Effect of high levels of dietary folic acid on folate metabolism in vitamin B12 deficiency

The effect of administering high levels of folic acid to vitamin B12-deficient animals was studied. In B12 deficiency histidine oxidation is decreased. This is the result of both decreased liver folate levels and increases in the proportion of methyltetrahydrofolates. The purpose of this study was to determine if the addition of very high levels of folic acid to B12-deficient diets could increase liver folates and thereby restore histidine oxidation. Rats were fed a soy protein B12-deficient diet containing 10% pectin which has been shown previously to accelerate B12 depletion. When this diet was supplemented with B12 and folic acid, histidine oxidation was 5.4% in 2 h and the livers contained 3.49 micrograms of folate/g. In the absence of B12, the histidine oxidation rate was 0.34% and the liver folate level was 1.33 micrograms/g. When 200 mg/kg of folic acid was added to the B12-deficient diet there was no increase in histidine oxidation (0.35%) but the liver folates were increased to 3.68 micrograms which is about the same as that with B12 supplementation. The percentage tetrahydrofolate of the total liver folates was the same with and without a high level of dietary folic acid. Thus there was an increase in the absolute level of tetrahydrofolate without any increase in folate function as measured by histidine oxidation. Red cell folate levels were the same with and without B12, which is in contrast to the markedly lower liver folate levels in B12 deficiency. These data suggest a difference between B12 regulation of folate metabolism in the liver and in the bone marrow.     

Effects of dietary zinc deficiency on homocysteine and folate metabolism in rats

In rats, zinc deficiency has been reported to result in elevated hepatic methionine synthase activity and alterations in folate metabolism. We investigated the effect of zinc deficiency on plasma homocysteine concentrations and the distribution of hepatic folates. Weanling male rats were fed ad libitum a zinc-sufficient control diet (382.0 nmol zinc/g diet), a low-zinc diet (7.5 nmol zinc/g diet), or a control diet pair-fed to the intake of the zinc-deficient rats. After 6 weeks, the body weights of the zinc-deficient and pair-fed control groups were lower than those of controls, and plasma zinc concentrations were lowest in the zinc-deficient group. Plasma homocysteine concentrations in the zinc-deficient group (2.3 +/- 0.2 micromol/L) were significantly lower than those in the ad libitum-fed and pair-fed control groups (6.7 +/- 0.5 and 3.2 +/- 0.4 micromol/L, respectively). Hepatic methionine synthase activity in the zinc-deficient group was higher than in the other two groups. Low mean percentage of 5-methyltetrahydrofolate in total hepatic folates and low plasma folate concentration were observed in the zinc-deficient group compared with the ad libitum-fed and pair-fed control groups. The reduced plasma homocysteine and folate concentrations and reduced percentage of hepatic 5-methyltetrahydrofolate are probably secondary to the increased activity of hepatic methionine synthase in zinc deficiency.     

Immunization of male mice with luteinizing hormone-releasing hormone fusion proteins reduces testicular and accessory sex gland function

Genes for ovalbumin-luteinizing hormone-releasing hormone 7 (LHRH-7) and thioredoxin-LHRH-7 fusion proteins (containing seven LHRH inserts) were constructed by cassette and mismatch mutagenesis and expressed in Escherichia coli. In experiment 1, 10 microgram of either ovalbumin-LHRH-7 or thioredoxin-LHRH-7 were suspended in Z-max adjuvant and injected three times at 4-wk intervals into postpubertal male BALB/c mice. In experiment 2, the fusion proteins were suspended in Immumax adjuvant and administered in equimolar quantities (0.4 nmol per injection) to postpubertal male BALB/c mice. In addition to injection of these two proteins alone, the proteins were also administered in different sequences or together in a mixture. Both LHRH fusion proteins induced significant antibody titers, which resulted in a significant decrease in vesicular gland and anterior prostate weight (measure of biological response) in both experiments. Vesicular gland and anterior prostate weight and LHRH antibody titers were significantly correlated in experiments 1 (r = -0.64) and 2 (r = -0.53). Percentage of animals responding to treatment varied from 40-60% in experiment 1 and from 11-89% in experiment 2, with the highest responses in treatments that used a combination of both fusion proteins. The variation in responders and nonresponders was evaluated by estimating antibody K(D) from displacement curves. Part, but not all, of the high antibody nonresponders can be explained by antibody affinity.     

