Thiocyanate supplementation decreases atherosclerotic plaque in mice expressing human myeloperoxidase.

Elevated levels of the heme enzyme myeloperoxidase (MPO) are associated with adverse cardiovascular outcomes. MPO predominantly catalyzes formation of the oxidants hypochlorous acid (HOCl) from Cl(-), and hypothiocyanous acid (HOSCN) from SCN(-), with these anions acting as competitive substrates. HOSCN is a less powerful and more specific oxidant than HOCl, and selectively targets thiols; such damage is largely reversible, unlike much HOCl-induced damage. We hypothesized that increased plasma SCN(-), and hence HOSCN formation instead of HOCl, may decrease artery wall damage. This was examined using high-fat fed atherosclerosis-prone LDLR(-/-) mice transgenic for human MPO, with and without SCN(-) (10 mM) added to drinking water. Serum samples, collected fortnightly, were analyzed for cholesterol, triglycerides, thiols, MPO, and SCN(-); study-long exposure was calculated by area under the curve (AUC). Mean serum SCN(-) concentrations were elevated in the supplemented mice (200-320 µM) relative to controls (< 120 µM). Normalized aortic root plaque areas at sacrifice were 26% lower in the SCN(-)-supplemented mice compared with controls (P = 0.0417), but plaque morphology was not appreciably altered. Serum MPO levels steadily increased in mice on the high-fat diet, however, comparison of SCN(-)-supplemented versus control mice showed no significant changes in MPO protein, cholesterol, or triglyceride levels; thiol levels were decreased in supplemented mice at one time-point. Plaque areas increased with higher cholesterol AUC (r = 0.4742; P = 0.0468), and decreased with increasing SCN(-) AUC (r = - 0.5693; P = 0.0134). These data suggest that increased serum SCN(-) levels, which can be achieved in humans by dietary manipulation, may decrease atherosclerosis burden.

A mutation in a case of early onset narcolepsy and a generalized absence of hypocretin peptides in human narcoleptic brains.

We explored the role of hypocretins in human narcolepsy through histopathology of six narcolepsy brains and mutation screening of Hcrt, Hcrtr1 and Hcrtr2 in 74 patients of various human leukocyte antigen and family history status. One Hcrt mutation, impairing peptide trafficking and processing, was found in a single case with early onset narcolepsy. In situ hybridization of the perifornical area and peptide radioimmunoassays indicated global loss of hypocretins, without gliosis or signs of inflammation in all human cases examined. Although hypocretin loci do not contribute significantly to genetic predisposition, most cases of human narcolepsy are associated with a deficient hypocretin system.

Enamel defects in primary and permanent teeth of children born prematurely.

The objective of this study was to determine the prevalence of enamel defects in both primary and permanent dentitions of the same preterm children, and to elucidate the role of early dietary mineral and vitamin D intake in the etiology of the enamel defects. The status of the primary and permanent teeth was evaluated in 32 preterm children and in 64 control children. The prevalence of enamel defects in children born preterm was clearly higher as compared with controls in both the primary (78% vs 20%, P<0.001) and permanent (83% vs 36%, P<0.001) dentitions. Neither the mineral supplementation used nor a vitamin D dose of 1000 IU/day, as compared with a lower dose of 500 IU/day, reduced the prevalence of enamel defects in the primary or permanent dentitions. Further studies are needed to clarify whether achieving near optimum intra-uterine mineral retention would lower the prevalence of subsequent enamel defects in infants born prematurely.

Maturation of primary and permanent teeth in preterm infants.

AIMS: To elucidate the development of primary and permanent teeth and to interpret the effect of different calcium, phosphorus, and vitamin D supplementation in the neonatal period on dental maturation in preterm children. METHODS: Preterm infants were randomised to four groups to receive a vitamin D dose of 500 or 1000 IU/day and calcium and phosphorus supplemented or unsupplemented breast milk. The maturity of the primary and permanent teeth was recorded in 30 preterm children. Sixty children aged 2 years and 60 children aged 9-11 years served as controls. Bone mineral content/density was assessed in the preterm infants. RESULTS: The median (range) corrected teething age was 7 (2-16) months in preterm infants and 6 (2-12) months in controls (p = 0.43). The median (range) number of erupted teeth at 2 years of age was 16 (11-19) in preterm infants and 16 (12-20) in controls (p = 0.16). Maturation of the permanent teeth in the preterm infants was not delayed compared with the controls (mean Demirjian SDS 0.16 v 0.49, p = 0.14). Early dietary intake of either mineral or vitamin D did not affect maturation of the primary dentition in preterm children. Children receiving the higher vitamin D dose in the neonatal period had more mature permanent dentition than those receiving the lower dose, but mineral intake did not affect maturation of the permanent teeth. Dental maturation did not correlate with bone mineral status. CONCLUSIONS: This is the first longitudinal study to follow primary and permanent tooth maturation in the same preterm children. Premature birth has no appreciable late sequelae in tooth maturation.

