MR1-restricted T cells (MR1Ts) are a T cell subset that recognize and mediate host defense to a broad array of microbial pathogens, including respiratory pathogens (e.g., Mycobacterium tuberculosis, Streptococcus pyogenes, and Francisella tularensis) and enteric pathogens (e.g., Escherichia coli and Salmonella species). Mucosal-associated invariant T (MAIT) cells, a subset of MR1Ts, were historically defined by the use of a semi-invariant T cell receptor (TCR) and recognition of small molecules derived from the riboflavin biosynthesis pathway presented on MR1. We used mass spectrometry to identify the repertoire of ligands presented by MR1 from the microbes E. coli and Mycobacterium smegmatis We found that the MR1 ligandome is unexpectedly broad, revealing functionally distinct ligands derived from E. coli and M. smegmatis The identification, synthesis, and functional analysis of mycobacterial ligands reveal that MR1T ligands can be distinguished by MR1Ts with diverse TCR usage. These data demonstrate that MR1 can serve as an immune sensor of the microbial ligandome.

Silymarin, a known standardized extract obtained from seeds of Silybum marianum is widely used in treatment of several diseases of varying origin. In the present paper, we clarified the antioxidant activity of silymarin by employing various in vitro antioxidant assay such as 1,1-diphenyl-2-picryl-hydrazyl free radical (DPPH(.)) scavenging, 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) radical scavenging activity, total antioxidant activity determination by ferric thiocyanate, total reducing ability determination by Fe3+ - Fe2+ transformation method and Cuprac assay, superoxide anion radical scavenging by riboflavin/methionine/illuminate system, hydrogen peroxide scavenging and ferrous ions (Fe2+) chelating activities. Silymarin inhibited 82.7% lipid peroxidation of linoleic acid emulsion at 30 microg/mL concentration; butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), alpha-tocopherol and trolox indicated inhibition of 83.3, 82.1, 68.1 and 81.3% on peroxidation of linoleic acid emulsion at the same concentration, respectively. In addition, silymarin had an effective DPPH(.) scavenging, ABTS(.)+ scavenging, superoxide anion radical scavenging, hydrogen peroxide scavenging, ferric ions (Fe3+) reducing power by Fe3+-Fe2+ transformation, cupric ions (Cu2+) reducing ability by Cuprac method, and ferrous ions (Fe2+) chelating activities. Also, BHA, BHT, alpha-tocopherol and trolox, were used as the reference antioxidant and radical scavenger compounds. Moreover, this study, which clarifies antioxidant mechanism of silymarin, brings new information on the antioxidant properties of silymarin. According to the present study, silymarin had effective in vitro antioxidant and radical scavenging activity. It could be used in the pharmacological and food industry because of its antioxidant properties.

Bdellovibrio and like organisms (BALO) are obligate predators of Gram-negative bacteria, belonging to the α- and δ-proteobacteria. BALO prey using either a periplasmic or an epibiotic predatory strategy, but the genetic background underlying these phenotypes is not known. Here we compare the epibiotic Bdellovibrio exovorus and Micavibrio aeruginosavorus to the periplasmic B. bacteriovorus and Bacteriovorax marinus. Electron microscopy showed that M. aeruginosavorus, but not B. exovorus, can attach to prey cells in a non-polar manner through its longitudinal side. Both these predators were resistant to a surprisingly high number of antibiotic compounds, possibly via 26 and 19 antibiotic-resistance genes, respectively, most of them encoding efflux pumps. Comparative genomic analysis of all the BALOs revealed that epibiotic predators have a much smaller genome (ca. 2.5 Mbp) than the periplasmic predators (ca. 3.5 Mbp). Additionally, periplasmic predators have, on average, 888 more proteins, at least 60% more peptidases, and one more rRNA operon. Fifteen and 219 protein families were specific to the epibiotic and the periplasmic predators, respectively, the latter clearly forming the core of the periplasmic 'predatome', which is upregulated during the growth phase. Metabolic deficiencies of epibiotic genomes include the synthesis of inosine, riboflavin, vitamin B6 and the siderophore aerobactin. The phylogeny of the epibiotic predators suggests that they evolved by convergent evolution, with M. aeruginosavorus originating from a non-predatory ancestor while B. exovorus evolved from periplasmic predators by gene loss.

BACKGROUND

Vitamin B(2) exists in blood as riboflavin and its cofactors, flavin mononucleotide (FMN) and FAD. The erythrocyte glutathione reductase activation coefficient (EGRAC) has traditionally been used to assess vitamin B(2) status in humans. We investigated the relationships of EGRAC and plasma and erythrocyte concentrations of riboflavin, FMN, and FAD in elderly volunteers and their responses to riboflavin administration.

