OBJECTIVE

To assess the influence of smoking on serum parathyroid hormone (PTH), serum vitamin D metabolites, serum ionized calcium, serum phosphate, and biochemical markers of bone turnover in a cohort of 510 healthy Danish perimenopausal women.

METHODS

A cross-sectional study.

METHODS

Copenhagen, Denmark.

METHODS

Five-hundred-and-ten healthy women aged 45-58 y, included 3-24 months after last menstrual bleeding. None were using hormone replacement therapy.

METHODS

The women were grouped according to their current smoking status. The two groups were compared with regard to serum levels of 25-hydroxyvitamin D (25OHD) and 1, 25-dihydroxyvitamin D (1,25-(OH)2D), intact PTH, ionized calcium and phosphate, osteocalcin, as well as urine pyridinolines. Bone mineral density (BMD) was measured with DEXA-scans. Multiple regression analyses were performed to detect the effect of potentially confounding lifestyle factors, such as calcium and vitamin D intakes, alcohol and coffee consumption, sunbathing, and physical exercise.

RESULTS

Fifty percent were current smokers. Smokers had significantly reduced levels of serum 25OHD (P=0.02), 1,25(OH)2D (P=0.001), and PTH (P<0.001). There was no difference in serum ionized calcium between smokers and non-smokers. We found a negative effect of smoking on serum osteocalcin (P=0.01), while urinary pyridinolines were similar in the two groups. The small differences in lifestyle between the two groups could not explain these findings. Smokers had small but significant reductions in bone mineral density.

CONCLUSIONS

Smoking has a significant effect on calcium and vitamin D metabolism, which is not likely to be explained by other confounding lifestyle factors. The depression of the vitamin D-PTH system seen among smokers may represent another potential mechanism for the deleterious effects of smoking on the skeleton, and may contribute to the reported risk of osteoporosis among smokers.

BACKGROUND

Grants from the Karen Elise Jensens Foundation.

The main regulator of the human tumor suppresser gene p21(waf1/cip1) is the transcription factor p53, but more recently it has been suggested to be a primary anti-proliferative target for the nuclear receptor VDR in the presence of its ligand 1alpha,25-dihydroxyvitamin D3 (1alpha,25(OH)2D3). To identify VDR responding regions, we analyzed 20 overlapping regions covering the first 7.1 kb of the p21(waf1/cip1) promoter in MCF-7 human breast cancer cells using chromatin immuno-precipitation assays (ChIP) with antibodies against p53 and VDR. We confirmed two known p53 binding regions at approximate positions -1400 and -2300 and identified a novel site at position -4500. In addition, we found three VDR-associated promoter regions at positions -2300, -4500 and -6900, i.e. two regions showed binding for both p53 and VDR. In silico screening and in vitro binding assays using recombinant and in vitro translated proteins identified five p53 binding sites within the three p53-positive promoter regions and also five 1alpha,25(OH)2D3 response elements within the three VDR-positive regions. Reporter gene assays confirmed the expected responsiveness of the respective promoter regions to the p53 inducer 5-fluorouracil and 1alpha,25(OH)2D3. Moreover, re-ChIP assays confirmed the functionality of the three 1alpha,25(OH)2D3-reponsive promoter regions by monitoring simultaneous occupancy of VDR with the co-activator proteins CBP, SRC-1 and TRAP220. Taken together, we demonstrated that the human p21((waf1/cip1)) gene is a primary 1alpha,25(OH)2D3-responding gene with at least three VDR binding promoter regions, in two of which also p53 co-localizes.

The active form of vitamin D, 1,25-dihydroxyvitamin D(3)[1,25(OH)(2)D(3)], is a secosteroid hormone that regulates calcium and bone metabolism, controls cell proliferation and differentiation, and exerts immunoregulatory activities. This range of functions has been exploited clinically to treat a variety of conditions, from secondary hyperparathyroidism to osteoporosis, to autoimmune diseases such as psoriasis. Recent advances in understanding 1,25(OH)(2)D(3) functions and novel insights into the mechanisms of its immunomodulatory properties suggest a wider applicability of this hormone in the treatment of autoimmune diseases and allograft rejection.

