The renin-angiotensin system (RAS) plays a central role in the regulation of blood pressure, electrolyte, and volume homeostasis. Epidemiological and clinical studies have long suggested an association of inadequate sunlight exposure or low serum 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] levels with high blood pressure and/or high plasma renin activity, but the mechanism is not understood. Our recent discovery that 1,25(OH)(2)D(3) functions as a potent negative endocrine regulator of renin gene expression provides some insights into the mechanism. The concept of vitamin D regulation of blood pressure through the RAS opens a new avenue to our understanding of the physiological functions of the vitamin D endocrine system, and provides a basis for exploring the potential use of vitamin D analogues in prevention and treatment of hypertension.

BACKGROUND

Clues from the epidemiology of schizophrenia suggest that low levels of developmental vitamin D may be associated with increased risk of schizophrenia.

OBJECTIVE

To directly examine the association between neonatal vitamin D status and risk of schizophrenia.

METHODS

Individually matched case-control study drawn from a population-based cohort.

METHODS

Danish national health registers and neonatal biobank.

METHODS

A total of 424 individuals with schizophrenia and 424 controls matched for sex and date of birth.

METHODS

The concentration of 25 hydroxyvitamin D(3) (25[OH]D3) was assessed from neonatal dried blood samples using a highly sensitive liquid chromatography tandem mass spectroscopy method. Relative risks were calculated for the matched pairs when examined for quintiles of 25(OH)D3.

RESULTS

Compared with neonates in the fourth quintile (with 25[OH]D3 concentrations between 40.5 and 50.9 nmol/L), those in each of the lower 3 quintiles had a significantly increased risk of schizophrenia (2-fold elevated risk). Unexpectedly, those in the highest quintile also had a significantly increased risk of schizophrenia. Based on this analysis, the population-attributable fraction associated with neonatal vitamin D status was 44%. The relationship was not explained by a wide range of potential confounding or interacting variables.

CONCLUSIONS

Both low and high concentrations of neonatal vitamin D are associated with increased risk of schizophrenia, and it is feasible that this exposure could contribute to a sizeable proportion of cases in Denmark. In light of the substantial public health implications of this finding, there is an urgent need to further explore the effect of vitamin D status on brain development and later mental health.

PTEN, a tumor suppressor frequently inactivated in many human cancers, directly antagonizes the activity of phosphatidylinositol-3-OH kinase (PI3K) by dephosphorylating phosphoinositides. We show here that PTEN interacts directly with the NHERF1 and NHERF2 (Na+/H+ exchanger regulatory factor) homologous adaptor proteins through the PDZ motif of PTEN and the PDZ1 domain of NHERF1 or both PDZ domains of NHERF2. NHERFs were shown to interact directly with platelet-derived growth factor receptor (PDGFR), and we demonstrate the assembly of a ternary complex between PTEN, NHERFs and PDGFR. The activation of the PI3K pathway after PDGFR stimulation was prolonged in NHERF1(-/-) mouse embryonic fibroblasts as compared to wild-type cells, consistent with defective PTEN recruitment to PDGFR in the absence of NHERF1. Depletion of NHERF2 by small interfering RNA similarly increased PI3K signaling. Phenotypically, the loss of NHERF1 enhanced the PDGF-induced cytoskeletal rearrangements and chemotactic migration of the cells. These data indicate that, in normal cells, NHERF proteins recruit PTEN to PDGFR to restrict the activation of the PI3K.

To study the characteristics of molecular damage induced by ionizing radiation at the DNA level, Monte Carlo track simulation of energetic electrons and ions in liquid water, a canonical model of B-DNA, and a comprehensive classification of DNA damage in terms of the origin and complexity of damage were used to calculate the frequencies of simple and complex strand breaks. A threshold energy of 17.5 eV was used to model the damage by direct energy deposition, and a probability of 0.13 was applied to model the induction of a single-strand break produced in DNA by OH radical reactions. For preliminary estimates, base damage was assumed to be induced by the same direct energy threshold deposition or by the reaction of an OH radical with the base, with a probability of 0.8. Computational data are given on the complexity of damage, including base damage by electrons with energies of 100-4500 eV and ions with energies of 0.3-4.0 MeV/nucleon (59-9 keV microm(-1) protons and 170-55 keV microm(-1) alpha particles). Computational data are presented on the frequencies of single- and double-strand breaks induced as a function of the LET of the particles, and on the relative frequencies of complex single- and double-strand breaks for electrons. The modeling and calculations of strand breaks show that: (1) The yield of strand breaks per unit absorbed dose is nearly constant over a wide range of LET. (2) The majority of DNA damage is of a simple type, but the majority of the simple single-strand breaks are accompanied by at least one base damage. (3) For low-energy electrons, nearly 20-30% of the double-strand breaks are of a complex type by virtue of additional breaks. The proportion of this locally clustered damage increases with LET, reaching about 70% for the highest-LET alpha particles modeled, with the complexity of damage increasing further, to about 90%, when base damage is considered. (4) The extent of damage in the local hit region of the DNA duplex is mostly limited to a length of a few base pairs. (5) The frequency of base damage when no strand breaks are present in the hit segment of DNA varies between 20-40% as a function of LET for protons and alpha particles.