METHODS

EGRAC and plasma and erythrocyte concentrations of riboflavin, FMN, and FAD were determined in 124 healthy individuals with a mean age of 69 years. The same measurements were made in a subgroup of 46 individuals with EGRAC>> or =1.20 who participated in a randomized double-blind 12-week intervention study and received riboflavin (1.6 mg/day; n = 23) or placebo (n = 23).

RESULTS

Median plasma concentrations were 10.5 nmol/L for riboflavin, 6.6 nmol/L for FMN, and 74 nmol/L for FAD. In erythrocytes, there were only trace amounts of riboflavin, whereas median FMN and FAD concentrations were 44 and 469 nmol/L, respectively. Erythrocyte FMN and FAD correlated with each other and with EGRAC and plasma riboflavin (P <0.05). All variables except plasma FAD responded significantly to riboflavin supplementation compared with placebo (P < or =0.04). The strongest increases were for riboflavin in plasma (83%) and for FMN in erythrocytes (87%).

CONCLUSIONS

Concentrations of all B(2) vitamers except plasma FAD are potential indicators of vitamin B(2) status, and plasma riboflavin and erythrocyte FMN may be useful for the assessment of vitamin B(2) status in population studies.

Frequent consumption of supplements is a common behavior in adults, as well as children and adolescents. We report on vitamin intake from consuming supplements, taking into account the vitamin intake from fortified and usual foods. A total of 5990 3-d weighed dietary records from 931 German subjects (452 males, 479 females, 2-18 y of age) between 1986 and 2003 from the Dortmund Nutritional and Anthropometric Longitudinally Designed (DONALD) Study were evaluated. We identified 133 different vitamin-containing supplements in 451 records (7.5%). Slightly more males (8.0%) than females (7.1%) consumed supplements, with highest frequencies found between ages 15 and 18 y. For the majority (10 of 13) of vitamins (vitamin A, vitamin E, vitamin K, thiamin, riboflavin, vitamin B-6, vitamin B-12, niacin, biotin, and vitamin C), intake from usual and fortified food already reached or exceeded 80% of the recommended dietary allowances in all age groups. In the case of folate and pantothenic acid, intake from supplements was necessary to achieve at least 80% of recommended dietary allowances for half of the age groups, especially in females. Supplements with vitamin D considerably helped increase vitamin intake but failed to achieve 80% of references in almost all age groups. Intakes of vitamin A and folic acid exceeded the tolerable upper level most frequently in many age groups and were most pronounced in up to 32% and 13% of children aged 2-3 y. Intake of vitamin D, vitamin E, and vitamin C exceeded the tolerable upper level in single age groups only (<7% of subjects). For the other vitamins, no exceeded limits were identified. The ubiquitous availability of supplements might indicate that even healthy children and adolescents profit by taking them. However, it is difficult to evaluate whether consumers receive more benefits or risks from the unrestricted consumption of supplements as they are marketed today.

OBJECTIVE

To evaluate the relation between nutrient intake and psychiatric functioning in adults with confirmed mood disorders.

METHODS

A cross-sectional study was conducted of the intake of major (that is, carbohydrates, fat, and protein) and minor (that is, vitamins and minerals) nutrients (from 3-day food records and a Food Frequency Questionnaire), Global Assessment of Functioning (GAF) scores, and symptoms of depression and mania (the Hamilton Depression Rating Scale and the Young Mania Rating Scale) in 97 community-based adults with mood disorders whose diagnoses were confirmed with structured interviews.

RESULTS

Significant correlations were found between GAF scores and energy (kilocalories), carbohydrates, fibre, total fat, linoleic acid, riboflavin, niacin, folate, vitamin B6, vitamin B12, pantothenic acid, calcium, phosphorus, potassium, and iron (all P values < 0.05), as well as magnesium (r = 0.41, P < 0.001) and zinc (r = 0.35, P < 0.001). Though modest in magnitude, the pattern of correlations was consistent, indicating higher levels of mental function associated with a higher intake of nutrients. Depression and mania scores, which were generally mild or moderate, did not individually show consistent patterns. When dietary supplement use was added to nutrient intakes from food, GAF scores remained positively correlated (P < 0.05) with all dietary minerals.

CONCLUSIONS

This detailed analysis in a clinically diagnosed sample was consistent with prior epidemiologic surveys, revealing an association between higher levels of nutrient intakes and better mental health. Nutrient intakes warrant further consideration in the treatment of people with mood disorders.