Nonhomologous end joining (NHEJ) is the major DNA double-strand break (DSB) repair pathway in mammalian cells. A critical step in this process is DNA ligation, involving the Xrcc4-DNA ligase IV complex. DNA end processing is often a prerequisite for ligation, but the coordination of these events is poorly understood. We show that polynucleotide kinase (PNK), with its ability to process ionizing radiation-induced 5'-OH and 3'-phosphate DNA termini, functions in NHEJ via an FHA-dependent interaction with CK2-phosphorylated Xrcc4. Analysis of the PNK FHA-Xrcc4 interaction revealed that the PNK FHA domain binds phosphopeptides with a unique selectivity among FHA domains. Disruption of the Xrcc4-PNK interaction in vivo is associated with increased radiosensitivity and slower repair kinetics of DSBs, in conjunction with a diminished efficiency of DNA end joining in vitro. Therefore, these results suggest a new role for Xrcc4 in the coordination of DNA end processing with DNA ligation.

In metazoans, certain calmodulin-related calcium-binding proteins (recoverins, neurocalcins and frequenins) are found at highest levels in excitable cells, but their physiological roles are largely uncharacterized. Here we show that Saccharomyces cerevisiae contains a frequenin homologue, Frq1, and that its target is Pik1, a phosphatidylinositol-4-OH kinase. Frq1 binds to a conserved sequence motif in Pik1 outside Pik1's catalytic domain and stimulates its activity in vitro. N-myristoylated Frq1 may also assist in Pik1 localization.

The tumor suppressor p53 is commonly inhibited under conditions in which the phosphatidylinositide 3'-OH kinase/protein kinase B (PKB)Akt pathway is activated. Intracellular levels of p53 are controlled by the E3 ubiquitin ligase Mdm2. Here we show that PKB inhibits Mdm2 self-ubiquitination via phosphorylation of Mdm2 on Ser(166) and Ser(188). Stimulation of human embryonic kidney 293 cells with insulin-like growth factor-1 increased Mdm2 phosphorylation on Ser(166) and Ser(188) in a phosphatidylinositide 3'-OH kinase-dependent manner, and the treatment of both human embryonic kidney 293 and COS-1 cells with phosphatidylinositide 3'-OH kinase inhibitor LY-294002 led to proteasome-mediated Mdm2 degradation. Introduction of a constitutively active form of PKB together with Mdm2 into cells induced phosphorylation of Mdm2 at Ser(166) and Ser(188) and stabilized Mdm2 protein. Moreover, mouse embryonic fibroblasts lacking PKBalpha displayed reduced Mdm2 protein levels with a concomitant increase of p53 and p21(Cip1), resulting in strongly elevated apoptosis after UV irradiation. In addition, activation of PKB correlated with Mdm2 phosphorylation and stability in a variety of human tumor cells. These findings suggest that PKB plays a critical role in controlling of the Mdm2.p53 signaling pathway by regulating Mdm2 stability.

Vitamin D functions in the body through both an endocrine mechanism (regulation of calcium absorption) and an autocrine mechanism (facilitation of gene expression). The former acts through circulating calcitriol, whereas the latter, which accounts for more than 80% of the metabolic utilization of the vitamin each day, produces, uses, and degrades calcitriol exclusively intracellularly. In patients with end-stage kidney disease, the endocrine mechanism is effectively disabled; however, the autocrine mechanism is able to function normally so long as the patient has adequate serum levels of 25(OH)D, on which its function is absolutely dependent. For this reason, calcitriol and its analogs do not constitute adequate replacement in managing vitamin D needs of such patients. Optimal serum 25(OH)D levels are greater than 32 ng/mL (80 nmol/L). The consequences of low 25(OH)D status include increased risk of various chronic diseases, ranging from hypertension to diabetes to cancer. The safest and most economical way to ensure adequate vitamin D status is to use oral dosing of native vitamin D. (Both daily and intermittent regimens work well.) Serum 25(OH)D can be expected to rise by about 1 ng/mL (2.5 nmol/L) for every 100 IU of additional vitamin D each day. Recent data indicate that cholecalciferol (vitamin D(3)) is substantially more potent than ergocalciferol (vitamin D(2)) and that the safe upper intake level for vitamin D(3) is 10,000 IU/d.

