Commercial Rabbit Research Articles

Ectoadenylate Kinase and Plasma Membrane ATP Synthase Activities of Human Vascular Endothelial Cells

Formation of ATP from ADP on the external surface of vascular endothelial cells has been attributed to plasma membrane ATP synthase, ectoadenylate kinase (ecto-AK), and/or ectonucleoside diphosphokinase. These enzymes or their catalytic products have been causatively linked to the elaboration of vascular networks and the regulation of capillary function. The amount of ATP generated extracellularly is small, requiring sensitive analytical methods for quantification. Human umbilical vein endothelial cells were used to revisit extracellular ATP synthesis using a reliable tetrazolium reduction assay and multiwell plate cultures. Test conditions compatible with AK stability were established. Extracellular AK activity was found to be <1% of the total (intracellular and extracellular), raising the possibility that the external enzyme could have leaked from living cells and/or a few dying cells. To determine whether AK inadvertently leaked from the cells, the activity of another cytoplasmic enzyme, glucose-6-phosphate dehydrogenase (G6PD), was also measured. G6PD is present in the cytoplasm in similar abundance to AK. The activity ratio of G6PD (extracellular/total) was found to be similar to that of AK. Because G6PD in the medium was probably due to leakage, other cytoplasmic macromolecules, including AK, should be released proportionately from the cells. The role of plasma membrane ATP synthase in extracellular ATP formation was examined using Hanks' balanced salt solution with and without selective inhibitors of AK and ATP synthase activities. With P(1),P(5)-di(adenosine 5')-pentaphosphate (inhibitor of AK activity), no extracellular ATP synthesis was detected, whereas with oligomycin, piceatannol, and aurovertin (inhibitors of F(1)F(0)-ATP synthase and F(1)-ATPase activities), no inhibition of extracellular ATP synthesis was observed. AK activity alone could account for the observed extracellular ATP synthesis. The possible impact of ADP impurity in the assays is discussed.

Unravelling the paradox of rabbit haemorrhagic disease virus emergence, using phylogenetic analysis; possible implications for rabbit conservation strategies

During the past 50 years two readily distinguishable rabbit-specific diseases caused by Myxoma virus (MYXV) and Rabbit haemorrhagic disease virus (RHDV) respectively, have decimated wild rabbit populations worldwide. Combined with the use of these viruses as biocontrol agents, the consequences for farming, commercial rabbit breeding and rare habitat conservation dependent on rabbit grazing, have been both positive and negative. Moreover, rare predators that rely on rabbits as a food resource, and even hunters, have suffered the consequences of rabbit populations being affected by one or other of these viruses. Rabbit haemorrhagic disease virus was first identified after thousands of domestic rabbits died suddenly in China in 1984. Similar epidemics subsequently occurred in other regions of Asia, the Middle East, Europe and North America, suggesting that the virus had dispersed widely following its emergence in China. However, the discovery that RHDV had circulated apparently harmlessly for many years before the first recognised epidemic in China prompted us to investigate the evolution, emergence and dispersal of this virus in relation to its impact on conservation of wildlife species. Accordingly, we have sequenced new isolates of RHDV and combined these data with a global selection of available RHDV sequences. Using phylogenetic analysis we demonstrate that the Chinese epidemic virus diverged from European viruses that circulated many years prior to 1984. We also demonstrate that the lineages of the pathogenic viruses that emerged in the UK in the early 1990s, are distinct from and pre-date those of the 1984 Chinese virus. In other words, European strains of RHDV emerged from apparently harmless strains to cause epidemic outbreaks, independently of the Chinese 1984 epidemic virus. These studies demonstrate how understanding viral epidemiology can improve the development of strategies to conserve rabbits, rare predator species and the habitat.

Reuptake of Extracellular Amelogenin by Dental Epithelial Cells Results in Increased Levels of Amelogenin mRNA through Enhanced mRNA Stabilization