The kinetics of photolysis of aqueous riboflavin solutions on UV and visible irradiation has been studied in the pH range 1-12 using a specific multicomponent spectrophotometric method for the simultaneous determination of riboflavin and its major photoproducts (formylmethylflavin, lumichrome and lumiflavin). The apparent first-order rate constants for the photodegradation reactions in the pH range have been determined. The log k-pH profiles indicate that riboflavin has maximum photostability around pH 5-6, at which the rate of oxidation-reduction of the molecule is lowest. The cationic and anionic forms of riboflavin are non-fluorescent and less susceptible to photolysis than the non-ionised molecule as indicated by the relatively slow rates below pH 3.0 and above pH 10.0. The rate of photolysis is increased up to 80-fold at pH 10.0, compared to that at pH 5.0, due to increase in redox potentials with an increase in pH and consequently the ease with which the molecule is oxidised. The increase in rate at pH 3.0, compared to that at pH 5.0, appears to be due to the involvement of the excited singlet state as well as the triplet state in riboflavin degradation. The apparent first-order rate constants for the photolysis of riboflavin at pH 5.0-10.0 with UV and visible radiation are 0.185 x 10(-2) to 13.182 x 10(-2)min(-1) and 0.098 x 10(-2) to 7.762 x 10(-2)min(-1), respectively.

Novel mechanisms of action and new chemical scaffolds are needed to rejuvenate antibacterial drug discovery, and riboswitch regulators of bacterial gene expression are a promising class of targets for the discovery of new leads. Herein, we report the characterization of 5-(3-(4-fluorophenyl)butyl)-7,8-dimethylpyrido[3,4-b]quinoxaline-1,3(2H,5H)-dione (5FDQD)-an analog of riboflavin that was designed to bind riboswitches that naturally recognize the essential coenzyme flavin mononucleotide (FMN) and regulate FMN and riboflavin homeostasis. In vitro, 5FDQD and FMN bind to and trigger the function of an FMN riboswitch with equipotent activity. MIC and time-kill studies demonstrated that 5FDQD has potent and rapidly bactericidal activity against Clostridium difficile. In C57BL/6 mice, 5FDQD completely prevented the onset of lethal antibiotic-induced C. difficile infection (CDI). Against a panel of bacteria representative of healthy bowel flora, the antibacterial selectivity of 5FDQD was superior to currently marketed CDI therapeutics, with very little activity against representative strains from the Bacteroides, Lactobacillus, Bifidobacterium, Actinomyces, and Prevotella genera. Accordingly, a single oral dose of 5FDQD caused less alteration of culturable cecal flora in mice than the comparators. Collectively, these data suggest that 5FDQD or closely related analogs could potentially provide a high rate of CDI cure with a low likelihood of infection recurrence. Future studies will seek to assess the role of FMN riboswitch binding to the mechanism of 5FDQD antibacterial action. In aggregate, our results indicate that riboswitch-binding antibacterial compounds can be discovered and optimized to exhibit activity profiles that merit preclinical and clinical development as potential antibacterial therapeutic agents.

Photodynamic collagen crosslinking (CXL) using ultraviolet-A (UVA) irradiation combined with the photosensitizer riboflavin has been introduced as a new treatment for progressive keratoconus. The results of clinical studies are promising, but the efficacy of the treatment in halting the progression depends on the stability of the induced biomechanical effects. The effects of corneal CXL on corneal rigidity; collagen fiber diameter; and resistance to heat degradation, enzymatic digestion, and swelling due to hydration are reviewed in this paper. The collective results indicate that CXL using UVA and riboflavin enhances the biomechanical properties of the corneal tissue, which remain stable over time. Therefore, this treatment could become the future standard therapy for keratoconus or used to halt the progression of keratoconus and postpone the need for corneal transplantation. The increase in availability and popularity of the CXL technique accentuates the requirement for reliable and accurate techniques for measuring corneal biomechanical properties before and after treatment.

OBJECTIVE

To assess maternal diet and nutritional adequacy prior to conception and during pregnancy.

METHODS

Follow-up of a cohort of pregnant women with collection of questionnaire data throughout pregnancy and after delivery.

METHODS

Antenatal clinics at two public hospitals in Porto, Portugal.

METHODS

Two hundred and forty-nine pregnant women who reported a gestational age below 13 weeks at the time they attended their first antenatal visit.

