bluuebunny/crossref_metadata_2025 huggingface dataset:
Growth and metabolism of fetal and maternal muscles of adolescent sheep on adequate or high feed intakes: possible role of protein kinase C-α in fetal muscle growth
Authors: Palmer Robert M. , Thompson Michael G. , Meallet Chrystel , Thom Amanda , Aitken Raymond P. , Wallace Jacqueline M.
Published: September 2005

From days 4–104 of pregnancy, adolescent sheep, weighing 43·7 (SE 0·87)kg were offered a complete diet at two different intakes (approximately 5 or 15 kg/week) designed to meet slightly, or well above, maternal maintenance requirements. The fetal and maternal muscles were taken on day 104 of pregnancy and analysed for total DNA, RNA and protein. Ewes offered a high intake to promote rapid maternal weight gain, weighed more (76·5 (SE 4·5)v.50·0 (SE 1·7) kg) and had muscles with a greater fresh weight, whilst their fetuses had smaller muscles, than those fed at a lower intake. Plantaris muscle of the ewes fed at the high intake contained more RNA and protein; again the opposite situation was found in the fetal muscle. On the higher maternal intakes, the DNA, RNA and protein contents of the fetal plantaris muscle were less than in fetuses of ewes fed at the lower intake. To investigate the possible mechanisms involved in this decrease in fetal muscle mass, cytosolic and membrane-associated muscle proteins were subjected to Western immunoblotting with antibodies to nine isoforms of protein kinase C (PKC), a family of enzymes known to play an important role in cell growth. Five PKC isoforms (α, ε, θ, μ and ζ) were identified in fetal muscle. One of these, PKC-α, was located predominantly in the cytosolic compartment in the smaller fetuses of the ewes fed at a high plane of nutrition, but was present to a greater extent in the membranes of the more rapidly growing fetuses of the ewes fed at the lower intake. This was the only isoform to demonstrate nutritionally related changes in its subcellular compartmentation suggesting that it may mediate some aspects of the change in fetal growth rate.
Glutamine metabolism in the gastrointestinal tract of the rat assessed by the relative activities of glutaminase (EC 3.5.1.2) and glutamine synthetase (EC 6.3.1.2)
Authors: James L. A. , Lunn P. G. , Elia M.
Published: September 2005

The activities of the two key enzymes involved in glutamine metabolism, glutaminase (EC 3.5.1.2) and glutamine synthetase (EC 6.3.1.2), have been measured in the various tissues of the gastrointestinal (GI) tract of the rat, from the mouth to the rectum. Glutaminase activity was particularly high in the mucosa of the small intestine, where its activity accounted for more than 80% of the total activity of the GI tract. In contrast, the mouth and oesophagus had very low activities, accounting for less than 2% of the total. Glutamine synthetase was mainly confined to the lower part of the stomach, which accounted for almost 90% of the total activity of the GI tract. Activity in the small intestine was very low, accounting for less than 2% of the total, and similarly low levels were found in the mouth and oesophagus. The data provide the most complete information on the distribution of these enzymes in the GI tract of the rat and suggest: (a) that the mucosa of the small intestine has the highest capacity for glutamine breakdown but the lowest capacity for its synthesis, and so requires an external source of this amino acid; (b) that there is little potential for glutamine synthesis or breakdown in the mouth and oesophagus; and (c) that the lower stomach has a substantial capacity to synthesize glutamine, in contrast to the rest of the GI tract. The results of the investigation are relevant to sites of glutamine metabolism in therapeutic studies involving glutamine administration discussed with reference to reports of the effects of glutamine administration on GI tract injury.
Urinary excretion of purine derivatives as an index of microbial-nitrogen intake in growing rabbits
Authors: Balcells J. , Ganuza J. M. , Pérez J. F. , Martín-Orúe S. M. , González Ronquillo M.
Published: September 2005

Three experiments were carried out to establish a response model between intake and urinary excretion of purine compounds. In Expt 1 the relationship between the intake of purine bases (PB) and the excretion of total purine derivatives (PD) was determined in seven growing rabbits with a mean initial live weight (LW) of 2·03 (SE 0·185) kg, aged 70 d, each fitted with a wooden neck collar to prevent caecotrophagy. They were fed on five experimental diets formulated with different levels of nucleic acids (0·00, 3·75, 7·50, 11·25, 15·00 g yeast-RNA/kg diet). The relationship between intake of purine (x,μmol/kg W0·75) and total urinary PD excretion (y,μmol/kg W0·75),y= 0·56 + 0·67x(r20·86; RSD 0·338), indicated that about 70% of duodenal PB were recovered as urinary PD and that the endogenous contribution was constant and independent of dietary PB supply. Endogenous excretion of PD (allantoin and uric acid) was measured in a second experiment using six rabbits fed on a purine-free diet and fitted with neck collars to avoid caecotrophagy. Basal daily urinary excretion values for allantoin and uric acid were 532 (SE 33·9) and 55 (SE 7·3) μmol/kg W0·75respectively; xanthine and hypoxanthine were not found in urine samples and therefore the sum of allantoin and uric acid should comprise the total excretion of PD (588 (SE 40·1) μmol/kg W0·75). The xanthine oxidase (EC 1·2.3·2) activity in plasma, liver, duodenum, jejunum and kidney was measured in a third experiment. The activities of xanthine oxidase in duodenal and jejunal mucosa, liver and kidney were: 0·61 (SE 0·095), 0·37 (SE 0·045), 0·035 (SE 0·001) and 0 units/g fresh tissue respectively and in plasma 2·96 (SE 0·094) units/1. The results show that urinary excretion of PD may be a useful tool to estimate duodenal PB input and microbial protein intake once the relationship between PB and N has been established in caecal micro-organisms.
Metabolism of parenterally administered fat emulsions in the rat: studies of fatty acid oxidation with 1-13C- and 8-13C-labelled triolein
Authors: Bäurle Wolfgang , Brösicke Herbert , Matthews Dwight E. , Pogan Karin , Fürst Peter
Published: September 2005