The influence of the serum binding protein (DBP) for vitamin D and its metabolites on the concentration of its main ligands, 25-hydroxyvitamin D(3) (25-OHD(3)) and 1,25-dihydroxyvitamin D(3) (1,25-[OH](2)D(3)) was studied. The concentration of both 1,25-(OH)(2)D(3) and DBP in normal female subjects (45+/-14 ng/liter and 333+/-58 mg/liter, mean+/-SD, respectively; n = 58) increased during the intake of estro-progestogens (69+/-27 ng/liter and 488+/-90 mg/liter, respectively; n = 29), whereas the 25-OHD(3) concentration remained unchanged. A positive correlation was found between the concentrations of 1,25-(OH)(2)D(3) and DBP in these women. At the end of pregnancy, the total concentrations of 1,25-(OH)(2)D(3) (97+/-26 ng/liter, n = 40) and DBP (616+/-84 mg/liter) are both significantly higher than in nonpregnant females and paired cord serum samples (48+/-11 ng/liter and 266+/-41 mg/liter, respectively). A marked seasonal variation of 25-OHD(3) was observed in pregnant females and their infants, whereas in the same samples the concentrations of both DBP and 1,25-(OH)(2)D(3) remained constant throughout the year. The free 1,25-(OH)(2)D(3) index, calculated as the molar ratio of this steroid and DBP, remains normal in women taking estro-progestogens, however, and this might explain their normal intestinal calcium absorption despite a high total 1,25-(OH)(2)D(3) concentration. In pregnancy the free 1,25-(OH)(2)D(3) index remains normal up to 35 wk of gestation, but during the last weeks of gestation, the free 1,25-(OH)(2)D(3) index increases in both circulations. A highly significant correlation exists between the (total and free) 25-OHD(3) and 1,25-(OH)(2)D(3) concentrations in maternal and cord serum both at 35 and 40 wk of gestation.

The National Surgical Adjuvant Breast and Bowel Project (NSABP) implemented protocol B-15 to compare 2 months of Adriamycin (doxorubicin; Adria Laboratories, Columbus, OH) and cyclophosphamide (AC) with 6 months of conventional cyclophosphamide, methotrexate, and fluorouracil (CMF) in patients with breast cancer nonresponsive to tamoxifen (TAM, T). A second aim was to determine whether AC followed in 6 months by intravenous (IV) CMF was more effective than AC without reinduction therapy. Through 3 years of follow-up, findings from 2,194 patients indicate no significant difference in disease-free survival (DFS, P = .5), distant disease-free survival (DDFS, P = .5) or survival (S, P = .8) among the three groups. Since the outcome from AC and CMF was almost identical, the issue arises concerning which regimen is more appropriate for the treatment of breast cancer patients. AC seems preferable since, following total mastectomy, AC was completed on day 63 versus day 154 for conventional CMF; patients visited health professionals three times as often for conventional CMF as for AC; women on AC received therapy on each of 4 days versus on each of 84 days for conventional CMF; and nausea-control medication was given for about 84 days to conventional CMF patients versus for about 12 days to patients on AC. The difference in the amount of alopecia between the two treatment groups was less than anticipated. While alopecia was almost universally observed following AC therapy, 71% of the CMF patients also had hair loss and, in 41%, the loss was greater than 50%. This study and NSABP B-16, which evaluates the worth of AC therapy in TAM-responsive patients, indicate the merit of 2 months of AC therapy for all positive-node breast cancer patients.