RESULTS

Intakes of energy and macronutrients were within recommended levels for most women. Pregnancy was accompanied by increases in the dietary intake of vitamins A and E, riboflavin, folate, Ca and Mg, but declines in the intake of alcohol and caffeine. The micronutrients with higher inadequacy prevalences prior to pregnancy were vitamin E (83%), folate (58%) and Mg (19%). These three micronutrients, together with Fe, were also those with the highest inadequacy prevalences during pregnancy (91%, 88%, 73% and 21%, respectively, for folate, Fe, vitamin E and Mg). Ninety-seven per cent of the women reported taking supplements of folic acid during the first trimester, but the median gestational age at initiation was 6.5 (interquartile range 5, 9) weeks. Self-reported prevalences of Fe and Mg supplementation were high, and increased throughout pregnancy.

CONCLUSIONS

The study identified low dietary intakes of vitamin E, folate and Mg both in the preconceptional period and during pregnancy, and low intake of Fe during pregnancy only. The low dietary intake of folate and the late initiation of supplementation indicate that current national guidelines are unlikely to be effective in preventing neural tube defects.

OBJECTIVE

To determine if usual nutrient intake is related to a 5-year change in the amount of lens nuclear opacification assessed by computer-assisted image analysis.

METHODS

A sample of 408 Boston, Mass-area women from the Nurses' Health Study aged 52 to 74 years at baseline participated in a 5-year study related to nutrition and vision. Usual nutrient intake was calculated as the average intake from 5 food frequency questionnaires that were collected over a 13- to 15-year period before the baseline evaluation of lens nuclear density. Duration of vitamin supplement use before baseline was determined from 7 questionnaires collected during this same period. We assessed the degree of nuclear density (opacification) using computer-assisted image analysis of digital lens images with amount of nuclear density measured as a function of average pixel gray scale, ranging from 0 (clear) to 255 (black).

RESULTS

Median (range) baseline and follow-up nuclear densities were 44 (19 to 102) and 63 (32 to 213). The median (range) 5-year change in nuclear density was 18 (-29 to 134) and was positively correlated with the amount of opacification at baseline (Spearman correlation coefficient = 0.35; P<.001). Geometric mean 5-year change in nuclear density was inversely associated with the intake of riboflavin (P trend = .03) and thiamin (P trend = .04) and duration of vitamin E supplement use (P trend = .006).

CONCLUSIONS

Our results suggest that long-term use of vitamin E supplements and higher riboflavin and/or thiamin intake may reduce the progression of age-related lens opacification.

Many flavoproteins are non-covalent complexes between FMN and an apoprotein. To understand better the stability of flavoproteins, we have studied the energetics of the complex between FMN and the apoflavodoxin from Anabaena PCC 7119 by a combination of site-directed mutagenesis, titration calorimetry, equilibrium binding constant determinations, and x-ray crystallography. Comparison of the strength of the wild type and mutant apoflavodoxin-FMN complexes and that of the complexes between wild type apoflavodoxin and shortened FMN analogues (riboflavin and lumiflavin) allows the dissection of the binding energy into contributions associated with the different parts of the FMN molecule. The estimated contribution of the phosphate is greatest, at 7 kcal mol(-1); that of the isoalloxazine is of around 5-6 kcal mol(-1) (mainly due to interaction with Trp-57 and Tyr-94 in the apoprotein) and the ribityl contributes least: around 1 kcal mol(-1). The stabilization of the complex is both enthalpic and entropic although the enthalpy contribution is dominant. Both the phosphate and the isoalloxazine significantly contribute to the enthalpy of binding. The ionic strength does not have a large effect on the stability of the FMN complex because, although it weakens the phosphate interactions, it strengthens the isoalloxazine-protein hydrophobic interactions. Phosphate up to 100 mM does not affect the strength of the riboflavin complex, which suggests the isoalloxazine and phosphate binding sites may be independent in terms of binding energy. Interestingly, we find crystallographic evidence of flexibility in one of the loops (57-62) involved in isoalloxazine binding.