To reassess the hypothesis that fatty acid catabolism occurs to completion via β-oxidation, male Sprague–Dawley rats receiving continuous total parenteral nutrition (TPN) including 43% energy as fat were infused with [1-13C]- or [8-13C]triolein. Expired CO2was collected continuously for 4 h and its13C:12C ratio determined by isotope–ratio mass spectrometry. Bicarbonate retention was also assessed over 4 h by infusion of NaH14CO3and measurement of the expired14CO2. A possible loss of label from [8-13C]oleic acid from the citric acid cycle via labelled acetyl-CoA without oxidation to CO2was assessed by infusing further animals with acetate labelled with14C either at C atoms 1 or 2 and determination of its conversion to expired14CO2. At isotopic steady state, 63.2 (SE 1.6)% (n8) of the infused [1-14C]acetate and 46.0 (SE 1.2)% (n8) of [2-14C]acetate was recovered as expired14CO2. After correction for bicarbonate retention and non-oxidative isotope loss, 37.3 (SE 1.2)% (n20) of the [1-13C]triolein was found to have been oxidized, whereas 32.6 (SE 1.0)% (n20) of the [8-13C]triolein was oxidized (P≤ 0.01). The lower oxidation of the C atom at position 8 of oleic acid than that at position 1 indicates incomplete oxidative breakdown of the fatty acid after entering β-oxidation.
Phyto-oestrogens: where are we now?
Authors: Bingham S. A. , Atkinson C. , Liggins J. , Bluck L. , Coward A.
Published: September 2005

Phyto-oestrogens have emerged from their esoteric role in animal husbandry following the hypothesis that the human Western diet is relatively deficient in these substances compared with societies where large amounts of plant foods and legumes are eaten. Evidence is beginning to accrue that they may begin to offer protection against a wide range of human conditions, including breast, bowel, prostate and other cancers, cardiovascular disease, brain function, alcohol abuse, osteoporosis and menopausal symptoms. Of the two main classes of these weak oestrogens, the isoflavones are under intensive investigation due to their high levels in soyabean. Like the ‘anti-oestrogen’ Tamoxifen, these seem to have oestrogenic effects in human subjects in the cardiovascular system and bone. Although previously only available from food, isoflavones are now being marketed in health-food supplements or drinks, and tablets may soon be available over the counter as ‘natural’ hormone-replacement therapy. In cancer, anti-oestrogenic effects are thought to be important, although genistein especially has been shown to induce wide-ranging anti-cancer effects in cell lines independent of any hormone-related influence. There are few indications of harmful effects at present, although possible proliferative effects have been reported. In infants, the effects of high levels in soya milk formulas are uncertain. The second group, lignans, have been less investigated despite their known anti-oestrogenic effects and more widespread occurrence in foods. Investigation of the possible benefits ofphyto-oestrogensis hampered by lack of analytical standards and, hence, inadequate methods for the measurement of low levels in most foods. This problem may prove to be a major dilemma for regulatory authorities, clinicians and others wishing to advise the general public on whether these compounds really do have the health benefits attributed to them.
The effect of test meal monounsaturated fatty acid: saturated fatty acid ratio on postprandial lipid metabolism
Authors: Roche Helen M. , Zampelas Antonis , Jackson Kim G. , Williams Christine M. , Gibney Michael J.
Published: September 2005

Epidemiological evidence shows that a diet high in monounsaturated fatty acids (MUFA) but low in saturated fatty acids (SFA) is associated with reduced risk of CHD. The hypocholesterolaemic effect of MUFA is known but there has been little research on the effect of test meal MUFA and SFA composition on postprandial lipid metabolism. The present study investigated the effect of meals containing different proportions of MUFA and SFA on postprandial triacylglycerol and non-esterified fatty acid (NEFA) metabolism. Thirty healthy male volunteers consumed three meals containing equal amounts of fat (40g), but different proportions of MUFA (12, 17 and 24% energy) in random order. Postprandial plasma triacylglycerol, apolipoprotein B-48, cholesterol, HDL-cholesterol, glucose and insulin concentrations and lipoprotein lipase (EC3.1.1.34) activity were not significantly different following the three meals which varied in their levels of SFA and MUFA. There was a significant difference in the postprandial NEFA response between meals. The incremental area under the curve of postprandial plasma NEFA concentrations was significantly (P= 0·03) lower following the high-MUFA meal. Regression analysis showed that the non-significant difference in fasting NEFA concentrations was the most important factor determining difference between meals, and that the test meal MUFA content had only a minor effect. In conclusion, varying the levels of MUFA and SFA in test meals has little or no effect on postprandial lipid metabolism.
Rapid chylomicron appearance following sequential meals: effects of second meal composition
Authors: Evans Kevin , Kuusela Pertti J. , Cruz Martha L. , Wilhelmova Ingrid , Fielding Barbara A. , Frayn Keith N.
Published: September 2005