High hexokinase activity in tomato fruit perturbs carbon and energy metabolism and reduces fruit and seed size

Tomato (Lycopersicon esculentum var MP‐1) plants overexpressing Arabidopsis hexokinase 1 (AtHXK1) exhibited high hexokinase (HXK) activity in correlation with drastic phenotypic modifications in fruit. Transgenic fruit and seeds were reduced in size. Reduction in fruit size was due to decreased cell expansion, which could not be corrected by perfusion with sucrose (Suc). Neither could wild type (WT) fruit and seed size be obtained by grafting of transgenic flowers onto WT shoots. Starch and hexose contents were lower but organic and amino acids were higher in transgenic fruit. Lower respiratory rates measured in vitro accompanied by even lower ATP levels and ATP/ADP ratios indicated metabolic perturbations that may explain, in part, reduced fruit and seed size.     

Chronic effects of ethanol on muscle metabolism in the rat

Chronic ethanol feeding in the rat is associated with a skeletal myopathy involving primarily type-II muscle fibers, which is recognised to be mediated via a specific impairment in protein turnover. This paper investigates whether the cause of this myopathy may be related to abnormalities in carbohydrate and lipid metabolism in different muscles. [U-¹⁴C]Glucose metabolism was examined in two muscles with different fibre compsitions, the extensor digitorum longus (EDL) muscle, which contains predominantly type-II muscle fibres, and the soleus muscle, which is composed primarily of type-I muscle fibres. Feeding on the ethanol-supplemented Lieber-DeCarli liquid diet for 2 or 6 weeks was associated with profound distubances in glucose metabolism in both EDL and soleus muscles, particularly in relation to rates of glycogen and alanine formation. We discuss the importance of these metabolic changes in relation to the genesis of chronic alcoholic skeletal myopathy.     

Effect of salmon gonadotropic hormone on sex steroids in male rainbow trout: plasma levels and testicular secretion in vitro

Summary Male rainbow trout were treated with salmon gonadotropic hormone (GTH) at different stages of the circannual reproductive cycle; spawning fish were also treated with an antiserum against salmon GTH. Injection of GTH led to a several-fold increase of plasma sex steroid levels during spermatogenesis and in the spawning season but was without effect at early stages of testicular development. GTH neutralization during the spawning season was followed by a several-fold decrease of plasma sex steroid levels. During spermatogenesis and in the spawning season, both treatment regimes resulted in an increased sensitivity of testicular explants in response to a subsequent stimulation of steroid secretion in vitro. This up-regulatory response may facilitate and maintain the high sex steroid plasma levels observed during the spawning season. It may also be necessary to allow for concomitant peak values of plasma GTH and sex steroids in the spawning season, a situation difficult to understand within the negative feedback concept. The adaptive capacities of the testicular steroidogenic system indicate that it is not only an effector site for GTH but also an active part of the endocrine system controling reproduction.Abbreviations BSA bovine serum albumin - bw body weight - E₂ 17-estradiol - GnRH gonadotropin releasing-hormone - GTH gonadotropic hormone - LH luteinizing hormone - OHT 11-hydroxytestosterone - OT 11-ketotestosterone - 17-20P 17-hydroxy, 20-dihydroprogesterone - PE pituitary extract - raGTH rabbit anti-GTH antiserum - rPS rabbit preimmune serum - T testosterone     

Methylenetetrahydrofolate reductase deficiency and low dietary folate increase embryonic delay and placental abnormalities in mice

BACKGROUND: Despite extensive research on mild methylenetetrahydrofolate reductase (MTHFR) deficiency and low dietary folate in different disorders, the association of these metabolic disturbances with a variety of congenital defects and pregnancy complications remains controversial. In this study we investigated the effects of MTHFR and dietary folate deficiency at 10.5 days post coitum (dpc) in our mouse model of mild MTHFR deficiency. METHODS: Mthfr +/+ and +/? female mice were fed a control or folic acid–deficient diet for 6 weeks, then mated with Mthfr +/? males. At 10.5 dpc, embryos were examined and placentae were collected for histologic evaluation. RESULTS: Maternal MTHFR and folate deficiencies resulted in increased developmental delays and smaller embryos. We also observed a low frequency of a variety of embryonic defects in the experimental groups, such as neural tube, heart looping, and turning defects; these results mimic the low incidence and multifactorial nature of these anomalies in humans. Folate‐deficient mice also had increased embryonic losses and severe placental defects, including placental abruption and disturbed patterning of placental layers. Folate‐deficient placentae had decreased ApoA‐I expression, and there was a trend toward a negative correlation between ApoA‐I expression with maternal homocysteine concentrations. CONCLUSIONS: Our study provides biological evidence linking maternal MTHFR and dietary folate deficiencies to adverse pregnancy outcomes in mice. It underscores the importance of folate not only in reducing the incidence of early embryonic defects, but also in the prevention of developmental delays and placental abnormalities that may increase susceptibility to other defects and to reproductive complications. Birth Defects Research (Part A), 2009. © 2009 Wiley‐Liss, Inc.     