Flavodoxin from Desulfovibrio vulgaris (Hildenborough) has been expressed at a high level (3-4% soluble protein) in Escherichia coli by subcloning a minimal insert carrying the gene behind the tac promoter of plasmid pDK6. The recombinant protein was readily isolated and its properties were shown to be identical to those of the wild-type protein obtained directly from D. vulgaris, with the exception that the recombinant protein lacks the N-terminal methionine residue. Detailed measurements of the redox potentials of this flavodoxin are reported for the first time. The redox potential, E2, for the couple oxidized flavodoxin/flavodoxin semiquinone at pH 7.0 is -143 mV (25 degrees C), while the value for the flavodoxin semiquinone/flavodoxin hydroquinone couple (E1) at the same pH is -440 mV. The effects of pH on the observed potentials were examined; E2 varies linearly with pH (slope = -59 mV), while E1 is independent of pH at high pH values, but below pH 7.5 the potential becomes less negative with decreasing pH, indicating a redox-linked protonation of the flavodoxin hydroquinone. D. vulgaris apoflavodoxin binds FMN very tightly, with a value of 0.24 nM for the dissociation constant (Kd) at pH 7.0 and 25 degrees C, similar to that observed with other flavodoxins. In addition, the apoflavodoxin readily binds riboflavin (Kd = 0.72 microM; 50 mM sodium phosphate, pH 7.0, 5 mM EDTA at 25 degrees C) and the complex is spectroscopically very similar to that formed with FMN. The redox potentials for the riboflavin complex were determined at pH 6.5 (E1 = -262 mV, E2 = -193 mV; 25 degrees C) and are discussed in the light of earlier proposals that charge/charge interactions between different parts of the flavin hydroquinone play a crucial role in determining E1 in flavodoxin.

Hemidesmus indicus R. Br. (Asclepiadaceae) is a well known drug in Ayurveda system of medicine. In the present study, antioxidant activity of methanolic extract of H. indicus root bark was evaluated in several in vitro and ex vivo models. Further, preliminary phytochemical analysis and TLC fingerprint profile of the extract was established to characterize the extract which showed antioxidant properties. The in vitro and ex vivo antioxidant potential of root bark of H. indicus was evaluated in different systems viz. radical scavenging activity by DPPH reduction, superoxide radical scavenging activity in riboflavin/light/NBT system, nitric oxide (NO) radical scavenging activity in sodium nitroprusside/Greiss reagent system and inhibition of lipid peroxidation induced by iron-ADP-ascorbate in liver homogenate and phenylhydrazine induced haemolysis in erythrocyte membrane stabilization study. The extract was found to have different levels of antioxidant properties in the models tested. In scavenging DPPH and superoxide radicals, its activity was intense (EC50 = 18.87 and 19.9 microg/ml respectively) while in scavenging NO radical, it was moderate. It also inhibited lipid peroxidation of liver homogenate (EC50 = 43.8 microg/ml) and the haemolysis induced by phenylhydrazine (EC50 = 9.74 microg/ml) confirming the membrane stabilization activity. The free radical scavenging property may be one of the mechanisms by which this drug is effective in several free radical mediated disease conditions.

The effect of flavin deficiency on blue light responses in Neurospora crassa was studied through the use of two riboflavin mutants (rib-1 and rib-2). The photoresponses assayed were the suppression of circadian conidiation, the phase shifting of the circadian conidiation rhythm, and the induction of carotenoid synthesis. Flavin deficiency was induced in the rib-1 mutant by restrictive growth temperatures and in the rib-2 mutant by low levels of supplemental riboflavin. At 26 degrees C, a semirestrictive growth temperature, the rib-1 mutant is about 1/80th as sensitive to light for the photosuppression of circadian conidiation. Flavin deficiency in the rib-1 and rib-2 strains was effective in reducing the photosensitivity for phase shifting and carotenogenesis to about 1/16th and 1/4th of normal, respectively. Experiments with permissive temperatures, riboflavin supplementation, and revertants at the rib locus all indicated that the effects on light sensitivity were due solely to the presence of the rib mutations. These results provide evidence that one or more flavin photoreceptors are involved in the blue light responses of Neurospora.

OBJECTIVE

Photodynamic collagen cross-linking by using ultraviolet A (UVA) irradiation and the photosensitizer riboflavin has been recently introduced as a new possible treatment of progressive keratoconus. This is the first study, to our knowledge, investigating biochemical aspects of the new procedure. Its aim was to analyze the possible changes in the electrophoretic pattern of corneal collagen type I after collagen cross-linking treatment.

METHODS

Twenty fresh postmortem porcine corneas were cross-linked; another 20 porcine corneas treated with physiologic saline were used as controls. After removal of the central 10 mm of the epithelium, the corneas were treated with the photosensitizer riboflavin and UVA irradiation for 30 minutes by using a double UVA diode (370 nm, 3 mW/cm). For biochemical analysis, the central 10-mm corneal buttons were trephined, tissue was homogenized, and collagen type I was extracted. Subsequently, the collagen extracts were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

RESULTS

In the controls, the typical collagen pattern of normal cornea was found with 1 gamma trimer band, 2 beta dimer bands, and 2 alpha monomer bands. In the cross-linked samples, there was an additional intense polymer band in the stacking gel that was resistant to mercaptoethanol, heat, and pepsin treatment. Its molecular size was estimated to be at least 1000 kDa.