Steroid hormones are central regulators of a variety of biological processes. According to the free hormone hypothesis, steroids enter target cells by passive diffusion. However, recently we demonstrated that 25(OH) vitamin D(3) complexed to its plasma carrier, the vitamin D-binding protein, enters renal proximal tubules by receptor-mediated endocytosis. Knockout mice lacking the endocytic receptor megalin lose 25(OH) vitamin D(3) in the urine and develop bone disease. Here, we report that cubilin, a membrane-associated protein colocalizing with megalin, facilitates the endocytic process by sequestering steroid-carrier complexes on the cellular surface before megalin-mediated internalization of the cubilin-bound ligand. Dogs with an inherited disorder affecting cubilin biosynthesis exhibit abnormal vitamin D metabolism. Similarly, human patients with mutations causing cubilin dysfunction exhibit urinary excretion of 25(OH) vitamin D(3). This observation identifies spontaneous mutations in an endocytic receptor pathway affecting cellular uptake and metabolism of a steroid hormone.

OBJECTIVE

The hormone 1,25-dihydroxyvitamin D (1,25(OH)2D) promotes prostate epithelial cell differentiation in vitro and thus, several groups have hypothesized that men who systemically have lower levels of 1,25(OH)2D may be at increased risk for prostate cancer. To address this hypothesis, we evaluated the association of circulating concentrations of 1,25(OH)2D and its precursor 25-hydroxyvitamin D (25(OH)D) with subsequent risk of prostate cancer.

METHODS

Prostate cancer cases were 460 men in the Health Professionals Follow-up Study who were diagnosed through 1998 after providing a blood specimen in 1993/95. 90.2% of the cases were organ confined or had minimal extraprostatic extension. An equal number of controls who had had a screening PSA test after blood draw were individually matched to cases on age, history of a PSA test before blood draw, and time of day, season, and year of blood draw. Plasma 1,25(OH)2D and 25(OH)D concentrations were determined by radio-immunosorbant assay blindly to case-control status. Odds ratios (OR) of prostate cancer and 95% confidence intervals (CI) were estimated from conditional logistic regression models mutually adjusting for quartiles of 1,25(OH)2D and 25(OH)D concentrations and for suspected prostate cancer risk factors. Quartile cutpoints were determined separately by season of blood draw using the distributions among controls.

RESULTS

Mean concentrations of 1,25(OH)2D and 25(OH)D were slightly, but not statistically significantly (p = 0.06 and 0.20, respectively), higher in cases (34.3 +/- 7.1 pg/ml and 24.6 +/- 7.7 ng/ml, respectively) than in controls (33.5 +/- 7.1 pg/ml and 23.9 +/- 8.2 ng/ml, respectively). The OR of prostate cancer comparing men in the top to bottom quartile of 1,25(OH)2D was 1.25 (95% CI: 0.82-1.90, p-trend = 0.16). For 25(OH)D the OR of prostate cancer comparing the top and bottom quartiles was 1.19 (95% CI: 0.79-1.79, p-trend = 0.59). These findings did not vary by level of the other metabolite, age at diagnosis, family history of prostate cancer, or factors that are thought to influence 25(OH)D levels.

CONCLUSIONS

In this prospective study, we did not observe an inverse association between plasma concentrations of 1,25(OH)2D or 25(OH)D and incident prostate cancer, although we cannot rule out potential effects at later stages of the disease.

OBJECTIVE

There is a need to explore the processes of social support and self-efficacy change over time among individuals in recovery homes, and to assess the extent to which residents remain abstinent, obtain and maintain employment, refrain from criminal activity, and utilize health care systems both while within the and after leaving such settings.

METHODS

Residents were recruited and interviewed at an initial baseline phase and then re-interviewed at three subsequent 4-month intervals.

METHODS

Oxford Houses are recovery home residences for individuals with substance abuse and dependence problems who seek a supportive, democratic, mutual-help setting.

METHODS

A national US sample of Oxford House residents (n=897: 604 men, 293 women).

METHODS

Information was gathered on abstinence, social support, self-efficacy, employment, criminal history, and medical care utilization.

RESULTS

Change in cumulative abstinence was predicted by support for alcohol use, abstinence self-efficacy, and length of residency in OH (i.e., less than versus>>or=6 months), even after controlling for initial time spent in OH.

CONCLUSIONS

Results suggest that receiving abstinence support, guidance, and information from recovery home members committed to the goal of long-term sobriety may enhance residents' abstinence self-efficacy and enable persons recovering from alcohol and other drug addiction to reduce the probability of a relapse.