Non-invasive measurement of blood pressures in the Yucatan micropig (Sus scrofa domestica), with and without midazolam-induced sedation

Current literature suggests that the effects of midazolam, a water-soluble benzodiazepine, on blood pressure in swine are minimal. The hypothesis of the study reported here was that a light sedative dose would induce a decrease in blood pressure in this species. Healthy female Yucatan Micropigs (n = 20), 16 to 30 (mean, 22) kg, aged four six months, were individually placed in a humane restraint sling and allowed to acclimate. Systolic (SBP), diastolic (DBP), and mean (MBP) blood pressures (mmHg) and heart rate (HR; beats per min [bpm]) were measured by use of oscillometry. The pressure cuff was placed at the base of the tail, and five sets of values were recorded at five-min intervals, beginning at 10 and ending 30 min after cuff placement. Following a three- to four-day rest period, this procedure was repeated with the addition of a dose of 0.5 mg of midazolam HCl/kg of body weight given intramuscularly at the time of cuff placement. A paired one-way Student's t-test was used to compare the means of the five measures between control and midazolam treatment. Mean (+/- SD) differences for SBP, DBP, MBP, and HR were 18.9 (+/- 3.97), 17.8 (+/- 5.27), and 18.6 (+/- 5.09) mmHg and 20.7 (+/- 3.73) bpm, respectively. All four parameters were significantly reduced in the midazolam-sedated group (P < 0.001). The maximal decrease in SBP, DBP, and MBP occurred at 15 and 20 min after dosing. Mean values based on the means of the five measures were 128 (+/- 12.6), 80 (+/- 9.4), and 99 (+/- 9.2) mmHg and 135 (+/- 17.4) bpm, and 109 (+/- 15.4), 63 (+/- 12.6), and 80 (+/- 13.6) mmHg and 115 (+/- 15.5) bpm for SBP, DBP, MBP, and HR in the control (n = 20) and midazolam (n = 20) groups, respectively. The control values can serve as normal oscillometric values for this age, sex, and breed of Micropig. We conclude that midazolam, given intramuscularly at a sedative dosage, negatively affects cardiovascular parameters measured by use of a blood pressure cuff, in sexually mature female Micropigs, compared with values in untreated pigs, which is similar to reports for humans.     

Somatomedin levels in the diagnosis and therapy of growth hormone deficiency

Serum somatomedin-C (SM-C) and somatomedin (SM) concentrations were measured by, respectively, radioimmuno (SM-C RIA) and radioreceptor assays (SM RRA) in 3 groups of children with short stature. The patient population was different from previously reported series in that it was urban Brazilian, low income, and significantly older. Group A consisted of 6 male and 3 female children, aged 7.7-16.0 years, whose average peak plasma immunoreactive growth hormone (GH) was above 10 ng/ml. Group B contained 8 male and 5 female untreated GH-deficient patients, ranging in age from 9.5 to 21.0 years. In Group C there were 4 male and 1 female GH-deficient subjects treated with I.M. injections of GH (0.1 U/kg) from 1 month to 7 years. The mean +/- SE basal RIA SM-C (ng/ml) concentrations were significantly lower in groups B (34.2 +/- 8.8) and C (43.8 +/- 13.7) than A (214.3 +/- 42.7): A X B, P less than 0.001 and A X C, P less than 0.02. Likewise the mean +/- SE basal RRA SM (ng/ml) concentrations were significantly lower in groups B (78.9 +/- 17.6) and C (90.8 +/- 19.3) than group A (316.3 +/- 43.0): A X B, P less than 0.001 and A X C, P less than 0.002. A significant linear correlation was observed between RIA and RRA in group B (r = 0.84; P less than 0.001) and C (r = 0.96; P less than 0.01), but not for A (r = 0.61; P greater than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

The role of selenium in thyroid hormone metabolism and effects of selenium deficiency on thyroid hormone and iodine metabolism

