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Animal models suggest a key role for dihydroxylated vitamin D metabolites in fracture healing, as evidenced by increases in serum concentration of 24R,25-dihydroxyvitamin D (24R,25[OH]2D) after long bone fracture. Human studies investigating the kinetics of serum concentrations of 24R,25[OH]2D, 1,25-dihydroxyvitamin D (1,25[OH]2D) and their parent metabolite 25-hydroxyvitamin D (25[OH]D) are lacking. We, therefore, conducted a longitudinal study to determine whether total, free, or bioavailable concentrations of these vitamin D metabolites fluctuate in humans after long bone fracture. Twenty-eight patients with cross-shaft (diaphyseal) long bone fracture presenting to an emergency department in London, UK, were studied. Serum concentrations of 25(OH)D, 24R,25(OH)2D, 1,25(OH)2D, vitamin D binding protein, albumin, and calcium were determined within 48 hours of fracture and again at 1 and 6 weeks postfracture. Concentrations of free and bioavailable vitamin D metabolites were calculated using standard equations. No changes in mean serum concentrations of 25(OH)D or 24R,25(OH)2D were seen at either follow-up time point versus baseline. In contrast, mean serum 1,25(OH)2D concentration declined by 21% over the course of the study, from 68.5 pmol/L at baseline to 54.1 pmol/L at 6 weeks (p

It is largely through historical accident in the interval of 1920–1940 that vitamin D3 became classified as a vitamin rather than as a steroid hormone. The formal definition of a vitamin is that it is a trace dietary constituent required to produce the normal function of a physiological process or processes. The emphasis here is on trace and the fact that the vitamin must be supplied regularly in the diet; this implies that the body is unable to metabolically synthesize the vitamin in question. However, the ultraviolet exposure of 7-dehydrocholesterol present in the skin results in the photochemical production of vitamin D3. Thus, vitamin D3 becomes a true vitamin only when the animal or human does not have regular access to sunlight or ultraviolet light. Under normal physiological circumstances, all mammals, including humans, can generate, via ultraviolet exposure of 7-dehydrocholesterol present in the skin, adequate quantities of vitamin D3 to meet their nutritionally defined requirements. There is a vibrant historical record beginning in 1650 and culminating in 1963 concerned with the determination of the chemical structures of vitamin D3 and vitamin D2. A surprising aspect concerning vitamin D3 is that it is itself biologically inert. There are no known essential biological actions or contributions that rely specifically on the molecule vitamin D3. While chemists had certainly appreciated the strong structural similarity between the vitamins D and other steroids, this correlation was never widely acknowledged in the biological, clinical, or nutritional sciences until 1965–1970. The biological role of vitamin D3 is to serve as a substrate for the liver 25-hydroxylase which produces 25-hydroxyvitamin D3 [25(OH)D3]. 25(OH)D3 in turn serves as the substrate for the kidney proximal tubule 25(OH)D3-1α-hydroxylase enzyme which produces the steroid hormone 1α,25(OH)2-vitamin D3 [1α,25(OH)2D3].

Steroids exert their actions through several pathways. The classical genomic pathway, which involves binding of steroids to receptors and subsequent modulation of gene expression, is well characterized. Besides this, rapid actions of steroids have been shown to exist. Since 30 years, research on rapid actions of steroids is an emerging field of science. Today, rapid effects of steroids are well established, and are shown to exist for every type of steroid. The classical steroid receptors have been shown to be involved in rapid actions, but there is also strong evidence that unrelated structures mediate these rapid effects. Despite increasing knowledge about the mechanisms and structures which mediate these actions, there is still no unanimous acceptance of this category. This article briefly reviews the history of the field including current controversies and challenges. It is not meant as a broad review of literature, but should increase the awareness of the endocrinology society for rapid responses to steroids. As members of the organizing committee of the VI International Meeting on Rapid Responses to Steroid Hormones 2009, we propose a research agenda focusing on the identification of new receptoral structures and the identification of mechanisms of actions at physiological steroid concentrations. Additionally, efforts for the propagation of translational studies, which should finally lead to clinical benefit in the area of rapid steroid action research, should be intensified.