Targeting neuroendocrine tumors expressing somatostatin receptor subtypes (sst) with radiolabeled somatostatin agonists is an established diagnostic and therapeutic approach in oncology. While agonists readily internalize into tumor cells, permitting accumulation of radioactivity, radiolabeled antagonists do not, and they have not been considered for tumor targeting. The macrocyclic chelator 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) was coupled to two potent somatostatin receptor-selective peptide antagonists [NH(2)-CO-c(DCys-Phe-Tyr-DAgl(8)(Me,2-naphthoyl)-Lys-Thr-Phe-Cys)-OH (sst(3)-ODN-8) and a sst(2)-selective antagonist (sst(2)-ANT)], for labeling with (111/nat)In. (111/nat)In-DOTA-sst(3)-ODN-8 and (111/nat)In-DOTA-[4-NO(2)-Phe-c(DCys-Tyr-DTrp-Lys-Thr-Cys)-DTyr-NH(2)] ((111/nat)In-DOTA-sst(2)-ANT) showed high sst(3)- and sst(2)-binding affinity, respectively. They did not trigger sst(3) or sst(2) internalization but prevented agonist-stimulated internalization. (111)In-DOTA-sst(3)-ODN-8 and (111)In-DOTA-sst(2)-ANT were injected intravenously into mice bearing sst(3)- and sst(2)-expressing tumors, and their biodistribution was monitored. In the sst(3)-expressing tumors, strong accumulation of (111)In-DOTA-sst(3)-ODN-8 was observed, peaking at 1 h with 60% injected radioactivity per gram of tissue and remaining at a high level for >72 h. Excess of sst(3)-ODN-8 blocked uptake. As a control, the potent agonist (111)In-DOTA-[1-Nal(3)]-octreotide, with strong sst(3)-binding and internalization properties showed a much lower and shorter-lasting uptake in sst(3)-expressing tumors. Similarly, (111)In-DOTA-sst(2)-ANT was injected into mice bearing sst(2)-expressing tumors. Tumor uptake was considerably higher than with the highly potent sst(2)-selective agonist (111)In-diethylenetriaminepentaacetic acid-[Tyr(3),Thr(8)]-octreotide ((111)In-DTPA-TATE). Scatchard plots showed that antagonists labeled many more sites than agonists. Somatostatin antagonist radiotracers therefore are preferable over agonists for the in vivo targeting of sst(3)- or sst(2)-expressing tumors. Antagonist radioligands for other peptide receptors need to be evaluated in nuclear oncology as a result of this paradigm shift.

A number of studies have suggested that Vitamin D has a potential role in the development/treatment of diabetes. These effects may be mediated by circulating levels of 1alpha,25(OH)(2)D(3), but local production of 1alpha,25(OH)(2)D(3), catalysed by the enzyme 25-hydroxyvitamin D(3)-1alpha-hydroxylase (1alpha-OHase), is also likely to be important. RT-PCR analyses demonstrated that both isolated rat islets and MIN6 cells (mouse insulin-secreting cell line, characteristic of beta cells) expressed 1alpha-OHase mRNA. The transcript in both cell types was similar to that seen in HKC-8 cells (a renal cell line, which expresses 1alpha-OHase). Western blot analysis and immunolocalisation identified 1alpha-OHase protein in MIN6 cells and human pancreatic tissue. In addition, suspensions of rat islets were able to convert [3H]-25-hydroxyvitamin D(3) to [3H]-1alpha,25(OH)(2)D(3), demonstrating 1alpha-OHase activity. Both cell systems expressed the Vitamin D receptor and 1alpha,25(OH)(2)D(3) (50nM) evoked a rapid rise in [Ca(2+)](i) in MIN6 cells. This data clearly demonstrates islets are able to produce 1alpha,25(OH)(2)D(3) and respond rapidly to treatment with 1alpha,25(OH)(2)D(3). Therefore, we would postulate that local production of 1alpha,25(OH)(2)D(3) maybe an important autocrine link between Vitamin D status and pancreatic function.