Previous studies have noted the presence of an early postprandial peak in plasma triacylglycerol concentrations following successive fat-rich meals. An earlier study has shown that the triacylglycerol in this early peak originates from a previous meal. The present study was performed to investigate the effects of different second meals on the plasma triacylglycerol response. Six healthy subjects were studied on four occasions each. At 5 h following a fat-rich breakfast they ingested one of the following in a balanced design: a fat-rich meal, a low-fat meal, water or nothing by mouth. Blood samples were taken for 2.5 h following the second meal. An early peak in chylomicron and plasma triacylglycerol concentrations was seen following both low-fat and fat-rich second meals but not following water. During studies investigating postprandial lipaemia, further meals must be avoided, even if they contain no fat, although water may be allowed.
Drug–nutrient interactions: inhibition of amino acid intestinal absorption by fluoxetine
Authors: Urdaneta Elena , Idoate Isabel , Larralde Jesús
Published: September 2005

Fluoxetine is one of the most widely used antidepressants and nowadays it is also being used to manage obesity problems. In our laboratory we demonstrated that the drug inhibited sugar absorption (Monteiroet al.1993). The aim of the present work was to determine the effect of fluoxetine on intestinal leucine absorption. Using a procedure of successive absorptionsin vivothe drug diminished amino acid absorption by 30% (P< 0.001). Experimentsin vitroin isolated jejunum also revealed a reduction in leucine uptake of 37% (P< 0.001). In both cases fluoxetine only affected mediated transport without altering diffusion. In a preparation enriched in basolateral membrane, fluoxetine inhibited the Na+,K+-ATPase (EC3.6.1.37) activity (55%;P< 0.001) in a non-competitive manner with an inhibition constant (Ki) value of 0.92 mm. Leucine uptake by brush-border membrane vesicles was diminished by the drug (a reduction of 48% was observed at 30s,P< 0.001); only the apical Na+-dependent transport system of the amino acid was modified and the inhibition was non-competitive. Leucine uptake in the presence of lysine indicated that transporter B was involved. These results suggest that fluoxetine reduces leucine absorption by its action on the basolateral and apical membrane of the enterocyte; the nutritional status of the patients under drug treatment may be affected as neutral amino acid absorption is decreased.
Effect of protein restriction on messenger RNA of insulin-like growth factor-I and insulin-like growth factor-binding proteins in liver of ovariectomized rats
Authors: Higashi Yusuke , Takenaka Asako , Takahashi Shin-Ichiro , Noguchi Tadashi
Published: September 2005

Effects of dietary protein restriction and ovariectomy on plasma concentrations and hepatic messenger RNA (mRNA) of insulin-like growth factor-I (IGF-I) and insulin-like growth factor binding proteins (IGFBP) were investigated in young female rats. Ovariectomy increased plasma IGF-I concentration in rats fed on either a 50 g casein/kg diet (protein-restricted diet) or a 200 g casein/kg diet (control diet), but it increased IGF-I mRNA in liver only in the rats fed on the control diet. On the other hand, by Western ligand blot analysis, we observed that ovariectomy increased plasma IGFBP-3 concentration, and decreased plasma IGFBP-4 concentration. Ovariectomy did not affect IGFBP-1 and IGFBP-2 mRNA in liver, but dietary protein restriction significantly increased them, which may correspond to their plasma concentrations. The present results show that ovarian hormones and dietary protein content affect the plasma concentrations of IGF-I, IGFBP-3 and IGFBP-4 and hepatic mRNA of IGF-I, IGFBP-1, IGFBP-2, IGFBP-3 and IGFBP-4 in different manners.
Lowering of plasma triiodothyronine level and sympathetic activity does not alter hypoalbuminaemia in rats fed on a diet of low protein concentration
Authors: Sawaya Ana Lydia , Lunn Peter G.
Published: September 2005

Previous studies have described high plasma triiodothyronine (T3) concentrations and sympathetic activity in rats fed on low-protein diets. The present investigation examined how the nutritional status of rats fed on a low-protein diet was affected when these hormonal changes were reduced by drug administration. The low-protein diet (LP group) prevented growth, reduced plasma albumin levels, elevated plasma T3concentration, and increased both the weight of the interscapular brown adipose tissue (BAT) and the activity of BAT cytochrome c oxidase (EC1.9.3.1). Lowering the plasma T3concentration (with carbimazole; CA group) elevated the plasma insulin concentration, promoted a small increase in the plasma albumin concentration and caused weight gain in comparison with the LP group. Reduction of sympathetic activity (with α-methyl-p-tyrosine; MT group) promoted a small elevation in plasma albumin concentration accompanied by a diminished T3concentration, BAT weight, and an increase in fat deposition in relation to LP rats. In a second experiment, simultaneous lowering of the plasma T3concentration and sympathetic activity (CA/MT group) resulted in weight gain associated with elevated plasma insulin concentration and fat deposition and a marked reduction in BAT cytochrome c oxidase activity. However no change in the hypoalbuminaemia was observed. The results of the present study suggest that in spite of the previously described increase in metabolic rate in rats fed on a diet with low-protein concentration when compared with controls, the mechanisms involved in the control of BAT activity and fat deposition seem to be independent of those which cause liver protein depletion and hypoalbuminaemia.
Effect of different varieties of pectin and guar gum on plasma, hepatic and biliary lipids and cholesterol gallstone formation in hamsters fed on high-cholesterol diets
Authors: Trautwein Elke A. , Kunath-Rau Angelika , Erbersdobler Helmut F.
Published: September 2005