Amelogenin is an extracellular matrix protein secreted by ameloblasts and is a major component of enamel matrix. Recently, in addition to their role in enamel formation, the biological activity of enamel proteins in the process of cell differentiation has recently become widely appreciated. In this study, we examined the biological activity of amelogenin on ameloblast differentiation. Recombinant mouse amelogenin (rm-amelogenin) enhanced the expression of endogenous amelogenin mRNA in a cultured dental epithelial cell line (HAT-7), despite a lack of increased amelogenin promoter activity. To solve this discrepancy, we analyzed the effects of rm-amelogenin on the stability of amelogenin mRNA. The half-life of amelogenin mRNA is extremely short, but in the presence of rm-amelogenin its half-life was extended three times longer than the control. Furthermore, we showed the entry of exogenous fluorescein isothiocyanate-conjugated rm-amelogenin into the cytoplasm of HAT-7 cells. It follows from our results that exogenous amelogenin increases amelogenin mRNA levels through stabilization of mRNA in the cytoplasm of HAT-7 cells. Here we speculated that during differentiation, dental epithelial cells utilize a unique mechanism for increasing the production of amelogenin, the reuptake of secreted amelogenin.

PHOX2B Regulates Its Own Expression by a Transcriptional Auto-regulatory Mechanism

The specification of neuronal identity is a result of interactions between the following two distinct classes of determinants: extrinsic factors that include secreted or cell membrane-associated signals in the local environment, and intrinsic factors that generally consist of ordered cascades of transcription factors. Little is known about the molecular mechanisms underlying the interplay between these extrinsic and intrinsic factors and the transcriptional processes that establish and maintain a given neuronal phenotype. Phox2b is a vertebrate homeodomain transcription factor and a well established intrinsic factor in developing autonomic ganglia, where its expression is triggered by the bone morphogenic proteins secreted by the dorsal aorta. In this study we characterized its proximal 5'-regulatory region and found that it contained five putative DNA sites that potentially bind homeodomain proteins, including PHOX2B itself. Chromatin immunoprecipitation assays showed that PHOX2B could bind its own promoter in vivo, and electromobility gel shift assays confirmed that four of the five sites could be involved in PHOX2B binding. Functional experiments demonstrated that 65% of the transcriptional activity of the PHOX2B promoter in neuroblastoma cells depends on this auto-regulatory mechanism and that all four sites were required for full self-transactivation. Our data provide a possible molecular explanation for the maintenance of PHOX2B expression in developing ganglia, in which initially its expression is triggered by bone morphogenic proteins, but may become independent of external stimuli when it reaches a certain nuclear concentration and sustains its own transcription.

Effect of dietary fats on blood cholesterol and lipid and the development of atherosclerosis in rabbits

The objective of this study was to determine the effects of dietary fat on the development of atherosclerosis and changes in blood lipids in rabbits. Forty New Zealand White rabbits were divided into 5 groups and fed commercial rabbit chow with 0% oil (control), 2% corn oil, 2% fish oil, 2% oxidized oil, or 2% heated corn oil. Cholesterol (CHO, 1.5-g/kg diet) was added to all the experimental diets. The rabbits in the control and oxidized oil groups developed the highest scores for atherosclerotic lesions but those fed fish oil, corn oil, and the heated corn oil groups had the lowest lesion scores. Plasma triacylglycerol, total CHO, and low-density lipoprotein and high-density lipoprotein CHO levels ( P < .05) in rabbits followed the trend of higher levels consistent with scores for atherosclerotic lesions. Plasma total CHO level was one factor closely related to the development of atherogenic lesions in the rabbit. Dietary CHO and oxidation status of dietary lipids significantly impacted lipid metabolism and the development of atherosclerosis in rabbits. Oxidized dietary oil accelerated the progress of atherosclerotic lesions, but polymerization of oils by heating lowered plasma CHO level and the development of atherogenic lesions in rabbits. Heating of oil appeared to reduce the absorption of CHO in the rabbit.

The use of a cat-flap at the nest entrance to mimic natural conditions in the breeding of fattening rabbits (Oryctolagus cuniculus)