Selenium deficiency impairs thyroid hormone metabolism by inhibiting the synthesis and activity of the iodothyronine deiodinases, which convert thyroxine (T₄) to the more metabolically active 3,3′-5 triiodothyronine (T₃). Hepatic type I iodothyronine deiodinase, identified in partially purified cell fractions using affinity labeling with [¹²⁵I]N-bromoacetyl reverse triiodothyronine, is also labeled with⁷⁵Se by in vivo treatment of rats with⁷⁵Se-Na₂SeO₃. Thus, the type I iodothyronine 5′-deiodinase is a selenoenzyme. In rats, concurrent selenium and iodine deficiency produces greater increases in thyroid weight and plasma thyrotrophin than iodine deficiency alone. These results indicate that a concurrent selenium deficiency could be a major determinant of the severity of iodine deficiency.     

The role of selenium in thyroid hormone metabolism and effects of selenium deficiency on thyroid hormone and iodine metabolism

Selenium deficiency impairs thyroid hormone metabolism by inhibiting the synthesis and activity of the iodothyronine deiodinases, which convert thyroxine (T₄) to the more metabolically active 3,3′–5 triiodothyronine (T₃). Hepatic type I iodothyronine deiodinase, identified in partially purified cell fractions using affinity labeling with [¹²⁵I]N-bromoacetyl reverse triiodothyronine, is also labeled with⁷⁵Se by in vivo treatment of rats with⁷⁵Se−Na₂SeO₃. Thus, the type I iodothyronine 5′-deiodinase is a selenoenzyme. In rats, concurrent selenium and iodine deficiency produces greater increases in thyroid weight and plasma thyrotrophin than iodine deficiency alone. These results indicate that a concurrent selenium deficiency could be a major determinant of the severity of iodine deficiency.     

Serum and saliva sex hormone levels in !Kung San men

Serum concentrations of testosterone (Tser), 5 alpha-dihydrotestosterone (DHT), estradiol 17 beta (E2), and free testosterone in saliva (Tsal) were determined by means of the radioimmunoassay method in 114 !Kung San men living in the Bushmanland district of Namibia. The healthy men (mean age 26.4 years) were asked about their dietary habits over the last two months and their acute alcohol intake during the 24 hours preceding the blood and saliva sampling. Although the sex hormone status of the !Kung lies within the range of normal men reported for Caucasoid samples, both Tser and Tsal exhibit relatively low concentrations in comparison to the great majority of published mean values. On the other hand, comparatively high DHT levels point to an elevated 5 alpha-reduction of testosterone to DHT in our sample. Estradiol concentrations show no deviation from normal values reported elsewhere for healthy young men. Different dietary habits of the !Kung lead to significant differences in their sex hormone status: both levels of Tsal and the androgen ratio Tsal/Tser decrease with increasing supplement of the traditional hunter-gatherer diet with domestic and Western food products. The amount of alcohol consumed during the day before the blood and saliva sampling shows a significant effect on the DHT metabolism, and the shorter the time after drinking, the greater decrease of DHT and DHT/E2 can be observed.     

Impact of methylenetetrahydrofolate reductase deficiency and low dietary folate on the development of neural tube defects in splotch mice

BACKGROUND: The etiology of neural tube defects (NTDs) is multifactorial, with environmental and genetic determinants. Folate supplementation prevents the majority of NTDs, and a polymorphism in methylenetetrahydrofolate reductase (MTHFR) has become recognized as a genetic risk factor. The mechanisms by which folate affects NTD development are unclear. The Splotch (Sp) mouse is a well-characterized mouse model for studying spontaneous NTDs. To assess the potential interaction between folate metabolism and the Sp mutant in NTD development, we studied mice with both Sp and Mthfr mutations, as well as the interaction between Sp and low dietary folate. METHODS: Wild-type, single Mthfr+/-mutant, single Sp/+mutant, and double mutant (Mthfr+/-, Sp/+) female mice were mated with males of the same genotype. Embryos were examined for NTDs on gestational day (GD) 13.5. To investigate the effects of folate deficiency on Sp mice, Sp/+female mice were fed a control diet (CD), a moderately folic acid-deficient diet (MFADD), or a severely folic acid-deficient diet (SFADD). They were mated with Sp/+males and the embryos were examined. RESULTS: There were no differences in the incidence or severity of NTDs in embryos from double-mutant mating pairs compared to those from single Sp mutants. Embryos from Mthfr+/-dams did not exhibit NTDs. Diets deficient in folate did not influence the incidence or severity of NTDs in embryos from Sp/+mice. CONCLUSIONS: We did not observe an interaction between Sp and Mthfr mutations, or between the Sp mutation and low dietary folate, in NTD development in Splotch mice.     