The effect of high- (hePE) and low- (lePE) esterification pectin and high- (hvGG) and low-(lvGG) viscosity guar gum on plasma, hepatic and biliary lipids and on prevention of cholesterol gallstones was investigated in male golden Syrian hamsters (Mesocricetus auratus). Hamsters were fed on cholesterol-rich (4g/kg), gallstone-inducing diets for 6 weeks. The diets were supplemented with 80g hePE, lePE, hvGG or lvGG/kg or 80g additional cellulose/kg. No significant differences in plasma total cholesterol and triacylglycerol concentrations between hvGG and lvGG and the gallstone-inducing or cellulose-enriched diets were observed. The hePE diet produced a 16% (non-significant) reduction in total plasma cholesterol but significantly decreased the plasma triacylglycerol level by 45%. The lePE diet caused only minor changes in plasma lipids. Hepatic cholesterol concentrations were significantly higher in hamsters fed on hvGG, lvGG, hePE or lePE primarily due to the accumulation of esterified cholesterol. Supersaturated bile samples, with lithogenic indices ranging from 1.6 to 2.0, were determined with all diets. The hePE and lePE diets slightly altered the bile acid profile by increasing glycocholic acid and decreasing taurochenodeoxycholic acid concentrations resulting in a higher cholic: chenodeoxycholic acid ratio. Cholesterol gallstone formation was not substantially inhibited by the two varieties of pectin and guar gum. The hvGG, lvGG, hePE and lePE diets did not alter faecal weight and caused only minor increases in faecal bile acid excretion. In general, the present findings demonstrate that dietary pectins and guar gums had only minor effects on cholesterol metabolism and did not prevent cholesterol gallstone formation in this hamster model. Possible explanations for this lack of a distinct response to pectin and guar gum are discussed.
Are overweight women at increased risk of obesity following pregnancy?
Authors: Harris H. E. , Ellison G. T. H. , Richter L. M. , Wet T. De , Levin J.
Published: September 2005

Longitudinal studies suggest that women who already have a high BMI are at greater risk of maternal obesity than their lighter counterparts. The aim of the present study was to investigate this possibility by examining the relationship between reproductive history and maternal BMI in a community of 627 women from South Africa with a high prevalence of obesity. Standardized questionnaires were used to obtain detailed sociodemographic and behavioural information, while maternal weight and height were both measured at the time of the interview. Analysis of covariance (ANCOVA) showed that maternal age (r2 0·015, P = 0·001), smoking status (r2 0·012, P = 0·036), and social support (r2 0·011, P = 0·006) were all independently associated with maternal BMI. If overweight women were at increased risk of maternal obesity, then the positive relationship between reproductive history and maternal BMI should be enhanced in this relatively obese community, yet the ANCOVA models showed no independent association between gravidity and maternal BMI after controlling for the effects of confounding factors. Although previous longitudinal studies have found a positive association between prepregnant weight and long-term weight gain, this relationship might arise because overweight women gain more weight over a fixed period of time than normal weight women, and therefore they may appear to be at greater risk of pregnancy-related weight gains. Overweight women are at greater risk of weight gain generally, but there is little unequivocal evidence to suggest that they are at any increased risk of maternal obesity, when compared with women of lower BMI.
Faecal pH, bile acid and sterol concentrations in premenopausal Indian and white vegetarians compared with white omnivores
Authors: Reddy Sheela , Sanders T. A. B. , Owen R. W. , Thompson M. H.
Published: September 2005

Faecal bulk, pH, water content, the concentrations of neutral sterols and bile acids and dietary intakes were measured in twenty-two Indian vegetarian, twenty-two white omnivorous and eighteen white vegetarian premenopausal women. Faecal bulk and water content were greater and pH lower in the Indian vegetarians. Total faecal animal sterol and coprostanol concentrations expressed on a dry-weight basis were lower in the vegetarians compared with the omnivores. The faecal sterol concentrations were correlated with dietary cholesterol intake. Primary bile acids were detected in six Indian vegetarians, two white vegetarians and two white omnivores; secondary bile acids were detected in all the white omnivore and vegetarian subjects but not in two of the Indian vegetarians. Total faecal free bile acid and conjugated bile acid concentrations were lower in the white vegetarians compared with the omnivores. Faecal lithocholic acid concentrations were lower in both Indian and white vegetarians. The lithocholic: deoxycholic acid ratio and coprostanol: total animal sterols ratio were significantly lower in the Indian vegetarians compared with the omnivores. Both ratios were positively correlated with faecal pH. Stepwise multiple regression analyses were undertaken in order to identify which nutrients influenced faecal pH, lithocholic and deoxycholic acid concentrations. The intakes of starch and dietary fibre were negatively associated with faecal concentrations of lithocholic and deoxycholic acid. Starch intake alone was negatively associated with faecal pH. The results of this study confirm that diets high in dietary fibre decrease faecal bile acid concentrations and suggest that the complex carbohydrates present in Indian vegetarian diets influence faecal pH and inhibit the degradation of faecal steroids.
The use of cumulative gas and volatile fatty acid production to predict in vitro fermentation kinetics of Italian ryegrass leaf cell walls and contents at various time intervals
Authors: Groot Jeroen C. J. , Williams Barbara A. , Oostdam Arno J. , Boer Huug , Tamminga Seerp
Published: September 2005