OBJECTIVE

To assess the vitamin D status in homebound, community-dwelling elderly persons; sunlight-deprived elderly nursing home residents; and healthy, ambulatory elderly persons.

METHODS

A cohort analytic study.

METHODS

Of 244 subjects at least 65 years old, 116 subjects (85 women and 31 men) had been confined indoors for at least 6 months, either in private dwellings in the community (the Hopkins Elder Housecall Program) or in a teaching nursing home (The Johns Hopkins Geriatrics Center). The 128 control subjects, a healthy ambulatory group, came from the Baltimore Longitudinal Study on Aging. All subjects were free of diseases or medications that might interfere with their vitamin D status.

METHODS

Serum levels of 25-hydroxyvitamin D (25-OHD) and 1,25-dihydroxyvitamin D (1,25-[OH]2D) were measured in all subjects. In a subgroup of 80 subjects, serum levels of intact parathyroid hormone (PTH), ionized calcium, and osteocalcin and intake of vitamin D (through 3-day food records) were assessed. A randomly selected cohort of sunlight-deprived subjects also had serum levels of vitamin D binding protein measured.

RESULTS

In sunlight-deprived subjects overall, the mean 25-OHD level was 30 nmol/L (12 ng/mL) (range, < 10 to 77 nmol/L [< 4 to 31 ng/mL]) and the mean 1,25-(OH)2D level was 52 pmol/L (20 pg/mL) (range, 18 to 122 pmol/L [7 to 47 pg/mL]). In the sunlight-deprived subjects, 54% of community dwellers and 38% of nursing home residents had serum levels of 25-OHD below 25 nmol/L (10 ng/mL) (normal range, 25 to 137 nmol/L [10 to 55 ng/mL]). A significant inverse relationship existed between 25-OHD (ie, Log [25-OHD]) and PTH when they were analyzed together (r = -0.42; R2 = 0.18; P < .001) and for each cohort separately. All other parameters measured, except ionized calcium, differed significantly from the Baltimore Longitudinal Study Group means. The mean (SD) daily intakes of vitamin D (121 [132] IU) and calcium (583 [322] mg) were below the recommended dietary allowance only in the community-dwelling homebound population. The mean vitamin D binding protein level in the sunlight-deprived subgroup was in the normal range.

CONCLUSIONS

Despite a relatively high degree of vitamin supplementation in the United States, homebound elderly persons are likely to suffer from vitamin D deficiency.

Control of thermoregulatory effectors by the autonomic nervous system is a critical component of rapid cold-defense responses, which are triggered by thermal information from the skin. However, the central autonomic mechanism driving thermoregulatory effector responses to skin thermal signals remains to be determined. Here, we examined the involvement of several autonomic brain regions in sympathetic thermogenic responses in brown adipose tissue (BAT) to skin cooling in urethane-chloralose-anesthetized rats by monitoring thermogenic [BAT sympathetic nerve activity (SNA) and BAT temperature], metabolic (expired CO(2)), and cardiovascular (arterial pressure and heart rate) parameters. Acute skin cooling, which did not reduce either rectal (core) or brain temperature, evoked increases in BAT SNA, BAT temperature, expired CO(2), and heart rate. Skin cooling-evoked thermogenic, metabolic, and heart rate responses were inhibited by bilateral microinjections of bicuculline (GABA(A) receptor antagonist) into the preoptic area (POA), by bilateral microinjections of muscimol (GABA(A) receptor agonist) into the dorsomedial hypothalamic nucleus (DMH), or by microinjection of muscimol, glycine, 8-OH-DPAT (5-HT(1A) receptor agonist), or kynurenate (nonselective antagonist for ionotropic excitatory amino acid receptors) into the rostral raphe pallidus nucleus (rRPa) but not by bilateral muscimol injections into the lateral/dorsolateral part or ventrolateral part of the caudal periaqueductal gray. These results implicate the POA, DMH, and rRPa in the central efferent pathways for thermogenic, metabolic, and cardiac responses to skin cooling, and suggest that these pathways can be modulated by serotonergic inputs to the medullary raphe.

