ornithine decarboxylase activity 55‐fold over controls. After a single oral dose of mirex the maximal induction of ODC activity occurred at 36 hr. Mirex is an unusually potent and long‐lasting inducer of rat hepatic ornithine decarboxylase activity.
The effects of ions on the conjugation of xenobiotics by the aralkyl‐coa and arylacetyl‐coa N‐acyltransferases from bovine liver mitochondria
Authors: Kelley Michael , Vessey Donald A.
Published: October 2006

The aralkyl‐CoA:glycine N‐acyltransferase and the arylacetyl‐CoA:amino acid of N‐acyltransferase were purified from bovine liver mitochondria and their response to a variety of ions investigated. The activity of the aralkyl transferase was inhibited by divalent cations and high concentrations of monovalent cations with all substrates investigated. For benzoyl‐coenzyme A (CoA), K+ was a competitive inhibitor, competing for binding at the benzoyl‐CoA binding site. With salicylyl‐CoA, K+ did increase the dissociation constant (KD) for acyl‐CoA but it was not a competitive inhibitor and in addition, K+ increased the Michaelis constant for glycine (Kglym) tenfold. The data suggest that the increase in Kglym is due to bound K+ forcing reorientation of salicylyl‐CoA at the active site so that it impinges on the glycine binding site. Inorganic anions and cations did not affect the extent of product inhibition by hippuric acid with either acyl‐CoA and this was because they affected the binding of acyl‐CoA and hippuric acid to the same extent. Ions did, however, greatly reduce the extent of product inhibition by CoA. This is critical because under approximate in vivo conditions (2.5 mM CoA), the salt‐free enzyme would be almost completely inhibited by CoA. The arylacetyl transferase was activated by inorganic ions when assayed at saturating substrate concentrations. However, at physiologic concentrations of glycine certain salts were modestly inhibitory. The inhibitory effect of KCl was characterized by a large decrease in the affinity of the enzyme for phenylacetyl‐CoA, suggesting that the arylacetyl‐CoA region of the active site contained an inhibitory ion binding site. At low (physiologic) concentrations of substrate, the arylacetyl transferase was extensively inhibited by CoA and this inhibition was greatly reduced by ions. The 3′‐phosphate group on CoA was found to be important for binding to the salt‐free enzyme but in the presence of ions its importance was diminished. In the absence of inorganic ions the affinity of the enzyme for phenylacetyl‐CoA and naphthylacetyl‐CoA was so high that it could not be measured. In the presence of KCl the KD values for phenylacetyl‐CoA and naphthylacetyl‐CoA were similar, but the Km for glycine was extremely high for 1‐naphthylacetyl‐CoA conjugation, which accounts for its slow rate of metabolism. Conjugation with glutamine had a high Michaelis constant for glutamine (KGlum) and a low maximum velocity (Vmax) which accounts for the absence of glutamine conjugation in vivo.
Oxidatively stressed lymphocytes remain in Go/Gla on mitogenic stimulation
Authors: Duncan David D. , Lawrence David A.
Published: October 2006

Thiol modifiers and oxidants inhibit lymphocyte activation. To investigate which of the many cell functions sensitive to oxidation are critical in this inhibition, mouse splenic lymphocytes were treated with oxidants prior to exposure to mitogen, and progression into the cell cycle was assayed. Different treatments were used to chemically dissect different potential targets within the cell: copper phenanthroline (CuP), to oxidize surface sulfhydryls; N‐ethyl maleimide (NEM), to alkylate extra‐ and intracellular thiols; and hydrogen peroxide, which generates the highly reactive hydroxyl radical within the cell. Progression into the cell cycle was assayed with acridine orange (AO) and assays of interleukin‐2 (IL‐2) production and IL‐2 receptor (IL‐2R) expression. The contribution of ADP‐ribosylation to inhibition of mitogenesis was assessed using 3‐aminobenzamide (3AB) to inhibit adenosine 5′‐diphosphate (ADP)‐ribose transferases. The results indicate that the CuP and NEM treatments both produce two independent inhibitory effects, that is, a failure in the production of and response to IL‐2. Cells treated with these compounds were able to progress only through G1a upon mitogenic stimulation. H2O2 had more complex effects. Both ADP‐ribosylation and modulations of cytosolic Ca2+ were involved in the inhibitory effects. With lower inhibitory doses of H2O2, lymphocytes were completely unresponsive to mitogen and failed to exit Go upon mitogenic stimulation. If intra‐ and extracellular Ca2+ were buffered before treatment with H2O2, higher concentrations were required, and under these conditions cells were able to enter G1a but could not progress into G1b. Under neither of these conditions could cells produce IL‐2 or express IL‐2R.
Alteration in de novo pyrimidine biosynthesis during uridine reversal of pyrazofurin‐inhibited dna synthesis
Authors: Ringer David P. , Howell Boyd A. , Etheredge Janet L.
Published: October 2006