Differences between the fermentation characteristics of cell contents (CC) and protease-treated cell walls (CW) of young leaves of Italian ryegrass (Lolium multiflorumLam.) cultivar Multimo (tetraploid), were studiedin vitro.Gas and volatile fatty acid (VFA) production rates were measured at regular intervals, as was the degradability of organic matter (OM) of CW. The measured VFA were used to predict the gas production and fermentable OM using stoichiometric calculations. For CW the volume and kinetics of measured gas production were the same as those predicted from the VFA formed. In contrast, the measured gas production for CC was consistently less than predicted, indicating that the stoichiometric equations were not valid for rapidly fermenting substrates. For both CC and CW, the relative rate of acetic acid production levelled off more slowly than for other VFA, resulting in an increasing gas yield (in ml/g fermentable OM) after 12 (CW)-24 (CC)h incubation. Consequently, the fermentation of OM was not linearly related to gas production kinetics. For CW, the kinetics of decline of degradable OM and fermentable OM were the same, after correction for a constant ‘lost fraction’ of degradable OM of 205 g/kg OM. This work indicates the value of detailed studies of fermentation processes to evaluate herbage quality. In particular, the role of CC and the difference between degradation and fermentation require further attention.
Rat adipose tissue rapidly accumulates and slowly releases an orally-administered high vitamin D dose
Authors: Brouwer D. A. Janneke , Beek Jackelieng Van , Ferwerda Harri , Brugman Astrid M. , Klis Fiona R. M. van der , Heiden H. Jacoline van der , Muskiet Frits A. J.
Published: September 2005

We investigated the effect of oral high-dose cholecalciferol on plasma and adipose tissue cholecalciferol and its subsequent release, and on plasma 25-hydroxyvitamin D (25(OH)D). Female Wistar rats (n126) received 37·5 μg cholecalciferol/d for 14 d and were subsequently studied for a further 88 d. Two subgroups of eighteen rats each were fasted for 3 d immediately after treatment (days 14−17) and at the end of the study (days 98−101). During treatment, plasma cholecalciferol increased rapidly to reach a steady-state. Plasma 25(OH)D and adipose tissue cholecalciferol increased linearly for 1 - 2 d after treatment. Serum Ca and inorganic phosphate also increased. Subsequently half-lives of plasma cholecalciferol and 25(OH)D, and perirenal and subcutaneous adipose tissue were: 1·4, 22·5, 97·5 and 80·9 d respectively. Fasting, as compared withad libitumfeeding, caused increased plasma free fatty acids, weight loss up to 14% and increased adipose tissue cholecalciferol (nmol/g wet weight). It did not affect plasma cholecalciferol immediately after cholecalciferol treatment, but raised plasma 25(OH)D. Fasting at the end of the study decreased plasma cholecalciferol and increased plasma 25(OH)D. We conclude that orally-administered cholecalciferol rapidly accumulates in adipose tissue and that it is very slowly released while there is energy balance. Fasting causes preferential loss of triacylglycerols from adipose tissue, as opposed to cholecalciferol, but nevertheless augments plasma 25(OH)D. Adipose tissue may act as a ‘buffer to functional vitamin D status’ by preventing, to a certain extent, unregulated production of 25(OH)D from dietary vitamin D, and by slowly releasing vitamin D under fasting conditions.
Comparison of oral and intraperitoneal iron supplementation in anaemic rats: a re-evaluation of the mucosal block theory of iron absorption
Authors: Benito Paloma , House William , Miller Dennis
Published: September 2005

To evaluate the extent to which daily oral Fe supplements may block Fe absorption from a subsequent dose, we compared effects of oral and intraperitoneal (i.p.) Fe supplementation on Fe status in anaemic rats (haemoglobin (Hb) 90 g/l). A ligated duodenal loop technique was used to assess the effects of the Fe supplements administered either orally or i.p. at different frequencies on Fe absorption from a subsequent test dose. Anaemic Sprague–Dawley rats were assigned to seven groups of eight rats each and received either oral or i.p. Fe supplements for 3 d as follows: (1) 4 mg oral supplement daily (three doses in 3 d); (2) 4 mg oral supplement once (one dose on day 1, low-Fe dose on days 2 and 3); (3) 12mg oral supplement once (one dose on day 1, low-Fe dose on days 2 and 3); (4) 3.2 mg i.p. supplement daily (three doses in 3d); (5) 3.2 mg i.p. supplement once (one dose on day 1); (6) 9.6 mg i.p. supplement once (one dose on day 1); (7) low-Fe diet (control). The effectiveness of the supplements in treating Fe deficiency on each of the two test-factors, i.e. route of administration and frequency of dose, was assessed by determining Hb-Fe gain and liver-Fe stores after the 3 d test period. Oral supplementation was as effective as i.p. in improving the Fe status of the anaemic animals. However, a 15 min absorption of a radio-Fe test dose from a ligated loop in i.p.-supplemented groups was significantly higher (11.68 (sd1.70) %, 17.49 (sd4.59) %, 16.71 (sd3.39) %) than in orally supplemented groups (3.24 (sd1.35) %, 2.45 (sd1.05) %, 1.80 (sd0.35) %) despite equal body Fe stores. No significant difference in intestinal Fe absorption efficiency was detected within the oral groups but those supplemented only once were more effective than or as effective as the group receiving daily supplements for 3d in improving Fe status as indicated by Hb-regeneration efficiency. We conclude that there is a mucosal block with the administration of oral Fe supplements but the extent of this blocking effect during oral Fe supplementation is not as dramatic as currently thought in the context of the poor efficacy of daily Fe supplementation programmes.
Comparing Experimental and Matching Methods Using a Large-Scale Voter Mobilization Experiment
Authors: Arceneaux Kevin , Gerber Alan S. , Green Donald P.
Published: September 2005