OBJECTIVE

To examine cross-sectional associations of serum vitamin D [25-hydroxyvitamin D, 25(OH)D] concentration with insulin resistance (IR) and beta-cell dysfunction in 712 subjects at risk for type 2 diabetes.

METHODS

Serum 25(OH)D was determined using a chemiluminescence immunoassay. Insulin sensitivity/resistance were measured using the Matsuda insulin sensitivity index for oral glucose tolerance tests (IS(OGTT)) and homeostasis model assessment of insulin resistance HOMA-IR. beta-Cell function was determined using both the insulinogenic index (IGI) divided by HOMA-IR (IGI/IR) and the insulin secretion sensitivity index-2 (ISSI-2). RESULTS Linear regression analyses indicated independent associations of 25(OH)D with IS(OGTT) and HOMA-IR (beta = 0.004, P = 0.0003, and beta = -0.003, P = 0.0072, respectively) and with IGI/IR and ISSI-2 (beta = 0.004, P = 0.0286, and beta = 0.003, P = 0.0011, respectively) after adjusting for sociodemographics, physical activity, supplement use, parathyroid hormone, and BMI.

CONCLUSIONS

Vitamin D may play a role in the pathogenesis of type 2 diabetes, as 25(OH)D concentration was independently associated with both insulin sensitivity and beta-cell function among individuals at risk of type 2 diabetes.

Interleukin 15 (IL-15) promotes the survival of natural killer (NK) cells by preventing apoptosis through mechanisms unknown at present. Here we identify Bim, Noxa and Mcl-1 as key regulators of IL-15-dependent survival of NK cells. IL-15 suppressed apoptosis by limiting Bim expression through the kinases Erk1 and Erk2 and mechanisms dependent on the transcription factor Foxo3a, while promoting expression of Mcl-1, which was necessary and sufficient for the survival of NK cells. Withdrawal of IL-15 led to upregulation of Bim and, accordingly, both Bim-deficient and Foxo3a-/- NK cells were resistant to cytokine deprivation. Finally, IL-15-mediated inactivation of Foxo3a and cell survival were dependent on phosphotidylinositol-3-OH kinase. Thus, IL-15 regulates the survival of NK cells at multiple steps, with Bim and Noxa being key antagonists of Mcl-1, the critical survivor factor in this process.

Endocytosis is required for internalization of micronutrients and turnover of membrane components. Endophilin has been assigned as a component of clathrin-mediated endocytosis. Here we show in mammalian cells that endophilin marks and controls a fast-acting tubulovesicular endocytic pathway that is independent of AP2 and clathrin, activated upon ligand binding to cargo receptors, inhibited by inhibitors of dynamin, Rac, phosphatidylinositol-3-OH kinase, PAK1 and actin polymerization, and activated upon Cdc42 inhibition. This pathway is prominent at the leading edges of cells where phosphatidylinositol-3,4-bisphosphate-produced by the dephosphorylation of phosphatidylinositol-3,4,5-triphosphate by SHIP1 and SHIP2-recruits lamellipodin, which in turn engages endophilin. This pathway mediates the ligand-triggered uptake of several G-protein-coupled receptors such as α2a- and β1-adrenergic, dopaminergic D3 and D4 receptors and muscarinic acetylcholine receptor 4, the receptor tyrosine kinases EGFR, HGFR, VEGFR, PDGFR, NGFR and IGF1R, as well as interleukin-2 receptor. We call this new endocytic route fast endophilin-mediated endocytosis (FEME).

DNA or 2-deoxyguanosine reacts with hydroxyl free radical to form 8-hydroxy-deoxyguanosine (8-OH-dG). We found that 8-OH-dG can be effectively separated from deoxyguanosine by high pressure liquid chromatography and very sensitively detected using electrochemical detection. The sensitivity of electrochemical detection is about one-thousand fold enhanced over optical detection. Utilizing deoxyguanosine in bicarbonate buffer it was found that ferrous ion, but not ferric ion, was effective in forming 8-OH-dG. The hydroxyl free radical scavenging agents, thiourea and ethanol, were very effective in quenching Fe(11) mediated 8-OH-dG formation, but superoxide dismutase had very little effect.