A nonclassical 5-hydroxytryptamine (5-HT) receptor mediates the stimulation of adenylate cyclase activity in mouse embryo colliculi neurons in primary culture. The pharmacological profile characterized with agonists and antagonists suggests that this 5-HT receptor does not appear to correspond to a known 5-HT receptor. On this 5-HT receptor, 5-HT (EC50 = 109 +/- 17 nM) and 5-methoxytryptamine (5-MeOT) were equipotent agonists. The other tryptamine derivatives, 5-carboxamidotryptamine (5-CT) and 5-methoxy-N,N-dimethyltryptamine (5-MeOT-N,N-DMT), were full potent agonists, whereas tryptamine, bufotenine, and 2-CH3-5-HT were weak partial agonists. Two selective 5-HT1A agonists: 8-hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT) and ipsapirone, could not stimulate adenylate cyclase. RU 24969, a tetrahydropyridoindole derivative that is a potent 5-HT1A and 5-HT1B agonist was also inactive, whereas RU 28253, another member of this series, could stimulate cAMP production. The action of antagonists acting on 5-HT1 or 5-HT2 receptors, such as methiothepin (5-HT1 and 5-HT2), metergoline (5-HT1 and 5-HT2), spiperone (5-HT1A and 5-HT2), (-)-pindolol (5-HT1B), mesulergine (5-HT1C), and ketanserin (5-HT2), were almost inactive in reversing the 5-HT stimulating effect. The selective 5-HT3 antagonist ICS 205 930 was a full competitive antagonist at this receptor. Nevertheless, MDL 72222, which is also a 5-HT3 antagonist, was very weak in antagonizing the 5-HT stimulatory effect. A receptor with similar characteristics has also been found in guinea pig hippocampal membranes. In these membranes, the second receptor of low affinity for 5-HT, termed RL, which is positively coupled to adenylate cyclase, was also antagonized by ICS 205 930. The relatively low affinity of this hippocampal receptor for 5-CT, its stimulation by RU 28253 but not by RU 24969, and its previously reported pharmacological characteristics support the contention that this 5-HT receptor and the 5-HT receptor of mouse embryo colliculi neurons in primary culture (both positively coupled to cAMP formation) present great homologies. Inasmuch as none of the classical specific 5-HT1 and 5-HT2 agonists or antagonists interact with these 5-HT receptors, it is unlikely that they belong to 5-HT1 or 5-HT2 receptor categories.(ABSTRACT TRUNCATED AT 400 WORDS)

The Society for Sarcopenia, Cachexia, and Wasting Disease convened an expert panel to develop nutritional recommendations for prevention and management of sarcopenia. Exercise (both resistance and aerobic) in combination with adequate protein and energy intake is the key component of the prevention and management of sarcopenia. Adequate protein supplementation alone only slows loss of muscle mass. Adequate protein intake (leucine-enriched balanced amino acids and possibly creatine) may enhance muscle strength. Low 25(OH) vitamin D levels require vitamin D replacement.

An enzyme activity introducing an alkali-labile site at 2-hydroxyadenine (2-OH-A) in double-stranded oligonucleotides was detected in nuclear extracts of Jurkat cells. This activity co-eluted with activities toward adenine paired with guanine and 8-oxo-7,8-dihydroguanine (8-oxoG) as a single peak corresponding to a 55 kDa molecular mass on gel filtration chromatography. Further co-purification was then done. Western blotting revealed that these activities also co-purified with a 52 kDa polypeptide which reacted with antibodies against human MYH (anti-hMYH). Recombinant hMYH has essentially similar activities to the partially purified enzyme. Thus, hMYH is likely to possess both adenine and 2-OH-A DNA glycosylase activities. In nuclear extracts from Jurkat cells, a 52 kDa polypeptide was detected with a small amount of 53 kDa polypeptide, while in mitochondrial extracts a 57 kDa polypeptide was detected using anti-hMYH. With amplification of the 5'-regions of the hMYH cDNA, 10 forms of hMYH transcripts were identified and subgrouped into three types, each with a unique 5' sequence. These hMYH transcripts are likely to encode multiple authentic hMYH polypeptides including the 52, 53 and 57 kDa polypeptides detected in Jurkat cells.

BACKGROUND

Indirect evidence suggests that optimal vitamin D status is achieved with a serum 25-hydroxyvitamin D [25(OH)D] concentration >75 nmol/L.

OBJECTIVE

We aimed to determine the intake of vitamin D(3) needed to raise serum 25(OH)D to >75 nmol/L.

METHODS

The design was a 6-mo, prospective, randomized, double-blinded, double-dummy, placebo-controlled study of vitamin D(3) supplementation. Serum 25(OH)D was measured by radioimmunoassay. Vitamin D(3) intake was adjusted every 2 mo by use of an algorithm based on serum 25(OH)D concentration.