CONCLUSIONS

In the cross-linked corneas, a strong band of high-molecular-weight collagen polymers was shown as the biochemical correlate of the cross-linking effect, showing the efficiency of the new cross-linking procedure. This polymer band complies well with the morphologic correlate of an increased fiber diameter after cross-linking treatment. Its chemical stability supports hopes of a long-term effect of the new treatment.

OBJECTIVE

To examine if and how ready-to-eat cereals (RTEC) contribute to the quality of the diet of children, adolescents and young adults in a Mediterranean setting.

METHODS

A random sample of 3534 subjects aged 2 to 24 years in Spain was studied. Food and nutrient intakes were determined by a 24 hour recall. RTEC consumption was assessed by a quantitative food frequency questionnaire. Additional questions on socioeconomic level and nutritional knowledge were administered. Cereal consumption was classified into non-consumers and daily intakes between 1 and 20g, 21 and 40g, and more than 40g. After excluding the underreporters the final sample consisted of 2852 individuals.

RESULTS

About half of the population (49.8%) reported eating RTEC. Macronutrient profile improved with increasing cereal consumption. Intakes of thiamine, riboflavin and vitamin B6 increased significantly with increasing consumption of RTEC in all age-sex groups, whereas niacin and folate intake improved in almost all groups and calcium, iron and vitamin D in at least half of the groups. Except for magnesium, vitamin B12 and vitamin E in males, consumption of RTEC was significantly associated with increased coverage of the daily nutrient requirements for all micronutrients studied. Higher levels of RTEC consumption was associated with a greater consumption of dairy products, and related to better breakfast quality.

CONCLUSIONS

Level of RTEC consumption is associated with a better nutritional profile in the diets of Spanish children, adolescents and young adults and a lower risk for inadequate micronutrient intakes. RTEC consumers have better quality breakfasts, in terms of both food choices as well as energy and nutrient content.

Ethylmalonic encephalopathy (EE) is a rare, recently defined inborn error of metabolism which affects the brain, gastrointestinal system and peripheral blood vessels and is characterized by a unique constellation of clinical and biochemical features. A 7-month-old male, who presented with psychomotor retardation, chronic diarrhea and relapsing petechiae is described with the objective of highlighting the biochemical and neuroradiological features of this disorder as well as the effect of high-dose riboflavin therapy. Urinary organic acid analysis revealed markedly increased excretion of ethylmalonic acid, isobutyrylglycine, 2-methylbutyrylglycine and isovalerylglycine. Acylcarnitine analysis in dried blood spots showed increased butyrylcarnitine. Short-chain acyl-CoA dehydrogenase (SCAD) activity in muscle was normal as were mitochondrial OXPHOS enzyme activities in cultured skin fibroblasts. In skeletal muscle the catalytic activity of complex II was decreased. Brain MRI revealed bilateral and symmetrical atrophy in the fronto-temporal areas, massive enlargement of the subarachnoid spaces and hyperdensities on T (2) sequences of the basal ganglia. Mutation analysis of the ETHE1 gene demonstrated homozygosity for the Arg163Gly mutation, confirming the diagnosis of EE at a molecular level. On repeat MRI, a significant deterioration was seen, correlating well with the clinical deterioration of the patient.

Mutants of the electron-transfer protein flavodoxin from Desulfovibrio vulgaris were made by site-directed mutagenesis to investigate the role of glycine-61 in stabilizing the semiquinone of FMN by the protein and in controlling the flavin redox potentials. The spectroscopic properties, oxidation-reduction potentials, and flavin-binding properties of the mutant proteins, G61A/N/V and L, were compared with those of wild-type flavodoxin. The affinities of all of the mutant apoproteins for FMN and riboflavin were less than that of the wild-type apoprotein, and the redox potentials of the two 1-electron steps in the reduction of the complex with FMN were also affected by the mutations. Values for the dissociation constants of the complexes of the apoprotein with the semiquinone and hydroquinone forms of FMN were calculated from the redox potentials and the dissociation constant of the oxidized complex and used to derive the free energies of binding of the FMN in its three oxidation states. These showed that the semiquinone is destabilized in all of the mutants, and that the extent of destabilization tends to increase with increasing bulkiness of the side chain at residue 61. It is concluded that the hydrogen bond between the carbonyl of glycine-61 and N(5)H of FMN semiquinone in wild-type flavodoxin is either absent or severely impaired in the mutants. X-ray crystal structure analysis of the oxidized forms of the four mutant proteins shows that the protein loop that contains residue 61 is moved away from the flavin by 5-6 A. The hydrogen bond formed between the backbone nitrogen of aspartate-62 and O(4) of the dimethylisoalloxazine of the flavin in wild-type flavodoxin is absent in the mutants. Reliable structural information was not obtained for the reduced forms of the mutant proteins, but if the mutants change conformation when the flavin is reduced to the semiquinone, to facilitate hydrogen bonding between N(5)H and the carbonyl of residue 61, then the change must be different from that known to occur in wild-type flavodoxin.