We examined the cross-sectional association between plasma 25-hydroxyvitamin D [25(OH)D] and markers of the insulin resistant phenotype. Plasma 25(OH)D concentrations were measured in 808 nondiabetic participants of the Framingham Offspring Study. Outcome measures included fasting and 2-h post 75-g oral glucose tolerance test (OGTT) glucose and insulin; these were used to calculate the homeostatic model assessment-insulin resistance (HOMA-IR) and insulin sensitivity index (ISI(0,120)). We also measured plasma adiponectin, triacylglycerol, and HDL cholesterol concentrations as markers of the insulin-resistant phenotype. After adjusting for age, sex, BMI, waist circumference, and current smoking status, plasma 25(OH)D concentration was inversely associated with fasting plasma glucose and insulin concentrations, and HOMA-IR. Compared with the participants in the lowest tertile category of plasma 25(OH)D, those in the highest tertile category had a 1.6% lower concentration of fasting plasma glucose (P-trend = 0.007), 9.8% lower concentration of fasting plasma insulin (P-trend = 0.001), and 12.7% lower HOMA-IR score (P-trend < 0.001). After adjusting for age and sex, plasma 25(OH)D was positively associated with ISI(0,120), plasma adiponectin, and HDL cholesterol and inversely associated with plasma triacylglycerol, but these associations were no longer significant after further adjustment for BMI, waist circumference, and current smoking status. 25(OH)D and 2-h post-OGTT glucose were not associated. Among adults without diabetes, vitamin D status was inversely associated with surrogate fasting measures of insulin resistance. These results suggest that vitamin D status may be an important determinant for type 2 diabetes mellitus.

Viruses must enter host cells to replicate, assemble and propagate. Because of the restricted size of their genomes, viruses have had to evolve efficient ways of exploiting host cell processes to promote their own life cycles and also to escape host immune defence mechanisms. Many viral open reading frames (viORFs) with immune-modulating functions essential for productive viral growth have been identified across a range of viral classes. However, there has been no comprehensive study to identify the host factors with which these viORFs interact for a global perspective of viral perturbation strategies. Here we show that different viral perturbation patterns of the host molecular defence network can be deduced from a mass-spectrometry-based host-factor survey in a defined human cellular system by using 70 innate immune-modulating viORFs from 30 viral species. The 579 host proteins targeted by the viORFs mapped to an unexpectedly large number of signalling pathways and cellular processes, suggesting yet unknown mechanisms of antiviral immunity. We further experimentally verified the targets heterogeneous nuclear ribonucleoprotein U, phosphatidylinositol-3-OH kinase, the WNK (with-no-lysine) kinase family and USP19 (ubiquitin-specific peptidase 19) as vulnerable nodes in the host cellular defence system. Evaluation of the impact of viral immune modulators on the host molecular network revealed perturbation strategies used by individual viruses and by viral classes. Our data are also valuable for the design of broad and specific antiviral therapies.

The RabA4b GTPase labels a novel, trans-Golgi network compartment displaying a developmentally regulated polar distribution in growing Arabidopsis thaliana root hair cells. GTP bound RabA4b selectively recruits the plant phosphatidylinositol 4-OH kinase, PI-4Kbeta1, but not members of other PI-4K families. PI-4Kbeta1 colocalizes with RabA4b on tip-localized membranes in growing root hairs, and mutant plants in which both the PI-4Kbeta1 and -4Kbeta2 genes are disrupted display aberrant root hair morphologies. PI-4Kbeta1 interacts with RabA4b through a novel homology domain, specific to eukaryotic type IIIbeta PI-4Ks, and PI-4Kbeta1 also interacts with a Ca2+ sensor, AtCBL1, through its NH2 terminus. We propose that RabA4b recruitment of PI-4Kbeta1 results in Ca2+-dependent generation of PI-4P on this compartment, providing a link between Ca2+ and PI-4,5P2-dependent signals during the polarized secretion of cell wall components in tip-growing root hair cells.