Pyrazofurin, a pyrimidine nucleoside analogue with antineoplastic activity, inhibits cell proliferation and DNA synthesis in cells by inhibiting uridine 5′‐phosphate (UMP) synthase. It has been previously shown in concanavalin A (con A)‐stimulated guinea pig lymphocytes (23) that pyrazofurininhibited DNA synthesis could be selectively reversed by exogenous uridine (Urd). In this report, we have examined possible mechanisms for the Urd reversal with experiments that determine the ability of exogenous Urd to (a) interfere with either the intracellular transport of pyrazofurin, or the conversion of pyrazofurin to its intracellularly active form, pyrazofurin‐5′‐phosphate; (b) reverse the pyrazofurin block of [14C]orotic acid incorporation into DNA; and (c) alter the pattern of exogenous [3H]Urd incorporation into DNA‐thymine (DNA‐Thy) and DNA‐cytosine (DNA‐Cyt) during pyrazofurin inhibition of pyrimidine de novo biosynthesis. The results of these experiments showed that Urd reversal does not occur through altered pyrazofurin transport or intracellular conversion to pyrazofurin‐5′‐phosphate, nor does it alter the distribution of [3H]Urd in DNA‐Thy and DNA‐Cyt. Instead, these findings indicate that the primary mechanism for exogenous Urd reversal of pyrazofurin inhibition of DNA synthesis involves the reversal of pyrazofurin inhibition of UMP synthase, thus restoring orotic acid incorporation into lymphocyte DNA through the pyrimidine de novo pathway.
Fetotoxic alterations in the normal ontogenies of rat microsomal and lysosomal enzymes
Authors: Harris Craig , Thomas David E. , Carter Melvin W. , Bradshaw William S.
Published: October 2006

The activity patterns during development for acid phosphatase (Ac‐P), alkaline phosphatase (A1‐P), ß‐glucuronidase (ßG), and UDP‐glucuronyltransferase (UDPGT) have been determined in various tissues of the rat for corn oil and distilled water controls as well as in animals prenatally exposed to four fetotoxic chemicals. Postnatal assays were performed on both sexes separately. In control animals, tissue‐specific differences between male and female activity levels were found for UDPGT. In the liver of mature offspring, enzyme activity was greater in males than in females. Although no sex difference was observed in the intestine, the kidneys of females exhibited higher values than those of males. An original computer‐assisted methodology is presented, designed (a) to permit a mathematical description for the complex curves exhibited by these ontogeny profiles, and (b) to assess the statistical significance of chemical‐induced alterations in these complex developmental patterns, specifically, to target sensitive periods and subtle changes near the fetotoxic threshold. Oral administration (days 6–18 of gestation) of 3,3′,4,4′‐tetrachlorobiphenyl (4CB) to pregnant females resulted in an induction of liver UDPGT activity in offspring postnatally, and some alterations in the perinatal pattern of ßG in the same tissue. This treatment also produced differences in the intestinal patterns of Ac‐P and male UDPGT. No significant changes were observed in offspring exposed to diethylstilbestrol (DES). Treatment with zeranol (ZN) caused reductions in activity over the entire postnatal period for ßG in liver, brain, intestine, and kidney, for Al‐P in brain, and for Ac‐P in the intestine. Cadmium‐treated dams gave birth to offspring that exhibited slightly altered ontogenies only in intestine for UDPGT and AcP. The alterations in these developmental profiles indicate periods of increased sensitivity, and may be useful in directing more specific studies into the fetotoxic mechanisms of these compounds.
Pentachlorobutadienyl‐l‐cysteine (PCBC) toxicity: The importance of mitochondrial dysfunction
Authors: Groves Carlotta E. , Schnellmann Rick G. , Sokol Paul P. , Steffens Thomas G. , Lock Edward A.
Published: October 2006