Sertoli cell secretory activities are highly dependent on ion channel functions and critical to spermatogenesis. The steroid hormone 1alpha,25(OH)2-vitamin D3 (1,25(OH)2-D3) stimulates exocytosis in different cell systems by activating a nongenotropic vitamin D receptor (VDR). Here, we described 1,25(OH)2-D3 stimulation of secretion via Cl(-) channel activation in the mouse immature Sertoli cell line TM4. 1,25(OH)2-D3 potentiation of chloride currents was dependent on hormone concentration, and correlated with a significant increase in whole-cell capacitance within 20-40 min. In addition, Cl(-) currents were potentiated by the nongenomic VDR agonist 1alpha,25(OH)2 lumisterol D3 (JN), while 1,25(OH)2-D3 potentiation of channels was suppressed by nongenomic VDR antagonist 1beta,25(OH)2-vitamin D3 (HL). Treatment of TM4 cells with PKC and PKA activators PMA and forskolin respectively, increased Cl(-) currents significantly, while PKC and PKA inhibitors Go6983 and H-89, respectively, abolished 1,25(OH)2-D3 stimulation of Cl(-) currents, suggesting phosphorylation pathways in 1,25(OH))2-D3 mediated channel responses. RT-PCR demonstrated the expression of outwardly rectifying ClC-3 channels in TM4 cells. Taken together, our results demonstrate a PKA/PKC-dependent 1,25(OH)2-D3/VDR nongenotropic pathway leading to Cl(-) channel and exocytosis activation in Sertoli cells. We conclude that 1,25(OH)2-D3 appears to be a modulator of male reproductive functions at least in part by stimulating Sertoli cell secretory functions.

Transport of 45Ca from the lumen to the venous effluent was studied in duodena of normal, vitamin D3-replete (+D) chicks perfused through the celiac artery with 130 pM 1,25(OH)2D3 or vehicle. Administration of actinomycin D 3 h prior to perfusion did not alter the unstimulated transport rate or diminish the response to exogenous 1,25(OH)2D3: After 40 min exposure to the seco-steroid, 45Ca in the vascular effluent was 140% of control levels. The anti-microfilament agent cytochalasin b and the ionophore monensin, an inhibitor of Golgi function, similarly failed to suppress 1,25(OH)2D3-stimulated calcium transport. In pilot studies, Golgi and basal-lateral membrane fractions were prepared from duodenal epithelium of vitamin D-deficient (-D) chicks treated with vehicle or 650 pmol of 1,25(OH)2D3in vivo 2 h, 10 h, or 15 h before sacrifice, as well as from +D birds. Analyses of Golgi fractions for cathepsin B (CB) activity revealed a biphasic response with time, increasing to 200% of -D levels 2 h after 1,25(OH)2D3 administration and in equivalent preparations from +D birds. Less pronounced increases in acid phosphatase activity were observed in the same membrane fractions. In basal-lateral membranes, enhanced CB activity was detectable 10 h after 1,25(OH)2D3in vivo, rose to 155% of -D levels at 15 h, and to 245% of controls in fractions from +D birds, whereas acid phosphatase was 75%, 81%, and 125% of controls, respectively, at these times. Levels of CB in whole homogenates from which membranes were isolated rose to 200% of controls at 2 h and 10 h after 1,25(OH)2D3, but declined to 112% of controls in +D chicks. Acid phosphatase levels in the same homogenates were lower than or equivalent to control levels. These findings suggested additional perfusion studies in normal birds. Although vascular perfusion with iodoacetate and 1,25(OH)2D3, or leupeptin and 1,25(OH)2D3, failed to inhibit the effect of the vitamin D metabolite, 1,25(OH)2D3-enhanced calcium transport was abolished by addition of 1 mM leupeptin to the lumenal medium, but not by 1 mM pepstatin. The combined data suggest that lysosomes or their hydrolases may be involved in 1,25(OH)2D3-mediated transport of calcium across intestinal epithelial cells.