Modulation of alcohol dehydrogenase and ethanol metabolism by sex hormones in the spontaneously hypertensive rat. Effect of chronic ethanol administration

In young (4-week-old) male and female spontaneously hypertensive (SH) rats, ethanol metabolic rate in vivo and hepatic alcohol dehydrogenase activity in vitro are high and not different in the two sexes. In males, ethanol metabolic rate falls markedly between 4 and 10 weeks of age, which coincides with the time of development of sexual maturity in the rat. Alcohol dehydrogenase activity is also markedly diminished in the male SH rat and correlates well with the changes in ethanol metabolism. There is virtually no influence of age on ethanol metabolic rate and alcohol dehydrogenase activity in the female SH rat. Castration of male SH rats prevents the marked decrease in ethanol metabolic rate and alcohol dehydrogenase activity, whereas ovariectomy has no effect on these parameters in female SH rats. Chronic administration of testosterone to castrated male SH rats and to female SH rats decreases ethanol metabolic rate and alcohol dehydrogenase activity to values similar to those found in mature males. Chronic administration of oestradiol-17β to male SH rats results in marked stimulation of ethanol metabolic rate and alcohol dehydrogenase activity to values similar to those found in female SH rats. Chronic administration of ethanol to male SH rats from 4 to 11 weeks of age prevents the marked age-dependent decreases in ethanol metabolic rate and alcohol dehydrogenase activity, but has virtually no effect in castrated rats. In the intoxicated chronically ethanol-fed male SH rats, serum testosterone concentrations are significantly depressed. In vitro, testosterone has no effect on hepatic alcohol dehydrogenase activity of young male and female SH rats. In conclusion, in the male SH rat, ethanol metabolic rate appears to be limited by alcohol dehydrogenase activity and is modulated by testosterone. Testosterone has an inhibitory effect and oestradiol has a testosterone-dependent stimulatory effect on alcohol dehydrogenase activity and ethanol metabolic rate in these animals.     

Effects of dietary zinc deficiency on gonadotrophin secretion and testicular growth in young male sheep.

The hypothesis that the secretion of gonadotrophins would be reduced by zinc deficiency was tested in five groups of four young Merino rams (initial liveweight 22 kg). Four groups were fed ad libitum with diets containing 4, 10, 17 or 27 micrograms Zn g-1. The effects of loss of appetite on the deficient diet was controlled by feeding a fifth group (pair-fed control) at a rate of 27 micrograms Zn g-1, but the amount of feed offered was restricted to that eaten voluntarily by the deficient (4 micrograms Zn g-1) group. Blood was sampled every 20 min for 32 h on two occasions before the treatments were imposed and 96 days later, at the end of the experiment. The rams were injected with gonadotrophin-releasing hormone (GnRH; 10 ng kg-1 i.v.) after each serial sampling, and with naloxone (1 mg kg-1 i.v.) 24 h after the end of the final GnRH test. In the group that were fed the diet with the lowest zinc content, the concentration of zinc in blood plasma was reduced to 18% of that in the pair-fed controls (P < 0.05) and was within the deficient range. The appetite of the deficient rams was half that of the controls fed 27 micrograms Zn g-1 ad libitum and there was no increase in liveweight or testicular diameter during pubertal development. Similar, but smaller, effects were observed in the pair-fed controls. There were no significant differences between pair-fed and deficient groups in the frequency of the luteinizing hormone (LH) pulses or in the concentration of follicle-stimulating hormone (FSH), but the secretion of gonadotrophins was markedly lower in both groups than in the control rams fed ad libitum. The response to GnRH was not affected by treatment, but the increase in LH pulse frequency evoked by naloxone was lower in the deficient animals than in other groups. The animals fed zinc at intermediate rates (10-17 micrograms g-1) showed similar responses to the controls fed ad libitum. It is concluded that the specific effects of zinc deficiency on testicular function were small. Most of the reduction in testicular growth in rams fed a deficient diet was not specifically related to the trace element, but was due to the fall in energy and protein intake caused by the loss of appetite. This leads to a reduction in the frequency of GnRH pulses secreted by the hypothalamus, and to low rates of gonadotrophin secretion by the pituitary gland.