RESULTS

A total of 138 subjects entered the study. After 2 dose adjustments, almost all active subjects attained concentrations of 25(OH)D >75 nmol/L, and no subjects exceeded 220 nmol/L. The mean (+/-SD) slope at 9 wk [defined as 25(OH)D change/baseline dose] was 0.66 +/- 0.35 (nmol/L)/(microg/d) and did not differ statistically between blacks and whites. The mean daily dose was 86 microg (3440 IU). The use of computer simulations to obtain the most participants within the range of 75-220 nmol/L predicted an optimal daily dose of 115 microg/d (4600 IU). No hypercalcemia or hypercalciuria was observed.

CONCLUSIONS

Determination of the intake required to attain serum 25(OH)D concentrations >75 nmol/L must consider the wide variability in the dose-response curve and basal 25(OH)D concentrations. Projection of the dose-response curves observed in this convenience sample onto the population of the third National Health and Nutrition Examination Survey suggests a dose of 95 microg/d (3800 IU) for those above a 25(OH)D threshold of 55 nmol/L and a dose of 125 microg/d (5000 IU) for those below that threshold.

8-Hydroxyguanine (8-OH-G) is one of the major DNA oxidation products implicated in mutagenesis induced by oxygen radical-forming agents, including ionizing radiation. It is also believed to be involved in spontaneous mutation induced by metabolically produced oxygen radicals. A mammalian homologue of 8-OH-G glycosylase/apurinic, apyrimidinic lyase (mutM homologue, MMH) has been identified in the EST database (for expressed sequence tags) through a homology search with yeast OGG1 protein. The human MMH protein (hMMH), 34% identical to the yeast OGG1 protein, is a member of the DNA repair protein superfamily. The hMMH gene was composed of seven exons, with the alternate last exon, exon 8, producing three major alternative splicing isoforms, because splicing of the sixth intron was optional. The hMMH protein expressed in Escherichia coli revealed the glycosylase activity and apurinic, apyrimidinic lyase activity on duplex DNA containing 8-OH-G. The hMMH protein can rescue a spontaneous mutator strain of E. coli lacking mutM and mutY. By the introduction of recombinant hMMH, the rate of mutation, the formation of rifampicin-resistant revertants, was reduced by 4-7 fold. Genomic structure analysis showed that 3' exons of the hMMH gene are transcribed on the antisense strand of the calcium-dependent calmodulin kinase 1 gene.

Amine-functionalized polyvalent oligonucleotide gold nanoparticles (DNA-Au NPs) were derivatized with a cisplatin prodrug, and the resulting DNA-Au NP conjugates were used to internalize multiple platinum centers. A platinum(IV) complex, c,c,t-[Pt(NH(3))(2)Cl(2)(OH)(O(2)CCH(2)CH(2)CO(2)H)], was tethered to the surface of DNA-Au NPs through amide linkages. The platinum-tethered gold nanoparticles were taken into several cancer cells. The drop in intracellular pH facilitated reductive release of cisplatin from the prodrug, which then formed 1,2-d(GpG) intrastrand cross-links in the cell nuclei, as confirmed by an antibody specific for this adduct. The cytotoxicity of the platinum(IV) complex increases significantly in several cancer cell lines when the complex is attached to the surface of the DNA-Au NPs and in some instances exceeds that of cisplatin.

We examined the association of bone mineral density (BMD) with a polymorphism in the gene encoding the vitamin D receptor (VDR) that causes a change in the predicted protein sequence. The polymorphism results from a C-to-T transition and creates an initiation codon (ATG) three codons proximal to a downstream start site. The polymorphism can be defined by a restriction fragment length polymorphism (RFLP) using the restriction endonuclease FokI. The presence of a FokI site, designated f, allows protein translation to initiate from the first ATG. The allele lacking the site (designated F), initiates from a second ATG site. Thus, translation products from these alleles are predicted to differ by three amino acids with the f variant elongated. In a group of 100 postmenopausal Mexican-American Caucasian women, subjects with the ff genotype (15% of the study population) had a 12.8% lower BMD at the lumbar spine than FF subjects (37% of the population) (p = 0.01). Heterozygote (Ff) subjects (48% of the population) had an intermediate BMD. This association between BMD and genotype was not apparent at the femoral neck or forearm. Over a 2-year follow-up period, a decrease in BMD at the femoral neck was greater in ff compared with FF subjects (-4.7% vs. -0.5%, p = 0.005). This trend was not apparent at the lumbar spine or forearm. There were no differences between genotype groups in measurements of 25-hydroxyvitamin D (25(OH)D), calcitriol, parathyroid hormone (PTH), osteocalcin, or urinary pyridinolines. We conclude that the FokI polymorphism of the VDR gene correlates significantly with decreased BMD at the lumbar spine and with an increased rate of bone loss at the hip in ff subjects. We emphasize that these initial data should be interpreted with caution but that the utility of this polymorphism as a genetic marker to determine BMD and osteoporosis risk warrants further study in larger populations with subjects of diverse ethnic backgrounds.