AbstractManagement systems allowing free nest access are widely used in commercial rabbit breeding, but these produce a potential conflict with the doe's behavioural goal of a closed nest entrance. Furthermore, the restricted space in commercial breeding units prevents the doe from achieving a sufficient distance between her and the nest, another highly adaptive behavioural goal. This can lead to behavioural problems and pup mortality higher than 20%, attributable to hypothermia, injuries, weakness caused by the scattering and crushing of pups, or even cannibalism. In this study we tested a type of nest entrance (a metal cat-flap) that visually closed the nest box while still allowing the doe free access during the first 15 days after parturition. The effects of a cat-flap access (group CF) and of a permanently open nest box (group O) on nest-related behaviour, general activity, and plasma corticosterone concentration of the does, and on the mortality and weight of the pups, were compared using 15 ZIKA does in each group. Over 24 h, there was no difference between the groups in the frequency of ‘nest controls’ (approaches or entries to the nest without nursing of pups). However, outside nursing hours, does in group O showed more ‘head contacts’ with the nest, whereas does in group CF performed more nose contacts and nesting activities outside the nest. Does in group O performed twice as many potentially disturbing nest contacts in this time than does in group CF and had a higher increase of corticosterone after the administration of exogenous adrenocorticotrophic hormone. Pup mortality from days 16 to 35 was significantly higher in group O, and pups born to does in group O left the nest earlier. There was no significant difference in the weaning weights of pups between the two groups. As does with a cat-flap at the nest entrance still showed repeated nest approaches, the cat-flap possibly did not block all nest stimuli from the does. Alternatively, it is possible that the repeated approaches to the next box result from its being one of very few attractive or interesting features in an otherwise barren environment. Removal of the nest box from the cage is an effective method to eliminate nest stimuli, but this increases work for the staff without improving the barren environment for the rabbits. A better way of increasing the distance between the doe and the nest, as well as presenting a number of other attractive features, is group-housing of does.

Limitations of intensive meat rabbit production in North America: A review

This paper documents underlying causes for the poor track record of the commercial meat rabbit industry in North America, relative to the success of several other species (cattle, swine, chickens and turkeys). For over half a century, efforts have been ongoing to develop a viable commercial meat rabbit industry. The progress has not been significant; rather, an accumulation of serious obstacles has targeted the species (e.g., high labor demand, no tradition of rabbit meat consumption, and nutritional limitations and behavioral constraints). Critical biological behaviors associated with the doe rabbit [e.g., short gestation and (or) underdeveloped neonates, cannibalism, territorialism, and pseudo-pregnancy] require that does be permanently placed into individual cages. These behaviors underpin the inability of management to offset labor by employing cost-effective automated feeding and management systems. As a consequence, labor costs per rabbit are high; rabbit meat is generally not competitive with more widely consumed meats. A proposed alternative solution is a redirected focus on rabbits as a “microlivestock” species — reared in small numbers as a family enterprise to enhance quality of life in rural and periurban areas, as well as in lesser developed countries — as opposed to further exploitation of the species as a commercial agricultural commodity. Key words: Rabbit, commercial, contemporary issues, industry, production

Validated Sandwich Enzyme-Linked Immunosorbent Assay for Casein and Its Application to Retail and Milk-Allergic Complaint Foods

Cows' milk is a commonly allergenic food. Cross-contamination of milk proteins into nondairy, kosher-pareve foods prepared on shared processing equipment can cause severe, life-threatening reactions in milk-allergic individuals. A sandwich-type enzyme-linked immunosorbent assay (ELISA; 96-well plate format) was developed for the detection of undeclared casein in foods. Rabbit anti-casein antibodies were used as the capture reagent. Food samples and standards were ground, extracted in 0.01M phosphate-buffered saline, clarified by centrifugation, and added to the wells. Goat anti-casein antibodies were employed as the detector antibody, and the amount of antibody bound was determined with a commercial rabbit anti-goat immunoglobulin conjugated to alkaline phosphatase, with subsequent substrate reaction. Antibodies developed were specific to casein, with no cross-reaction observed with 30 foods and food ingredients. Non–milk-containing products such as fruit juices, fruit juice bars, sorbets, and dark and pareve-labeled chocolate were purchased from June 2002 through June 2003. In addition, samples allegedly causing eight milk-allergic consumer complaints were analyzed. The ELISA had a detection limit of less than 0.5 ppm of casein. The casein content in the analyzed foods ranged from less than 0.5 ppm to more than 40,000 ppm casein; undeclared casein residues were found in all of the samples implicated in allergic reactions. The levels of milk contamination in some of the other surveyed products could also be hazardous for milk-allergic consumers. This ELISA method provides a useful quality control tool for the food industry and could also be used as a validation of kosher-pareve status.