OBJECTIVE

To evaluate associations with biochemical indicators of nutritional and other risk factors in the Lens Opacities Case-Control Study.

METHODS

Case-control study.

METHODS

The Lens Opacities Case-Control Study determined risk factors for cortical, nuclear, and posterior subcapsular opacities among 1380 participants aged 40 to 79 years.

METHODS

Vitamin E, selenium, and biochemistry profile determinations were performed on all patients; red blood cell enzymes and amino acids were measured in systematic samples of about 25% of the Lens Opacities Case-Control Study population.

RESULTS

Laboratory test values in cases and controls were compared and expressed as odds ratios and 95% confidence intervals.

RESULTS

In polychotomous logistic regression analyses controlling for age and sex, the risk of opacities was reduced to less than one half in persons with higher levels of vitamin E (odds ratio, 0.44 for nuclear opacities), albumin-globulin ratio (odds ratio, 0.41 for mixed opacities), or iron (odds ratio, 0.43 for cortical opacities); higher uric acid levels increased risk (odds ratio, 1.74 for mixed opacities). Persons with opacities were twice as likely to have high glutathione reductase activity (with flavin adenine dinucleotide), which suggests low riboflavin status (odds ratio, 2.13). Most odds ratios for amino acids were under unity and were significantly decreased for glycine (0.36) and aspartic acid (0.31).

CONCLUSIONS

Lens opacities were associated with lower levels of riboflavin, vitamin E, iron, and protein nutritional status. Higher levels of uric acid increased risk of mixed opacities. The findings for riboflavin, vitamin E, iron, and uric acid are compatible with the dietary intake and medical history results of the Lens Opacities Case-Control Study.

Migraine prophylaxis is a stepwise procedure with lifestyle advice followed by consideration of medications. Patients should be advised to try to maintain a regular lifestyle, with regular sleep, meals, exercise, and management of stress, perhaps through relaxation techniques or other ways that are sensible for them. If this regimen does not adequately control their migraines, preventatives are indicated. Patients can choose between evidence-based nutraceuticals such as riboflavin, feverfew, butterbur, or coenzyme Q10, or more traditional pharmacotherapeutics. Medicine choices are somewhat limited by what is available in each country, but from the full range, the medicines of first choice are beta-adrenoceptor blockers, flunarizine, topiramate, and valproic acid. Beta-adrenoceptor blockers are particularly useful in patients also suffering from hypertension or tachycardia. Following recent studies, topiramate has become a first choice for episodic as well as chronic migraine. It is the only prophylactic drug that may lead to weight loss, but it is sometimes associated with adverse cognitive effects. Valproic acid and flunarizine also have very good prophylactic properties. However, valproic acid is often associated with adverse effects, and flunarizine is unavailable in many countries, including the United States. If sequential monotherapies are ineffective, combinations of first-line drugs should be tried before advancing to drugs of second choice, which are associated with more adverse effects or have less well-established prophylactic properties. Amitriptyline should be used carefully because of its anticholinergic effects, although it is useful in comorbid tension-type headache, depression, and sleep disorders. Methysergide is very effective, but it has been supplanted or even made unavailable in many countries because of its well-described association with retroperitoneal fibrosis. Pizotifen has a slightly better safety profile but is unavailable in the United States. Aspirin is particularly useful in patients needing platelet inhibitors for other medical conditions, but the risk of gastrointestinal bleeding must be considered. The prophylactic properties of magnesium, riboflavin, and coenzyme Q10 are low at best, but their lack of severe adverse effects makes them good treatment options. Magnesium may be particularly useful during pregnancy. Lisinopril and candesartan were shown to be effective in single trials and are preferable in patients with hypertension. Acupuncture may be another alternative; although controlled trials have failed to differentiate its effect from placebo, it is at least innocuous. Botulinum toxin A is not effective in the prophylaxis of episodic migraine.