A line of mice deficient in vitamin D binding protein (DBP) was generated by targeted mutagenesis to establish a model for analysis of DBP's biological functions in vitamin D metabolism and action. On vitamin D-replete diets, DBP-/- mice had low levels of total serum vitamin D metabolites but were otherwise normal. When maintained on vitamin D-deficient diets for a brief period, the DBP-/-, but not DBP+/+, mice developed secondary hyperparathyroidism and the accompanying bone changes associated with vitamin D deficiency. DBP markedly prolonged the serum half-life of 25(OH)D and less dramatically prolonged the half-life of vitamin D by slowing its hepatic uptake and increasing the efficiency of its conversion to 25(OH)D in the liver. After an overload of vitamin D, DBP-/- mice were unexpectedly less susceptible to hypercalcemia and its toxic effects. Peak steady-state mRNA levels of the vitamin D-dependent calbindin-D9K gene were induced by 1,25(OH)2D more rapidly in the DBP-/- mice. Thus, the role of DBP is to maintain stable serum stores of vitamin D metabolites and modulate the rates of its bioavailability, activation, and end-organ responsiveness. These properties may have evolved to stabilize and maintain serum levels of vitamin D in environments with variable vitamin D availability.

BACKGROUND

Adequate 25(OH)D levels are required to prevent adverse effects on bone health. Population-based data on factors associated with 25(OH)D levels of people with MS have been lacking.

OBJECTIVE

To examine the prevalence and determinants of vitamin D insufficiency in a population-based sample of MS cases and controls, and to compare 25(OH)D status between MS cases and controls, taking into account case disability.

METHODS

We conducted a population based case-control study in Tasmania, Australia (latitude 41-43 degrees S) on 136 prevalent cases with MS confirmed by magnetic resonance imaging and 272 community controls, matched on sex and year of birth. Measurements included serum 25(OH)D, sun exposure, skin type, dietary vitamin D intake and disability including EDSS.

RESULTS

A high prevalence of vitamin D insufficiency was found in MS cases and controls. Among MS cases, increasing disability was strongly associated with lower levels of 25(<em>OH</em>)D and with reduced sun exposure. Cases with higher disability (EDSS>> 3) were more likely to have vitamin D insufficiency than controls (OR = 3.07 (1.37, 6.90) for 25(<em>OH</em>)D </= 40 nmol/l), but cases with low disability were not (OR = 0.87 (0.41, 1.86)).

CONCLUSIONS

The strong associations between disability, sun exposure and vitamin D status indicate that reduced exposure to the sun, related to higher disability, may contribute to the high prevalence of vitamin D insufficiency found in this population-based MS case sample. Active detection of vitamin D insufficiency among people with MS and intervention to restore vitamin D status to adequate levels should be considered as part of the clinical management of MS.

A trifluoromethyl ketone analogue of arachidonic acid in which the COOH group is replaced with COCF3 (AACOCF3) was prepared and found to be a tight- and slow-binding inhibitor of the 85-kDa cytosolic human phospholipase A2 (cPLA2). Enzyme inhibition was observed when AACOCF3 was tested in assays using either phospholipid vesicles or phospholipid/Triton X-100 mixed micelles. The fact that the inhibition developed over several minutes in both assays establishes that AACOCF3 inhibits by direct binding to the enzyme rather than by decreasing the fraction of enzyme bound to the substrate interface. From the measured values of the inhibitor association and dissociation rate constants, an upper limit of the equilibrium dissociation constant for the Ca(2+).AACOCF3.PLA2 complex of 5 x 10(-5) mole fraction was obtained. Thus, detectable inhibition of cPLA2 by AACOCF3 occurs when this compound is present in the assay at a level of one inhibitor per several thousand substrates. Arachidonic acid analogues in which the COOH group is replaced by COCH3, CH(OH)CF3, CHO, or CONH2 did not detectably inhibit the cPLA2. The arachidonyl ketones AACOCF2CF3 and AACOCF2Cl were found by 19F NMR to be less hydrated than AACOCF3 in phospholipid/Triton X-100 mixed micelles, and compared to AACOCF3 these compounds are also weaker inhibitors of cPLA2. In keeping with the fact that cPLA2 displays substrate specificity for arachidonyl-containing phospholipids, the arachidic acid analogue C19H39COCF3 is a considerably less potent inhibitor compared to AACOCF3.(ABSTRACT TRUNCATED AT 250 WORDS)