Proteasomal Interference Prevents Zona Pellucida Penetration and Fertilization in Mammals1

The ubiquitin-proteasome pathway has been implicated in the penetration of ascidian vitelline envelope by the fertilizing spermatozoon (Sawada et al., Proc Natl Acad Sci U S A 2002; 99:1223-1228). The present study provides experimental evidence demonstrating proteasome involvement in the penetration of mammalian zona pellucida (ZP). Using porcine in vitro fertilization as a model, penetration of ZP was completely inhibited by specific proteasomal inhibitors MG-132 and lactacystin. Three commercial rabbit sera recognizing 20S proteasomal core subunits beta-1i, beta-2i, alpha-6, and beta-5 completely blocked fertilization at a very low concentration (i.e., diluted 1/2000 to 1/8000 in fertilization medium). Neither proteasome inhibitors nor antibodies had any effects on sperm-ZP binding and acrosome exocytosis in zona-enclosed oocytes or on fertilization rates in zona-free oocytes, which were highly polyspermic. Consistent with a possible role of ubiquitin-proteasome pathway in ZP penetration, ubiquitin and various alpha and beta type proteasomal subunits were detected in boar sperm acrosome by specific antibodies, immunoprecipitated and microsequenced by MALDI-TOF from boar sperm extracts. Antiubiquitin-immunoreactive substrates were detected on the outer face of ZP by epifluorescence microscopy. This study therefore provides strong evidence implicating the ubiquitin-proteasome pathway in mammalian fertilization and zona penetration. This finding opens a new line of acrosome/ZP research because further studies of the sperm acrosomal proteasome can provide new tools for the management of polyspermia during in vitro fertilization and identify new targets for contraceptive development.

Cellulase fermentation on a novel substrate (waste cardboard) and subsequent utilization of home-produced cellulase and commercial amylase in a rabbit feeding trial

Cellulase and amylase enzymes are increasingly being used as feed additives in animal nutrition. Those are mainly applied in order to improve the nutritional value of feed by partially degrading its fiber content, which has major importance during the early post-weaning period of young animals possessing inadequate digestive enzyme activities. Due to their commercial availability at low cost, the utilization of amylases as animal feed additive can be highly recommended, while the high price of cellulases actually makes their use as a feed additive unreasonable. A promising way to gain cheap cellulases is to conduct the enzyme fermentation process on novel substrates, which are readily available at no cost. In this study, the utilization of old corrugated cardboard (OCC) as a novel fermentation substrate has been investigated for cellulase enzyme fermentation by Trichoderma reesei Rut C-30. Batch fermentation experiments were carried out in shake-flasks and in a 30 l laboratory fermenter. The OCC feedstock proved to be a suitable fermentation substrate, a filter paper activity (FPA) of as high as 2.27 FPU ml −1 was achieved in the fermenter, which corresponds to a yield of 227 FPU g −1 cellulose. The maximum productivity achieved was 35.7 FPU l −1 h −1. The subsequent utilization of this cellulase enzyme preparation was investigated as additive of commercial rabbit compound feed in order to improve its digestibility during the early post-weaning period. In parallel, commercial amylase preparations were also tested for the same purposes. The results of animal feeding experiments showed, that enzyme supplementation of the complete feed improved production traits, as it increased the digestibility of nutrients and thus reduced the mortality caused by digestive disorders.

COMPARATIVE STUDY ON THE RABBIT FISHES SIGANUS CANALICULATUS INHABIT THE ARABIAN GULF AND SIGANUS RIVULA TUS INHABIT THE RED SEA IN SAUDI ARABIA

The present work was carried out to compare some biological aspects, chemical composition, fatty acids, minerals and heavy metals of the commercial Rabbit fishes Siganus canaliculatus inhabit the Arabian Gulf and Siganus rivulatus inhabit the Red Sea in Saudi Arabia. Fish samples were selected to represent all fish size categories in the catch and were collected during the period from November 2002 to March 2003. Each fish specimen was measured, weighted and recorded, then dissected to investigate the fullness of alimentary canal and the maturity stage of the gonads. The length-weight relationships were computed by simple regression analysis and graphically represented. Both of absolute and relative condition factors (Kc and Kn) were estimated and graphically represented. The muscles (flesh) were removed and weighted to estimate the flesh percentage. Samples of flesh were taken to determine moisture, protein, ether extract, ash, fatty acids, minerals and heavy metals. Some samples of the entire fish were used to proximate analysis. The present results indicated that the Kc, Kn, maximum total length and flesh percentage of Rabbit fish Siganus canaliculatus were'higher than that of Siganus rivulatus. In general, polyunsaturated fatty acids were relatively low and heavy metal concentrations in the flesh of the two Siganus species-were lower than the permit limits for Human consumption,