The relationship between the covalent binding, uptake, and toxicity produced by pentachlorobutadienyl‐L‐cysteine (PCBC) was examined in rabbit renal proximal tubules (RPT), renal basolateral membrane vesicles, and isolated renal cortical mitochondria. Renal proximal tubules rapidly metabolized PCBC to a reactive intermediate that bound to tubular protein. Approximately 70–90% of PCBC found in the cell at any given time was bound to protein. PCBC initially uncoupled oxidative phosphorylation, followed by a 45% reduction of state 3 respiration and a 90% decrease in cellular adenosine triphosphate (ATP) levels. These events preceded cell death. Isolated mitochondria also metabolized PCBC to a reactive intermediate that bound to mitochondrial protein and initiated mitochondrial toxicity. These results show that. PCBC‐induced mitochondrial dysfunction occurred as a result of mitochondrial bioactivation and that the mitochondrion is the critical subcellular target in PCBC toxicity. Aminooxyacetic acid (AOAA), an inhibitor of cysteine conjugate β‐lyase, reduced the covalent binding of PCBC‐equivalents to tubular protein by approximately 90% and decreased but did not prevent the toxic effects produced by PCBC on RPT respiration and cellular ATP levels. AOAA delayed but had no effect on the overall extent of cell death produced by PCBC. The protective effect of AOAA was independent of any effects on PCBC uptake. These results show that AOAA decreased but did not prevent the metabolism of PCBC by cysteine conjugate β‐lyase. The partial inhibition of PCBC metabolism, and hence, PCBC‐induced cell death by AOAA, may be related to limited concentrations of AOAA within the tubule cell or mitochondria.
Lead influences translational or posttranslational regulation of ia expression and increases invariant chain expression in mouse B cells
Authors: McCabe Michael J. , Dias James A. , Lawrence David A.
Published: October 2006

The molecular mechanisms governing the increased cell surface expression of major histocompatibility complex (MHC) class II molecules (Ia) on lead‐treated mouse B cells was investigated. Lead has been shown to directly cause a selective, two‐fold increase in the B cell's surface density of both products of the I region of the mouse MHC, I‐A and I‐E. In the present study, Western blot analysis showed that Pb increases the total cellular pool of I‐A β‐chain by twofold. The increase in cellular I‐A was not found to be due to increased messenger RNA (mRNA) for either the α‐ or the β‐chain of I‐A. Biosynthetic labeling studies showed that Pb increases the translation or the stability of the Ia‐associated invariant chain (Ii or γ) and possibly the β‐chain of Ia. Collectively these results suggest that Pb increases the B cell's surface Ia by influencing translational or posttranslational regulation of Ia and/or Ia‐associated chains.
Reactivity of Hg(II) with superoxide: Evidence for the catalytic dismutation of superoxide by HG(II)
Authors: Miller Dennis M. , Lund Bert‐Ove , Woods James S.
Published: October 2006

Mercuric ion, a well‐known nephrotoxin, promotes oxidative tissue damage to kidney cells. One principal toxic action of Hg(II) is the disruption of mitochondrial functions, although the exact significance of this effect with regard to Hg(II) toxicity is poorly understood. In studies of the effects of Hg(II) on superoxide (O) and hydrogen peroxide (H2O2) production by rat kidney mitochondria, Hg(II) (1–6 μM), in the presence of antimycin A, caused a concentration‐dependent increase (up to fivefold) in mitochondrial H2O2 production but an apparent decrease in mitochondrial O production. Hg(II) also inhibited O‐dependent cytochrome c reduction (IC50 ≈︁2–3 μM) when O was produced from xanthine oxidase. In contrast, Hg(I) did not react with O in either system, suggesting little involvement of Hg(I) in the apparent dismutation of O by Hg(II). Hg(II) also inhibited the reactions of KO2 (i.e., O) with hemin or horseradish peroxidase dissolved in dimethyl sulfoxide (DMSO). Finally, a combination of Hg(II) and KO2 in DMSO resulted in a stable UV absorbance spectrum [currently assigned Hg(II)‐peroxide] distinct from either Hg(II) or KO2. These results suggest that Hg(II), despite possessing little redox activity, enhances the rate of O dismutation, leading to increased production of H2O2 by renal mitochondria. This property of Hg(II) may contribute to the oxidative tissue‐damaging properties of mercury compounds.
Whether cytochrome p450 induction accompanies glucuronosyl and glutathione transferase induction by isomers of dipyridyl appears unrelated to dose and iron chelation properties
Authors: Franklin Michael R.
Published: October 2006

Administration of 4, 4′dipyridyl to rats induces the activities of xenobiotic transferases (phase II drug metabolizing enzymes), UDP‐glucuronosyl‐tranferase and glutathione‐S‐transferase, and also the concentration and activity of cytochrome P450 (a phase I drug metabolizing enzyme). 2, 2′Dipyridyl, an isomer possessing iron chelation properties, only induces the phase II enzymes. Although the magnitude of the phase II induction by 2, 2′dipyridyl increases with increasing dosages, the selective induction of only phase II activities remains inviolate. Co‐administration of 2, 2′dipyridyl does not prevent 4, 4′dipyridyl from inducing cytochrome P450, suggesting that the iron chelation property is not the factor that precludes 2, 2′dipyridyl from coordinately inducing cytochrome P450 with the transferases.