Based on electrophysiological, neurochemical, genetic and neuropharmacological approaches, it is currently accepted that serotonin (5-HT) functions predominantly to promote wakefulness (W) and to inhibit REM (rapid eye movement) sleep (REMS). Yet, under certain circumstances the neurotransmitter contributes to the increase in sleep propensity. Most of the serotonergic innervation of the cerebral cortex, amygdala, basal forebrain (BFB), thalamus, preoptic and hypothalamic areas, raphe nuclei, locus coeruleus and pontine reticular formation comes from the dorsal raphe nucleus (DRN). The 5-HT receptors can be classified into at least seven classes, designated 5-HT(1-7). The 5-HT(1A) and 5-HT(1B) receptor subtypes are linked to the inhibition of adenylate cyclase, and their activation evokes a membrane hyperpolarization. The actions of the 5-HT(2A), 5-HT(2B) and 5-HT(2C) receptor subtypes are mediated by the activation of phospholipase C, with a resulting depolarization of the host cell. The 5-HT(3) receptor directly activates a 5-HT-gated cation channel which leads to the depolarization of monoaminergic, aminoacidergic and cholinergic cells. The primary signal transduction pathway of 5-HT(6) and 5-HT(7) receptors is the stimulation of adenylate cyclase which results in the depolarization of the follower neurons. Mutant mice that do not express 5-HT(1A) or 5-HT(1B) receptor exhibit greater amounts of REMS than their wild-type counterparts, which could be related to the absence of a postsynaptic inhibitory effect on REM-on neurons of the laterodorsal and pedunculopontine tegmental nuclei (LDT/PPT). 5-HT(2A) and 5-HT(2C) receptor knock-out mice show a significant increase of W and a reduction of slow wave sleep (SWS) which has been ascribed to the increase of catecholaminergic neurotransmission involving mainly the noradrenergic and dopaminergic systems. Sleep variables have been characterized, in addition, in 5-HT(7) receptor knock-out mice; the mutants spend less time in REMS that their wild-type counterparts. Direct infusion of the 5-HT(1A) receptor agonists 8-OH-DPAT and flesinoxan into the DRN significantly enhances REMS in the rat. In contrast, microinjection of the 5-HT(1B) (CP-94253), 5-HT(2A/2C) (DOI), 5-HT(3) (m-chlorophenylbiguanide) and 5-HT(7) (LP-44) receptor agonists into the DRN induces a significant reduction of REMS. Systemic injection of full agonists at postsynaptic 5-HT(1A) (8-OH-DPAT, flesinoxan), 5-HT(1B) (CGS 12066B, CP-94235), 5-HT(2C) (RO 60-0175), 5-HT(2A/2C) (DOI, DOM), 5-HT(3) (m-chlorophenylbiguanide) and 5-HT(7) (LP-211) receptors increases W and reduces SWS and REMS. Of note, systemic administration of the 5-HT(2A/2C) receptor antagonists ritanserin, ketanserin, ICI-170,809 or sertindole at the beginning of the light period has been shown to induce a significant increase of SWS and a reduction of REMS in the rat. Wakefulness was also diminished in most of these studies. Similar effects have been described following the injection of the selective 5-HT(2A) receptor antagonists volinanserin and pruvanserin and of the 5-HT(2A) receptor inverse agonist nelotanserin in rodents. In addition, the effects of these compounds have been studied on the sleep electroencephalogram of subjects with normal sleep. Their administration was followed by an increase of SWS and, in most instances, a reduction of REMS. The administration of ritanserin to poor sleepers, patients with chronic primary insomnia and psychiatric patients with a generalized anxiety disorder or a mood disorder caused a significant increase in SWS. The 5-HT(2A) receptor inverse agonist APD-125 induced also an increase of SWS in patients with chronic primary insomnia. It is known that during the administration of benzodiazepine (BZD) hypnotics to patients with insomnia there is a further reduction of SWS and REMS, whereas both variables tend to remain decreased during the use of non-BZD derivatives (zolpidem, zopiclone, eszopiclone, zaleplon). Thus, the association of 5-HT(2A) antagonists or 5-HT(2A) inverse agonists with BZD and non-BZD hypnotics could be a valid alternative to normalize SWS in patients with primary or comorbid insomnia.