In the social sciences, randomized experimentation is the optimal research design for establishing causation. However, for a number of practical reasons, researchers are sometimes unable to conduct experiments and must rely on observational data. In an effort to develop estimators that can approximate experimental results using observational data, scholars have given increasing attention to matching. In this article, we test the performance of matching by gauging the success with which matching approximates experimental results. The voter mobilization experiment presented here comprises a large number of observations (60,000 randomly assigned to the treatment group and nearly two million assigned to the control group) and a rich set of covariates. This study is analyzed in two ways. The first method, instrumental variables estimation, takes advantage of random assignment in order to produce consistent estimates. The second method, matching estimation, ignores random assignment and analyzes the data as though they were nonexperimental. Matching is found to produce biased results in this application because even a rich set of covariates is insufficient to control for preexisting differences between the treatment and control group. Matching, in fact, produces estimates that are no more accurate than those generated by ordinary least squares regression. The experimental findings show that brief paid get-out-the-vote phone calls do not increase turnout, while matching and regression show a large and significant effect.
Sex chromosome evolution: Molecular aspects of Y-chromosome degeneration inDrosophila
Authors: Bachtrog Doris
Published: September 2005

Ancient Y-chromosomes of various organisms contain few active genes and an abundance of repetitive DNA. The neo-Y chromosome ofDrosophila mirandais in transition from an ordinary autosome to a genetically inert Y-chromosome, while its homolog, the neo-X chromosome, is evolving partial dosage compensation. Here, I compare four large genomic regions located on the neo-sex chromosomes that contain a total of 12 homologous genes. In addition, I investigate the partial coding sequence for 56 more homologous gene pairs from the neo-sex chromosomes. Little modification has occurred on the neo-X chromosome, and genes are highly constrained at the protein level. In contrast, a diverse array of molecular changes is contributing to the observed degeneration of the neo-Y chromosome. In particular, the four large regions surveyed on the neo-Y chromosome harbor several transposable element insertions, large deletions, and a large structural rearrangement. About one-third of all neo-Y-linked genes are nonfunctional, containing either premature stop codons and/or frameshift mutations. Intact genes on the neo-Y are accumulating amino acid and unpreferred codon changes. In addition, both 5′- and 3′-flanking regions of genes and intron sequences are less constrained on the neo-Y relative to the neo-X. Despite heterogeneity in levels of dosage compensation along the neo-X chromosome ofD. miranda, the neo-Y chromosome shows surprisingly uniform signs of degeneration.
De novo discovery of a tissue-specific gene regulatory module in a chordate
Authors: Johnson David S. , Zhou Qing , Yagi Kasumi , Satoh Nori , Wong Wing , Sidow Arend
Published: September 2005

We engage the experimental and computational challenges of de novo regulatory module discovery in a complex and largely unstudied metazoan genome. Our analysis is based on the comprehensive characterization of regulatory elements of 20 muscle genes in the chordate, Ciona savignyi. Three independent types of data we generate contribute to the characterization of a muscle-specific regulatory module: (1) Positive elements (PEs), short sequences sufficient for strong muscle expression that are identified in a high-resolution in vivo analysis; (2) CisModules (CMs), candidate regulatory modules defined by clusters of overrepresented motifs predicted de novo; and (3) Conserved elements (CEs), short noncoding sequences of strong conservation between C. savignyi and C. intestinalis. We estimate the accuracy of the computational predictions by an analysis of the intersection of these data. As final biological validation of the discovered muscle regulatory module, we implement a novel algorithm to search the genome for instances of the module and identify seven novel enhancers.
Coping with cold: The genome of the versatile marine Antarctica bacterium Pseudoalteromonas haloplanktis TAC125
Authors: Médigue Claudine , Krin Evelyne , Pascal Géraldine , Barbe Valérie , Bernsel Andreas , Bertin Philippe N. , Cheung Frankie , Cruveiller Stéphane , D'Amico Salvino , Duilio Angela , Fang Gang , Feller Georges , Ho Christine , Mangenot Sophie , Marino Gennaro , Nilsson Johan , Parrilli Ermenegilda , Rocha Eduardo P.C. , Rouy Zoé , Sekowska Agnieszka , Tutino Maria Luisa , Vallenet David , von Heijne Gunnar , Danchin Antoine
Published: September 2005

A considerable fraction of life develops in the sea at temperatures lower than 15°C. Little is known about the adaptive features selected under those conditions. We present the analysis of the genome sequence of the fast growing Antarctica bacterium Pseudoalteromonas haloplanktis TAC125. We find that it copes with the increased solubility of oxygen at low temperature by multiplying dioxygen scavenging while deleting whole pathways producing reactive oxygen species. Dioxygen-consuming lipid desaturases achieve both protection against oxygen and synthesis of lipids making the membrane fluid. A remarkable strategy for avoidance of reactive oxygen species generation is developed by P. haloplanktis, with elimination of the ubiquitous molybdopterin-dependent metabolism. The P. haloplanktis proteome reveals a concerted amino acid usage bias specific to psychrophiles, consistently appearing apt to accommodate asparagine, a residue prone to make proteins age. Adding to its originality, P. haloplanktis further differs from its marine counterparts with recruitment of a plasmid origin of replication for its second chromosome.
A genome-wide survey of structural variation between human and chimpanzee
Authors: Newman Tera L. , Tuzun Eray , Morrison V. Anne , Hayden Karen E. , Ventura Mario , McGrath Sean D. , Rocchi Mariano , Eichler Evan E.
Published: September 2005