Vascular Smooth Muscle α-Actin Gene Transcription during Myofibroblast Differentiation Requires Sp1/3 Protein Binding Proximal to the MCAT Enhancer

The conversion of stromal fibroblasts into contractile myofibroblasts is an essential feature of the wound-healing response that is mediated by transforming growth factor beta1 (TGF-beta1) and accompanied by transient activation of the vascular smooth muscle alpha-actin (SmalphaA) gene. Multiple positive-regulatory elements were identified as essential mediators of basal SmalphaA enhancer activity in mouse AKR-2B stromal fibroblasts. Three of these elements bind transcriptional activating proteins of known identity in fibroblasts. A fourth site, shown previously to be susceptible to single-strand modifying agents in myofibroblasts, was additionally required for enhancer response to TGF-beta1. However, TGF-beta1 activation was not accompanied by a stoichiometric increase in protein binding to any known positive element in the SmalphaA enhancer. By using oligonucleotide affinity isolation, DNA-binding site competition, gel mobility shift assays, and protein overexpression in SL2 and COS7 cells, we demonstrate that the transcription factors Sp1 and Sp3 can stimulate SmalphaA enhancer activity. One of the sites that bind Sp1/3 corresponds to the region of the SmalphaA enhancer required for TGF-beta1 amplification. Additionally, the TGF-beta1 receptor-regulated Smad proteins, in particular Smad3, are rate-limiting for SmalphaA enhancer activation. Whereas Smad proteins collaborate with Sp1 in activating several stromal cell-associated promoters, they appear to operate independently from the Sp1/3 proteins in activating the SmalphaA enhancer. The identification of Sp and Smad proteins as essential, independent activators of the SmalphaA enhancer provides new insight into the poorly understood process of myofibroblast differentiation.

Estrogenic isoflavones in rodent diets

Many rodent diets contain components such as soy isoflavones (daidzein and genistein) known to have estrogenic properties. The dietary background of phytoestrogens may modulate some responses to environmental estrogens when these compounds are tested in rodent bioassays. Thus, and since only few data were available on the phytoestrogen content of rodent diets commonly used in European laboratories, it was of interest to analyze the daidzein and genistein contents of our standard animal feeds. Isoflavone contents were determined in seven batches of rodent chow (from two suppliers in Germany, Altromin and Ssniff) by high-performance liquid chromatography, and also analyzed in six rodent diets from the United States. The soy-based rodent diets from Germany contained isoflavone (daidzein plus genistein) concentrations in the range of 0.3–0.55 mg/g feed. These isoflavone contents are similar to those analyzed in the US rodent diets, and similar to values reported by others, including one particular lot of feed (with 0.35 mg isoflavones per g) which produced a large uterotrophic response in immature ovariectomized rats [Environ. Health Perspect., 106 (1998) 369]. Coumestrol was found in a sample of commercial rabbit food at rather high levels (0.27 mg/g), but, this phytoestrogen was not detected (<1 μg/g feed) in any of the other samples we analyzed. The soy components in our rodent diet produce a measurable background of daidzein and genistein in blood of female DA/Han rats, total isoflavones (aglycone plus conjugates) ranging between 90 and 290 ng/ml plasma. The ovariectomized animals kept on this chow, showed no signs of estrogenization of the reproductive tract (uterus, vagina), and responded normally to (xeno-)estrogen administration in a uterotrophic assay [J. Steroid Biochem. Mol. Biol., 73 (2000) 1]. Moreover, ovariectomized Wistar rats on our standard rodent diet (Ssniff R/M H) had lower uterine weights than animals kept on the isoflavone-free (solvent extracted) chow; both groups of rats responded to genistein administration with an increase in uterine weights. These results suggest that—albeit the sensitivity of the rodent uterotrophic assay is not reduced by the use of a diet containing soy isoflavones at commonly encountered levels—attention should be given to a variable dietary phytoestrogen background.

Translational extracts active biologically in vitro obtained from eukaryotic monolayer cells: a versatile method for viral RNA studies

Preservation of enzymatic activities in biological samples, especially after freeze/thawing, is a crucial requirement in virological research. Theoretically, this preservation can be achieved with the presence of cryopreservative agents. In contrast to tedious methods, it was found that this might be readily achieved by using well-defined conditions, including sucrose in the samples. Hence, the generation of a translational extract obtained from eukaryotic cells that have grown as monolayers is described below. This versatile method could be used advantageously for the in vitro translation of messenger RNAs, added exogenously, including viral mRNAs. The translational extract can be prepared freshly on a daily basis, or more conveniently it can be frozen and thawed subsequently for further use, without loss of activity. It can replace the Krebs ascites fluid and the commercial rabbit reticulocyte lysate. The procedure employed for the preservation of the biological activity of the translational extract can be extended to various other biological samples.