The long-term effect of early mineral, vitamin D, and breast milk intake on bone mineral status in 9- to 11-year-old children born prematurely.

BACKGROUND: Although the short-term benefits of mineral supplementation in preterm infants has been established, the long-term benefits are less clear. The purpose of the study was to evaluate effects of early-life mineral, vitamin D, and breast milk intake on bone mineral status in children 9 to 11 years of age who were born prematurely. METHODS: Seventy preterm infants born 1985 through 1987 were randomized into four groups: to receive a vitamin D dose of 500 or 1000 IU/day and calcium- and phosphorus-supplemented or unsupplemented breast milk. At 3 months of age, radial bone mineral content was determined by single-photon absorptiometry and vitamin D metabolites were assessed. At 9 to 11 years of age, the bone mineral status of the radius and lumbar spine was assessed using dual energy x-ray absorptiometry. RESULTS: At the age of 3 months, the preterm infants with diets supplemented with minerals had 36% higher bone mineral content than the preterm infants whose diet was not supplemented with minerals. At the age of 9 to 11 years, in contrast, bone mineral status was comparable among the groups, irrespective of different mineral supplementation during the neonatal period. Interestingly, the lumbar bone mineral apparent density was positively related to lactation in mineral-supplemented children. There was neither short-term nor long-term benefit to bone mineral status of a vitamin D dose of 1000 IU/day compared with 500 IU/day. CONCLUSIONS: The short-term benefit to bone mineral density in preterm infants of mineral supplementation of the early diet is obvious, but, in the long term, the effects seem to disappear. The results also imply that a relatively long period of breast-feeding may be needed to optimize long-term bone mineral acquisition in the lumbar spine.

Nephroblastoma overexpressed gene (NOV) codes for a growth factor that induces protein tyrosine phosphorylation.

NOV (nephroblastoma overexpressed gene) is a member of the CCN (connective tissue growth factor [CTGF], Cyr61/Cef10, NOV) family of proteins. These proteins are cysteine-rich and are noted for having growth-regulatory functions. We have isolated the rat NOV gene, and the DNA sequence shares 90% identity with the mouse and 80% identity with the human sequences. The rat NOV gene was expressed in all rat tissues examined, including brain, lung, heart, kidney, liver, spleen, thymus and skeletal muscle. Higher levels of rat NOV mRNA were seen in the brain, lung and skeletal muscle compared to the other tissues. Examination of NOV expression in various human cell lines revealed that NOV was expressed in U87, 293, T98G, SK-N-MC and Hs683 but not in HepG2, HL60, THP1 and Jurkat. The human NOV gene was transfected into 293 cells and the expressed protein purified. When 3T3 fibroblasts were treated with this recombinant NOV protein, a dose-dependent increase in proliferation was observed. Analysis of tyrosine-phosphorylated proteins revealed that when 3T3 cells were treated with NOV, a 221 kDa protein was phosphorylated. These data suggest that NOV can act as a growth factor for some cells and binds to a specific receptor that leads to the phosphorylation of a 221 kDa protein.

Expression profile of the copper homeostasis gene, rAtox1, in the rat brain.

In humans the regulation of cellular copper homeostasis is essential for proper organ development and function. A novel cytosolic protein, named Atox 1, was recently identified in yeast that functions in shuttling intracellular mononuclear copper [Cu(I)] to copper-requiring proteins. Atox 1 and its human homolog, hAtox1, are members of an emerging family of proteins termed copper chaperones that are involved in the maintenance of copper homeostasis. Northern blot analysis demonstrates that Atox 1 is widely expressed at varying levels in a variety of rat tissues including brain. Using in situ hybridization histochemistry, we characterized the expression profile for the rat homolog of Atox1 (rAtox1) in the normal adult rat brain. There is widespread expression within the brain that appears to be primarily neuronal. The highest levels of Atox1 message consists of distinct neuronal subtypes that are also characterized by their high levels of metals like copper, iron, and zinc, which include the pyramidal neurons of the cerebral cortex and hippocampus in addition to the neurons of the locus coeruleus. The high levels of a metal chaperone like Atox1 in subsets of neurons that also sequester metals suggests that Atox1 may be important in maintaining the functionality of metal requiring enzymes. A detailed analysis of the restricted expression profile for a novel copper chaperone, rAtox1, is described in the adult rat CNS. Further analysis shows that Atoxl expression is associated with neuronal populations that sequester copper.

Randomised controlled trial of vitamin D supplementation on bone density and biochemical indices in preterm infants.