Structural changes (deletions, insertions, and inversions) between human and chimpanzee genomes have likely had a significant impact on lineage-specific evolution because of their potential for dramatic and irreversible mutation. The low-quality nature of the current chimpanzee genome assembly precludes the reliable identification of many of these differences. To circumvent this, we applied a method to optimally map chimpanzee fosmid paired-end sequences against the human genome to systematically identify sites of structural variation ≥12 kb between the two species. Our analysis yielded a total of 651 putative sites of chimpanzee deletion (n = 293), insertions (n = 184), and rearrangements consistent with local inversions between the two genomes (n = 174). We validated a subset (19/23) of insertion and deletions using PCR and Southern blot assays, confirming the accuracy of our method. The events are distributed throughout the genome on all chromosomes but are highly correlated with sites of segmental duplication in human and chimpanzee. These structural variants encompass at least 24 Mb of DNA and overlap with >245 genes. Seventeen of these genes contain exons missing in the chimpanzee genomic sequence and also show a significant reduction in gene expression in chimpanzee. Compared with the pioneering work of Yunis, Prakash, Dutrillaux, and Lejeune, this analysis expands the number of potential rearrangements between chimpanzees and humans 50-fold. Furthermore, this work prioritizes regions for further finishing in the chimpanzee genome and provides a resource for interrogating functional differences between humans and chimpanzees.
AtPEX2 and AtPEX10 Are Targeted to Peroxisomes Independently of Known Endoplasmic Reticulum Trafficking Routes
Authors: Sparkes Imogen Averil , Hawes Chris , Baker Alison
Published: September 2005

Controversy exists in the literature over the involvement of the endoplasmic reticulum (ER) in the delivery of membrane proteins to peroxisomes. In this study, the involvement of the ER in the trafficking of two Arabidopsis (Arabidopsis thaliana) peroxisomal membrane proteins was investigated using confocal laser scanning microscopy of living cells expressing fusions between enhanced yellow fluorescent protein (eYFP) and AtPEX2 and AtPEX10. The fusion proteins were always detected in peroxisomes and cytosol irrespective of the location of the eYFP tag or the level of expression. The cytosolic fluorescence was not due to cleavage of the eYFP reporter from the C-terminal fusion proteins. Blocking known ER transport routes using the fungal metabolite Brefeldin A or expressing dominant negative mutants of Sar1 or RabD2a had no effect on the trafficking of AtPEX2 and AtPEX10 to peroxisomes. We conclude that AtPEX2 and AtPEX10 are inserted into peroxisome membranes directly from the cytosol.
Regulation of the Expression of Intracellular β-Carbonic Anhydrase in Response to CO2 and Light in the Marine Diatom Phaeodactylum tricornutum
Authors: Harada Hisashi , Nakatsuma Daisuke , Ishida Maki , Matsuda Yusuke
Published: September 2005

Cells of the marine diatom Phaeodactylum tricornutum Bohlin (UTEX 642) grown in 5% CO2 were transferred to air-level CO2 in the light or dark and allowed to acclimate to air. No accumulation of the transcript of the P. tricornutumβ-carbonic anhydrase 1 (ptca1) was detected in 5% CO2-grown cells, but ptca1 mRNA accumulated and reached a peak after 6 h acclimation to air but decreased over the next 18 h. A similar accumulation time course was observed in cells air-acclimated in the dark, except that levels of mRNA were <50% those in the light. These results suggest that air-level [CO2] is required to trigger the transcription of ptca1 and that light affects the extent of acclimation. During acclimation to air for 120 h in the light, levels of ptca1 mRNA exhibited a periodic oscillation with a cycle of about 24 h, which, however, was not reflected in protein accumulation levels. A 5′-upstream region from the transcription-start site toward −1,292 bp of ptca1 was cloned by inverse polymerase chain reaction, and 5′-truncations were carried out on this fragment. The truncated promoter regions were fused with the β-glucuronidase gene (uidA) and introduced into P. tricornutum. The promoter fragments, truncated at positions −1,292, −824, −484, −225, and −70 bp, conferred on transformants clear CO2-responsive β-glucuronidase expressions. In contrast, the CO2-responsive regulation was severely impaired or completely abolished by truncations, respectively, at position −50 or −30 bp. These results indicate that critical cis-elements required for CO2-responsive transcription of ptca1 may be located between −70 and −30 bp relative to the transcription start site.
Arabidopsis RAD51C Gene Is Important for Homologous Recombination in Meiosis and Mitosis
Authors: Abe Kiyomi , Osakabe Keishi , Nakayama Shigeki , Endo Masaki , Tagiri Akemi , Todoriki Setsuko , Ichikawa Hiroaki , Toki Seiichi
Published: September 2005

Rad51 is a homolog of the bacterial RecA recombinase, and a key factor in homologous recombination in eukaryotes. Rad51 paralogs have been identified from yeast to vertebrates. Rad51 paralogs are thought to play an important role in the assembly or stabilization of Rad51 that promotes homologous pairing and strand exchange reactions. We previously characterized two RAD51 paralogous genes in Arabidopsis (Arabidopsis thaliana) named AtRAD51C and AtXRCC3, which are homologs of human RAD51C and XRCC3, respectively, and described the interaction of their products in a yeast two-hybrid system. Recent studies showed the involvement of AtXrcc3 in DNA repair and functional role in meiosis. To determine the role of RAD51C in meiotic and mitotic recombination in higher plants, we characterized a T-DNA insertion mutant of AtRAD51C. Although the atrad51C mutant grew normally during vegetative developmental stage, the mutant produced aborted siliques, and their anthers did not contain mature pollen grains. Crossing of the mutant with wild-type plants showed defective male and female gametogeneses as evidenced by lack of seed production. Furthermore, meiosis was severely disturbed in the mutant. The atrad51C mutant also showed increased sensitivity to γ-irradiation and cisplatin, which are known to induce double-strand DNA breaks. The efficiency of homologous recombination in somatic cells in the mutant was markedly reduced relative to that in wild-type plants.
A Lily ASR Protein Involves Abscisic Acid Signaling and Confers Drought and Salt Resistance in Arabidopsis
Authors: Yang Chin-Ying , Chen Yu-Chuan , Jauh Guang Yuh , Wang Co-Shine
Published: September 2005