Electron paramagnetic resonance spectroscopy has been applied to measure radical generation in the postischemic heart; however, there is controversy regarding the methods used and the conclusion as to whether radicals are generated. In order to resolve this controversy, direct and spin trapping measurements of the time course and mechanisms of radical generation were performed in isolated perfused rabbit hearts. In reperfused tissue, 3 prominent radical signals are observed: A, isotropic g = 2.004 suggestive of a semiquinone; B, anisotropic g parallel = 2.033 and g perpendicular = 2.005 suggestive of ROO.; and C, a triplet g = 2.000 and aN = 24 G suggestive of a nitrogen centered radical. B and C, however, are highly labile and disappear at temperatures probably encountered in some previous studies. In normally perfused hearts, A is observed with only small amounts of B and C. During ischemia, B and C increase reaching a maximum after 45 min while A decreases. On reflow with oxygenated perfusate all 3 signals increase. With varying duration of ischemia and reflow, peak signal intensities occurred after 15 s of reflow following 30 min of ischemia. Reperfusion with superoxide dismutase, deferoxamine, or mannitol abolished the reperfusion increase of B. Measurements performed with the spin trap 5,5'-dimethyl-1-pyrroline-N-oxide (DMPO) demonstrated a similar time course of radical generation with prominent DMPO-OH and DMPO-R signals peaking between 10 and 20 s of reflow. Superoxide dismutase and deferoxamine also quenched these signals. Thus, .O2- derived .OH, R., and ROO. radicals are generated in postischemic myocardium. While the experimental techniques used can result in loss of intrinsic radicals and generation of extraneous radicals, with proper care and controls valid measurements of free radicals in biological tissues can be performed.

Transcription by RNA polymerase preceding the initiation of replication from the E. coli chromosomal origin (oriC) in vitro enables dnaA protein to open the DNA duplex under conditions when its action alone is insufficient. The RNA polymerases of phages T7 and T3 are as effective as the E. coli enzyme in activating initiation. The persistent RNA transcript hybridized to the template creates an R-loop that is responsible for activation. The activating RNA need not cross oriC, but must be less then 500 bp away. Transcripts lacking a 3' OH group are effective, proving that priming of DNA synthesis is not involved in the activation. Thus, transcription activates the origin of an otherwise inert plasmid by altering the local DNA structure, facilitating its opening by dnaA protein during the assembly of replication forks.

Several non-hypercalcemic analogs of 1alpha,25-dihydroxyvitamin D3 (1,25(OH)(2)D(3)) show antitumor activity in a subset of cancer patients. High vitamin D receptor (VDR) expression, which is associated with good prognosis but is lost during tumor progression. We show that the SNAIL transcription factor represses VDR gene expression in human colon cancer cells and blocks the antitumor action of EB1089, a 1,25(OH)(2)D(3) analog, in xenografted mice. In human colon cancers, elevated SNAIL expression correlates with downregulation of VDR.

Fibroblastic growth factor 23 (FGF23) regulates renal phosphate reabsorption and 1alpha-hydroxylase activity. Ablation of FGF23 results in elevated serum phosphate, calcium, and 1,25-dihydroxyvitamin D3 [1,25(OH)(2)D] levels; vascular calcifications; and early death. For determination of the independent roles of hyperphosphatemia and excess vitamin D activity on the observed phenotypic abnormalities, FGF23 null mice were fed a phosphate- or vitamin D-deficient diet. The phosphate-deficient diet corrected the hyperphosphatemia, prevented vascular calcifications, and rescued the lethal phenotype in FGF23 null mice, despite persistent elevations of serum 1,25(OH)(2)D and calcium levels. This suggests that hyperphosphatemia, rather than excessive vitamin D activity, is the major stimulus for vascular calcifications and contributes to the increased mortality in the FGF23-null mouse model. In contrast, the vitamin D-deficient diet failed to correct either the hyperphosphatemia or the vascular calcifications in FGF23 null mice, indicating that FGF23 independently regulates renal phosphate excretion and that elevations in 1,25(OH)(2)D and calcium are not sufficient to induce vascular calcifications in the absence of hyperphosphatemia. The vitamin D-deficient diet also improved survival in FGF23 null mice in association with normalization of 1,25(OH)(2)D and calcium levels and despite persistent hyperphosphatemia and vascular calcifications, indicating that excessive vitamin D activity can also have adverse effects in the presence of hyperphosphatemia and absence of FGF23. Understanding the independent and context-dependent interactions between hyperphosphatemia and excessive vitamin D activity, as well as vascular calcifications and mortality in FGF23 null mice, may ultimately provide important insights into the management of clinical disorders of hyperphosphatemia and excess vitamin D activity.