Benefits of Eating Conspecifics: Effects of Background Diet on Survival and Metamorphosis in the Cuban Treefrog (Osteopilus septentrionalis)

Abstract Cannibalism provides a potential mechanism for enhanced growth and survival for species with complex lifecycles developing in habitats of unpredictable duration. We asked whether background resource quality affected survival and metamorphosis of Cuban Treefrog (Osteopilus septentrionalis) tadpoles fed conspecifics. In a factorial experiment, we crossed two levels of background resources, a high-quality (commercial rabbit chow) or low-quality (algae and periphyton) diet with no conspecifics, the presence of a dead conspecific (in a bag), or ad libitum availability of dead conspecifics. Survival of tadpoles did not differ significantly among treatments. Availability of conspecifics resulted in larger mass and decreased larval periods for tadpoles on low-quality background treatments; however, for tadpoles on high-quality background treatments, availability of conspecifics resulted in prolonged larval periods. These results suggest that profitability of feeding on conspecifics is dependent on backgr...

Preference Test on Feed and Nutrient Intakes in Male and Female Lesser Mouse Deer (Tragulus Javanicus) in Captivity

A preference test on feed and nutrient intakes were conducted on four male (1.25±0.08 kg) and four female (1.21±0.15 kg) lesser mouse deer (Tragulus javanicus) in captivity. Each animal was kept in individual cages placed in a well-ventilated animal house. The experiment was conducted in two weeks, where the first week was for adaptation to the feeds and the second week for measurements of nutrient intake, nutrient digestibility and nitrogen balance. The feeds offered were kangkong (Ipomoea aquatica), long bean (Vigna sinensis) and french bean (Phaseolus vulgaris) as roughages and proteinaceous feeds; sweet potato (Ipomoea batatas) and carrot (Daucus carota) as carbohydrate-rich feeds; and commercial rabbit pellet (0.3 cm diameter and 0.5 cm long) as a complete feed. The dry matter (DM) content of each feed in the order mentioned above was 7.1, 6.1, 3.9, 18.5, 6.2 and 87.6%, respectively. Long bean had the highest protein (CP) content (29.7%), while sweet potato had the lowest (6.2%). The CP contents of other feeds were within the range of 14.2 -25.1%. Among the feeds, carrot had the lowest energy content (3.83 kcal/g) and long bean the highest (4.67 kcal/g). When fresh weight of the feed was considered, the male mouse deer consumed sweet potato the most (86.3±12.90 g/d), but the female had a high preference for carrot (79.2±9.76 g/d). The other feeds were consumed in lesser amounts. However, in terms of DM of the feed, the amount of commercial pellet consumed was the highest for both male (45.0±5.10%) and female (44.7±7.38%) mouse deer, followed by sweet potato (33.1±4.43% and 22.4±7.73% for male and female, respectively), Significant (p<0.05) differences in DM, organic matter (OM) and gross energy (GE) intakes were observed between male and female mouse deer. The male consumed higher amount of DM, OM and GE than the female. The total DM intake was 40.7±2.24 g/d/kg W0.75 for male and 35.9±1.72 g/d/kg W0.75 for female mouse deer. Percentage digestibilities of DM, OM, CP and GE were within 72.7-80.8% and were not significantly different between male and female mouse deer. However, male mouse deer had significantly (p<0.05) higher digestible DM, OM and GE intakes than the female. Both male and female mouse deer were in positive nitrogen balance (0.6 g N/d/kg W0.75). The male mouse deer gained 7.6±3.45 g/d, while the female gained 4.3±2.40 g/d.

Fructose 1,6-Bisphosphate Aldolase from Staphylococcus carnosus: Overexpression, Structure Prediction, Stereoselectivity, and Application in the Synthesis of Bicyclic Sugars

The unusually stable fructose 1,6-bisphosphate aldolase from Staphylococcus carnosus (FruAsca) was overproduced in E. coli. An α=β-barrel structure and highly conserved active site was inferred from sequence analysis in comparison to known class I aldolases. A preparative study with generic aliphatic and hydroxylated aldehydes confirmed a high level of stereoselectivity, and thus a functional equivalence of the Staphylococcus enzyme to the commercial rabbit muscle aldolase for asymmetric synthesis. Despite their high enzyme cross-linking reactivity, glutardialdehyde derivatives could be converted by the Staphylococcus aldolase to give novel bicyclic sugars.