OBJECTIVE

To determine mean intake of energy and protein, total fat, saturated fat, percent energy from total and saturated fat, cholesterol, carbohydrate, calcium, iron, zinc, folate, vitamins A, C, E, B-6 and B-12, thiamin, niacin, riboflavin, magnesium, sodium and fiber of preschool Head Start children at school and away from school.

METHODS

Twenty-four-hour food intakes for 358 Head Start children were obtained by observing food intake at school and acquiring intake recalls from parents or guardians specifying food their children consumed for the balance of the day. After determining group estimates of energy and nutrient intake, mean intake was compared to standard nutrient recommendations for the entire 24-hour day, i.e., for the time the children were in school and for the remaining hours away from school ("home" intake).

METHODS

The 358 Head Start children attended school either half-day (2- to 3-hour AM and PM sessions) or all-day (5 to 6 hours).

METHODS

Differences in nutrient intake among class times were analyzed using one-way analysis of variance (ANOVA) followed by Tukey's multiple comparison test. Differences with a p-value <0.05 (two-tailed) were considered to be statistically significant. Total energy, protein, calcium, iron, zinc, vitamins A, C, E, B6, and B12, thiamin, niacin, riboflavin as well as folate and magnesium were compared to the Recommended Dietary Allowances for the 4- to 6-year-old age group. Other standards that were used for comparisons included the National Cholesterol Education Program (fat, saturated fat and cholesterol), the 1989 National Research Council's Diet and Health Report (carbohydrate and sodium) and the recommendation for fiber proposed by the American Health Foundation.

RESULTS

At school, half-day children consumed up to 25% of the daily recommendation for energy and nutrients, while all-day children achieved at least a third of the recommended intakes. When intakes at home and school were combined, all three groups of children (AM, PM and all-day) exceeded dietary recommendations for protein, vitamins and minerals. Energy intake remained below 100% of the recommendation, while intake of total fat, saturated fat and cholesterol exceeded recommendations.

CONCLUSIONS

Further research is required to explore energy needs and determine nutritional status and nutrient needs of minority and low-income preschool children. Strategies are required to increase nutrient density, but not fat density, of meals and snacks served to children who attend day care for part of the day. Finally, school meals and nutrition education programs such as Team Nutrition should broaden their base to include healthful eating habits for all school children, including the very youngest children in preschool programs.

A mildly elevated homocysteine (Hcy) level is a novel and potentially modifiable risk factor for age-related osteoporotic fractures. Elevated Hcy levels can have a nutritional cause, such as inadequate intake of folate, riboflavin, pyridoxine or cobalamin, which serve as cofactors or substrates for the enzymes involved in the Hcy metabolism. We examined the association between intake of Hcy-related B vitamin (riboflavin, pyridoxine, folate and cobalamin) and femoral neck bone mineral density BMD (FN-BMD) and the risk of fracture in a large population-based cohort of elderly Caucasians. We studied 5304 individuals aged 55 years and over from the Rotterdam Study. Dietary intake of nutrients was obtained from food frequency questionnaires. Incident non-vertebral fractures were recorded during a mean follow-up period of 7.4 years, and vertebral fractures were assessed by X-rays during a mean follow-up period of 6.4 years. We observed a small but significant positive association between dietary pyridoxine (beta = 0.09, p = 1 x 10(-8)) and riboflavin intake (beta = 0.06, p = 0.002) and baseline FN-BMD. In addition, after controlling for gender, age and BMI, pyridoxine intake was inversely correlated to fracture risk. As compared to the three lowest quartiles, individuals in the highest quartile of age- and energy-adjusted dietary pyridoxine intake had a decreased risk of non-vertebral fractures (HR = 0.77, 95% CI = 0.65-0.92, p = 0.005) and of fragility fractures (HR = 0.55, 95% CI = 0.40-0.77, p = 0.0004). Further adjustments for other dietary B vitamins (riboflavin, folate and cobalamin), dietary intake of calcium, vitamin D, vitamin A and vitamin K, protein and energy intake, smoking and BMD did not essentially modify these results. We conclude that increased dietary riboflavin and pyridoxine intake was associated with higher FN-BMD. Furthermore, we found a reduction in risk of fracture in relation to dietary pyridoxine intake independent of BMD. These findings highlight the importance of considering nutritional factors in epidemiological studies of osteoporosis and fractures.