LLA23, an abscisic acid-, stress-, and ripening-induced protein, was previously isolated from lily (Lilium longiflorum) pollen. The expression of LLA23 is induced under the application of abscisic acid (ABA), NaCl, or dehydration. To provide evidence on the biological role of LLA23 proteins against drought, we used an overexpression approach in Arabidopsis (Arabidopsis thaliana). Constitutive overexpression of LLA23 under the cauliflower mosaic virus 35S promoter confers reduced sensitivity to ABA in Arabidopsis seeds and, consequently, a reduced degree of seed dormancy. Transgenic 35S∷LLA23 seeds are able to germinate under unfavorable conditions, such as inhibitory concentrations of mannitol and NaCl. At the molecular level, altered expression of ABA/stress-regulated genes was observed. Thus, our results provide strong in vivo evidence that LLA23 mediates stress-responsive ABA signaling. In vegetative tissues, it is intriguing that Arabidopsis 35S∷LLA23 stomata remain opened upon drought, while transgenic plants have a decreased rate of water loss and exhibit enhanced drought and salt resistance. A dual function of the lily abscisic acid-, stress-, and ripening-induced protein molecule is discussed.
Cell-to-Cell Movement of Green Fluorescent Protein Reveals Post-Phloem Transport in the Outer Integument and Identifies Symplastic Domains in Arabidopsis Seeds and Embryos
Authors: Stadler Ruth , Lauterbach Christian , Sauer Norbert
Published: September 2005

Developing Arabidopsis (Arabidopsis thaliana) seeds and embryos represent a complex set of cell layers and tissues that mediate the transport and partitioning of carbohydrates, amino acids, hormones, and signaling molecules from the terminal end of the funicular phloem to and between these seed tissues and eventually to the growing embryo. This article provides a detailed analysis of the symplastic domains and the cell-to-cell connectivity from the end of the funiculus to the embryo, and within the embryo during its maturation. The cell-to-cell movement of the green fluorescent protein or of mobile and nonmobile green fluorescent protein fusions was monitored in seeds and embryos of plants expressing the corresponding cDNAs under the control of various promoters (SUC2, SUC3, TT12, and GL2) shown to be active in defined seed or embryo cell layers (SUC3, TT12, and GL2) or only outside the developing Arabidopsis seed (AtSUC2). Cell-to-cell movement was also analyzed with the low-molecular-weight fluorescent dye 8-hydroxypyrene-1,3,6-trisulfonate. The analyses presented identify a phloem-unloading domain at the end of the funicular phloem, characterize the entire outer integument as a symplastic extension of the phloem, and describe the inner integument and the globular stage embryo plus the suspensor as symplastic domains. The results also show that, at the time of hypophysis specification, the symplastic connectivity between suspensor and embryo is reduced or interrupted and that the embryo develops from a single symplast (globular and heart stage) to a mature embryo with new symplastic domains.
Lipid Signaling in Plants. Cloning and Expression Analysis of the Obtusifoliol 14α-Demethylase fromSolanum chacoenseBitt., a Pollination- and Fertilization-Induced Gene with Both Obtusifoliol and Lanosterol Demethylase Activity
Authors: O'Brien Martin , Chantha Sier-Ching , Rahier Alain , Matton Daniel P.
Published: September 2005

The sterol 14α-demethylase (CYP51) is the most widely distributed cytochrome P450 gene family being found in all biological kingdoms. It catalyzes the first step following cyclization in sterol biosynthesis, leading to the formation of precursors of steroid hormones, including brassinosteroids, in plants. Most enzymes involved in the plant sterol biosynthesis pathway have been characterized biochemically and the corresponding genes cloned. Genes coding for enzymes promoting substrate modifications before 24-methylenelophenol lead to embryonic and seed defects when mutated, while mutants downstream the 24-methylenelophenol intermediate show phenotypes characteristic of brassinosteroid mutants. By a differential display approach, we have isolated a fertilization-induced gene, encoding a sterol 14α-demethylase enzyme, named CYP51G1-Sc. Functional characterization of CYP51G1-Sc expressed in yeast (Saccharomyces cerevisiae) showed that it could demethylate obtusifoliol, as well as nontypical plant sterol biosynthetic intermediates (lanosterol), in contrast with the strong substrate specificity of the previously characterized obtusifoliol 14α-demethylases found in other plant species. CYP51G1-Sc transcripts are mostly expressed in meristems and in female reproductive tissues, where they are induced following pollination. Treatment of the plant itself with obtusifoliol induced the expression of the CYP51G1-Sc mRNA, suggesting a possible role of this transient biosynthetic intermediate as a bioactive signaling lipid molecule. Furthermore, treatments of leaves with 14C-labeled obtusifoliol demonstrated that this sterol could be transported in distal parts of the plant away from the sprayed leaves. Arabidopsis (Arabidopsis thaliana) CYP51 homozygous knockout mutants were also lethal, suggesting important roles for this enzymatic step and its substrate in plant development.