Fructose 1,6-Bisphosphate Aldolase fromStaphylococcus carnosus: Overexpression, Structure Prediction, Stereoselectivity, and Application in the Synthesis of Bicyclic Sugars

The unusually stable fructose 1,6-bisphosphate aldolase from Staphylococcus carnosus (FruAsca) was overproduced in E. coli. An α=β-barrel structure and highly conserved active site was inferred from sequence analysis in comparison to known class I aldolases. A preparative study with generic aliphatic and hydroxylated aldehydes confirmed a high level of stereoselectivity, and thus a functional equivalence of the Staphylococcus enzyme to the commercial rabbit muscle aldolase for asymmetric synthesis. Despite their high enzyme cross-linking reactivity, glutardialdehyde derivatives could be converted by the Staphylococcus aldolase to give novel bicyclic sugars.

Rectal temperature and prostaglandin E2 increase in cerebrospinal fluid of conscious rabbits after intracerebroventricular injection of hemoglobin.

Fever accompanies subarachnoid hemorrhage (SAH) in the majority of patients. In a previous study, hemoglobin (Hb) was shown to catalyze in vitro, under aerobic conditions, the conversion of arachidonic acid to prostaglandin E2 (PGE2) and prostaglandin F2alpha. The aim of the present work was to assess whether this pathway also operates in vivo and to provide a mechanism to explain post-SAH fever. To this end, PGE2 concentration was determined in cerebrospinal fluid (CSF) of conscious rabbits chronically cannulated in the lateral ventricle and cisterna magna, following intracerebroventricular (i.c.v.) injection of 10 microg or 100 microg of commercial rabbit bicrystallized Hb as a model of SAH. Before i.c.v. injection, Hb solutions were filtered on a polimixin-B column to remove substantially, by over 90%, endotoxin-like substances. Results show that in nine rabbits injection of 10 microg Hb did not significantly modify body temperature or significantly alter CSF PGE2 content. On the contrary, in nine rabbits, injection of 100 microg Hb produced a significant increase in core temperature which was accompanied by a significant increase in CSF PGE2. When data related to these two parameters from the 9 control and 18 Hb-treated rabbits were analyzed as a single group, a linear, positive, and highly significant correlation was found. These findings indicate that, once Hb is released into the subarachnoid space during SAH, it enhances CSF PGE2 content and elicits hyperthermia, thus offering an explanation for the fever that is an aggravating condition in most SAH patients.

Cadmium-bound species in human urine using high-performance liquid chromatography-vesicular hydride generation-inductively coupled plasma mass spectrometry

Anion-exchange liquid chromatography was coupled on-line with inductively coupled plasma mass spectrometry (ICP-MS) via a vesicular hydride generation (HG) interface. This hybrid arrangement was assessed for Cd speciation in standards of rabbit liver metallothionein (MT) and human urine. Commercial rabbit liver MTs were readily resolved using a concentration gradient (2-200 mM) of Tris-HCl buffer solution at pH 7.4 and a polymeric Protein-Pak DEAE-5PW anion-exchange column. Three Cd species were detected in rabbit liver MT 1 [retention times of 3.2 (I), 5.6 (II) and 9.3 (III) min] and two species in MT 2 (retention times of 16.7 and 24 min). In rabbit liver MT 1 and MT 2, mixtures of Cd and Zn were found to be evenly distributed between the two isoforms, whereas Cu was predominantly associated with the MT 1 isoform.The chromatographic behaviour of the Cd-containing species separated and detected in basal human urine seems to be similar to that of the Cd fractions I and II of rabbit liver MT 1. The efficiency of CdH 2 generation for Cd bound to both the main isoforms of rabbit liver was the same as for inorganic Cd 2+ from a vesicular medium. This highly selective and sensitive on-line HG-ICP-MS detection system provided a 4-fold (aqueous) or 16-fold (urine) sensitivity enhancement in comparison with ICP-MS detection with conventional nebulization. Unfortunately, only a 2-5-fold improvement was observed for the corresponding detection limits due to the high values of the blanks observed using volatile species generation.