AIMS: To test the hypothesis that a vitamin D dose of 200 IU/kg, maximum 400 IU/day, given to preterm infants will maintain normal vitamin D status and will result in as high a bone mineral density as that attained with the recommended dose of 960 IU/day. METHODS: Thirty nine infants of fewer than 33 weeks of gestational age were randomly allocated to receive vitamin D 200 IU/kg of body weight/day up to a maximum of 400 IU/day or 960 IU/day until 3 months old. Vitamin D metabolites, bone mineral content and density were determined by dual energy x-ray absorptiometry, and plasma ionised calcium, plasma alkaline phosphatase, and intact parahormone measurements were used to evaluate outcomes. RESULTS: The 25 hydroxy vitamin D concentrations tended to be higher in infants receiving 960 IU/day, but the differences did not reach significance at any age. There was no difference between the infants receiving low or high vitamin D dose in bone mineral content nor in bone mineral density at 3 and 6 months corrected age, even after taking potential risk factors into account. CONCLUSIONS: A vitamin D dose of 200 IU/kg of body weight/day up to a maximum of 400 IU/day maintains normal vitamin D status and as good a bone mineral accretion as the previously recommended higher dose of 960 IU/day. Vitamin D is a potent hormone which affects organs other than bone and should not be given in excess to preterm infants.

Expression and characterization of a putative high affinity human soluble leptin receptor.

Leptin, a circulating 16-kDa protein secreted by adipocytes, decreases body weight by reducing food intake and enhancing energy utilization. Leptin receptors that share homology to the glycoprotein gp130 have been recently cloned. In addition, differentially spliced leptin receptor messenger RNAs have been identified. Functional mutations in either the leptin or leptin receptor gene cause obesity. In the present study, expression of the full length human leptin receptor complementary DNA encoding the long cytoplasmic domain of leptin receptor in COS7 cells resulted in high affinity membrane binding of 125I-leptin (Ki approximately 200 pM); no detectable binding was present in the medium. In addition, we expressed the extracellular domain of human leptin receptor in COS7 cells and identified a soluble leptin receptor in the conditioned medium that binds human and mouse leptin with high affinity comparable with the full length membrane receptor. Transfected COS7 cells expressing the soluble leptin receptor also demonstrated modest specific 125I-leptin binding in whole cells, presumably due to association of the soluble leptin receptor to cell membrane proteins. Data from cross-linking studies identified two specific bands in the 125I-leptin/soluble leptin receptor complex with molecular masses of approximately 130-150 kDa and 300 kDa. The 130-150 kDa molecular mass was confirmed in Western blot analysis and Coomassie staining of the purified soluble receptor and probably represents the glycosylated form of the receptor. The 300-kDa band most likely represents a homodimer of the soluble leptin receptor complex because HPLC gel filtration analysis of the 125I-leptin/soluble leptin receptor complex identified a single peak corresponding to a molecular mass of approximately 340 kDa. The soluble leptin receptor antagonized 125I-leptin binding to the membrane receptor, suggesting its potential utility as a functional tool for determining the role of endogenous leptin.

Ophthalmic side effects of hyperbaric oxygen therapy.

Hyperbaric oxygen (HBO) treatments are documented to cause ocular side effects. According to Palmquist et al., HBO therapy has been used for many years, yet there are only a few reports of its effects on the eye. With current studies reporting lens changes, as well as hyperopic and myopic fluctuations, the role of the nurse in assessing and reporting vision changes needs to be defined and clarified.

Metabolic bone disease of prematurity.

Metabolic bone disease is recognized with increasing frequency in very-low-birth-weight infants. Radiological changes characteristic of rickets have been found in 55% of infants with a birth weight of less than 1000 g and in 23% of infants weighing less than 1500 g at birth. Twenty-four per cent of infants with a birth weight of less than 1500 g have fractures. The main aetiological factor is insufficient phosphorus supplementation. The aetiology is, however, multifactorial and also includes calcium deficiency, vitamin D deficiency, certain drugs, aluminium loading and immobilisation. The method of choice in detecting subclinical mineral bone disease of prematurity is measurement of bone mineral density, but there is as yet no single good diagnostic method available for premature infants. The optimal mineral and vitamin D requirement of the premature infant must be established so that proper recommendations can be given. The current recommended vitamin D dose in Europe (ESPGAN 800-1000 IU/day) is probably too high when extra minerals are supplied. Moreover, the duration of mineral supplementation may need to be continued until the infant has reached a body weight of 3.5 kg. This article deals with the aetiology, pathogenesis, diagnosis and future prospects of metabolic bone disease of prematurity.

Serum vitamin A levels in mothers and their breast-fed term infants with or without supplemental vitamin A.

Serum concentrations of vitamin A were measured in term infants (n = 72) and their mothers at delivery and after 20 weeks of breast-feeding (n = 48). During the 20 weeks the infants received either no supplemental vitamin A (but the mothers were given 3,000 IU vitamin A daily) (n = 16) or a daily vitamin A supplementation of 600 (n = 17) or 1,500 IU (n = 15). After 20 weeks of breast-feeding the vitamin A levels in the unsupplemented infants were similar to those at birth. The infants supplemented either with 600 or 1,500 IU had higher vitamin A serum levels than at birth (p less than 0.01), however, there was no difference between the two supplemented groups. During lactation, the serum vitamin A concentrations of the mothers increased significantly in all groups with or without vitamin A supplementation.