The recent emergence of the novel pathogenic SARS-coronavirus 2 (SARS-CoV-2) is responsible for a global pandemic. In face of the health emergency, drug repositioning is the most reliable option to design an efficient therapy for infected patients without delay. The first step of the viral replication cycle, i.e. the attachment to the surface of respiratory cells mediated by the spike (S) viral protein, offers several potential therapeutic targets. The S protein uses the ACE-2 receptor for entry, but also sialic acids linked to host cell surface gangliosides. Using a combination of structural and molecular modeling approaches, we showed that chloroquine (CLQ), one of the drugs currently under investigation for SARS-CoV-2 treatment, binds sialic acids and gangliosides with high affinity. We identified a new type of ganglioside-binding domain at the tip of the N-terminal domain of the SARS-CoV-2 spike (S) protein. This domain (aa 111-158), which is fully conserved among clinical isolates worldwide, may improve the attachment of the virus to lipid rafts and facilitate the contact with the ACE-2 receptor. We showed that in presence of CLQ (or of the more active derivative hydroxychloroquine, CLQ-OH), the viral spike is no longer able to bind gangliosides. The identification of this new mechanism of action of CLQ and CLQ-OH supports the use of these repositioned drugs to cure SARS-CoV-2 infected patients and stop the pandemic. Our in silico approaches might also be used to assess the efficiency of a broad range of repositioned and/or innovative drug candidates before their clinical evaluation.

Fibroblast growth factor-23 (FGF23) is a phosphaturic hormone that contributes to several hypophosphatemic disorders by reducing the expression of the type II sodium-phosphate cotransporters (NaPi-2a and NaPi-2c) in the kidney proximal tubule and by reducing serum 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] levels. The FGF receptor(s) mediating the hypophosphatemic action of FGF23 in vivo have remained elusive. In this study, we show that proximal tubules express FGFR1, -3, and -4 but not FGFR2 mRNA. To determine which of these three FGFRs mediates FGF23's hypophosphatemic actions, we characterized phosphate homeostasis in FGFR3(-/-) and FGFR4(-/-) null mice, and in conditional FGFR1(-/-) mice, with targeted deletion of FGFR1 expression in the metanephric mesenchyme. Basal serum phosphorus levels and renal cortical brush-border membrane (BBM) NaPi-2a and NaPi-2c expression were comparable between FGFR1(-/-), FGFR3(-/-), and FGFR4(-/-) mice and their wild-type counterparts. Administration of FGF23 to FGFR3(-/-) mice induced hypophosphatemia in these mice (8.0 +/- 0.4 vs. 5.4 +/- 0.3 mg/dl; p < or = 0.001) and a decrease in renal BBM NaPi-2a and NaPi-2c protein expression. Similarly, in FGFR4(-/-) mice, administration of FGF23 caused a small but significant decrease in serum phosphorus levels (8.7 +/- 0.3 vs. 7.6 +/- 0.4 mg/dl; p < or = 0.001) and in renal BBM NaPi-2a and NaPi-2c protein abundance. In contrast, injection of FGF23 into FGFR1(-/-) mice had no effects on serum phosphorus levels (5.6 +/- 0.3 vs. 5.2 +/- 0.5 mg/dl) or BBM NaPi-2a and NaPi-2c expression. These data show that FGFR1 is the predominant receptor for the hypophosphatemic action of FGF23 in vivo, with FGFR4 likely playing a minor role.

The active metabolite of vitamin D, 1α, 25-dihydroxyvitamin D3 [1,25(OH)(2) D3], is involved in calcium and phosphate metabolism and exerts a large number of biological effects. Vitamin D3 inhibits parathyroid hormone secretion, adaptive immunity and cell proliferation, and at the same time promotes insulin secretion, innate immunity and stimulates cellular differentiation. The role of vitamin D3 in immunoregulation has led to the concept of a dual function as both as an important secosteroid hormone for the regulation of body calcium homeostasis and as an essential organic compound that has been shown to have a crucial effect on the immune responses. Altered levels of vitamin D3 have been associated, by recent observational studies, with a higher susceptibility of immune-mediated disorders and inflammatory diseases. This review reports the new developments with specific reference to the metabolic and signalling mechanisms associated with the complex immune-regulatory effects of vitamin D3 on immune cells.

Stretching of cardiac muscle modulates contraction through the enhancement of the Ca2+ transient, but how this occurs is still not known. We found that stretching of myocytes modulates the elementary Ca2+ release process from ryanodine-receptor Ca2+-release channels (RyRCs), Ca2+ sparks and the electrically stimulated Ca2+ transient. Stretching induces PtdIns-3-OH kinase (PI(3)K)-dependent phosphorylation of both Akt and the endothelial isoform of nitric oxide synthase (NOS), nitric oxide (NO) production, and a proportionate increase in Ca2+-spark frequency that is abolished by inhibiting NOS and PI(3)K. Exogenously generated NO reversibly increases Ca2+-spark frequency without cell stretching. We propose that myocyte NO produced by activation of the PI(3)K-Akt-endothelial NOS axis acts as a second messenger of stretch by enhancing RyRC activity, contributing to myocardial contractile activation.

It is unclear whether optimal levels of 25-hydroxyvitamin D (25(OH)D) in whites are the same as in minorities. In adult participants of NHANES, the relationships between 25(OH)D, bone mineral density (BMD), and parathyroid hormone (PTH) differed in blacks as compared to whites and Mexican-Americans, suggesting that optimal 25(OH)D levels for bone and mineral metabolism may differ by race.

BACKGROUND

Blacks and Hispanics have lower 25-hydroxyvitamin D concentrations than whites. However, it is unclear whether 25(OH)D levels considered "optimal" for bone and mineral metabolism in whites are the same as those in minority populations.

METHODS

We examined the relationships between 25(OH)D and parathyroid hormone in 8,415 adult participants (25% black and 24% Mexican-American) in the National Health and Nutrition Examination Surveys 2003-2004 and 2005-2006; and between 25(OH)D and bone mineral density in 4,206 adult participants (24% black and 24% Mexican-American) in the 2003-2004 sample.

RESULTS

Blacks and Mexican-Americans had significantly lower 25(OH)D and higher PTH concentrations than whites (P < 0.01 for both). BMD significantly decreased (P < 0.01) as serum 25(OH)D and calcium intake declined among whites and Mexican-Americans, but not among blacks (P = 0.2). The impact of vitamin D deficiency (25(OH)D ≤ 20 ng/ml) on PTH levels was modified by race/ethnicity (P for interaction, 0.001). Whereas inverse relationships between 25(OH)D and PTH were observed above and below a 25(OH)D level of 20 ng/ml in whites and Mexican-Americans, an inverse association between 25(OH)D and PTH was only observed below this threshold in blacks, with the slope of the relationship being essentially flat (P = 0.7) above this cut-point, suggesting that PTH may be maximally suppressed at lower 25(OH)D levels in blacks than in whites or Mexican-Americans.

CONCLUSIONS

The relationships between 25(OH)D, BMD, and PTH may differ by race among US adults. Whether race-specific ranges of optimal vitamin D are needed to appropriately evaluate the adequacy of vitamin D stores in minorities requires further study.

The abilities of 15 flavonoids as a scavenger of active oxygens (hydroxyl radical and superoxide anion) were studied. Hydroxyl radical (.OH) was generated by the Fenton system, and assayed by the determination of methanesulfonic acid (MSA) formed from the reaction of dimethyl sulfoxide (DMSO) with .OH. (+)-Catechin, (-)-epicatechin, 7,8-dihydroxy flavone, and rutin showed the .OH scavenging effect 100-300 times superior to that of mannitol, a typical .OH scavenger. The other flavonoids showed no .OH scavenging effect at their concentrations up to 50 microM. Baicalein, quercetin, morin, and myricetin unexpectedly increased the .OH production in the Fenton system. The flavonoids tested now, except monohydroxy flavones, were more or less inhibitive to the superoxide anion (O2) generation in the hypoxanthine-xanthine oxidase system. A great part of this inhibitory effect was likely owing to suppression of xanthine oxidase activity by the flavonoids. The flavonoids, which scavenged .OH or O2-, were necessarily antioxidants to the peroxidation of methyl linoleate. However, there was a type of flavonoid such as morin, which have neither .OH nor O2- scavenging effect, but was a strong antioxidant.