Fifteen hours after the intracardiac administration of a physiological dose (0.65 nmole) of radioactive cholecalciferol (vitamin D3) to rachitic chicks, the bulk of the radioactivity in the plasma, liver, and intestine exists as three polar metabolites. These are designated Metabolites 4A, 4B, and 4C on the basis of their chromatographic mobility on silicic acid columns. After doses of 0.65 to 6.5 nmoles of cholecalciferol, the predominant metabolite found in the plasma is 4A with smaller amounts of 4B and 4C. In the intestine, a primary target tissue of cholecalciferol, the predominant metabolite is 4B, with much smaller amounts of 4A and 4C. Metabolite 4A from both the plasma and intestine is tentatively identified as 25-hydroxycholecalciferol on the basis of its migration with 25-hydroxycholecalciferol during silicic acid column chromatography. The biological activity of Metabolites 4A and 4B was examined with an assay which measures intestinal calcium transport, in vivo. Both 4A and 4B have biological activity significantly greater than equivalent amounts of cholecalciferol. The kinetics of the appearance of Metabolite 4B in the intestine and its binding to intestinal chromatin are consistent with the lag in the physiological response to cholecalciferol. When radioactive Metabolite 4A is administered to rachitic chicks, the bulk of the plasma radioactivity remains as 4A, but the predominant metabolite found in the intestine is 4B. These data support the concept that Metabolite 4B may be the biologically active form of cholecalciferol in the intestine and that 25-hydroxycholecalciferol is an intermediate in its formation.

New knowledge of the biological and clinical importance of the steroid hormone 1alpha,25-dihydroxyvitamin D(3) [1alpha,25(OH)(2)D(3)] and its receptor, the vitamin D receptor (VDR), has resulted in significant contributions to good bone health. However, worldwide reports have highlighted a variety of vitamin D insufficiency and deficiency diseases. Despite many publications and scientific meetings reporting advances in vitamin D science, a disturbing realization is growing that the newer scientific and clinical knowledge is not being translated into better human health. Over the past several decades, the biological sphere of influence of vitamin D(3), as defined by the tissue distribution of the VDR, has broadened at least 9-fold from the target organs required for calcium homeostasis (intestine, bone, kidney, and parathyroid). Now, research has shown that the pluripotent steroid hormone 1alpha,25(OH)(2)D(3) initiates the physiologic responses of >/=36 cell types that possess the VDR. In addition to the kidney's endocrine production of circulating 1alpha,25(OH)(2)D(3,) researchers have found a paracrine production of this steroid hormone in >/=10 extrarenal organs. This article identifies the fundamentals of the vitamin D endocrine system, including its potential for contributions to good health in 5 physiologic arenas in which investigators have clearly documented new biological actions of 1alpha,25(OH)(2)D(3) through the VDR. As a consequence, the nutritional guidelines for vitamin D(3) intake (defined by serum hydroxyvitamin D(3) concentrations) should be reevaluated, taking into account the contributions to good health that all 36 VDR target organs can provide.

Vitamin D deficiency is associated with susceptibility to tuberculosis, and its biologically active metabolite, 1alpha,25 dihydroxyvitamin D(3) (1alpha,25(OH)(2)D(3)), has pleiotropic immune effects. The mechanisms by which 1alpha,25(OH)(2)D(3) protects against tuberculosis are incompletely understood. 1alpha,25(OH)(2)D(3) reduced the growth of mycobacteria in infected human PBMC cultures in a dose-dependent fashion. Coculture with agonists or antagonists of the membrane or nuclear vitamin D receptors indicated that these effects were primarily mediated by the nuclear vitamin D receptors. 1alpha,25(OH)(2)D(3) reduced transcription and secretion of protective IFN-gamma, IL-12p40, and TNF in infected PBMC and macrophages, indicating that 1alpha,25(OH)(2)D(3) does not mediate protection via these cytokines. Although NOS2A was up-regulated by 1alpha,25(OH)(2)D(3), inhibition of NO formation marginally affected the suppressive effect of 1alpha,25(OH)(2)D(3) on bacillus Calmette Guérin in infected cells. By contrast, 1alpha,25(OH)(2)D(3) strongly up-regulated the cathelicidin hCAP-18 gene, and some hCAP-18 polypeptide colocalized with CD14 in 1alpha,25(OH)(2)D(3) stimulated PBMC, although no detectable LL-37 peptide was found in supernatants from similar 1alpha,25(OH)(2)D(3)-stimulated PBMC cultures. A total of 200 mug/ml of the active peptide LL-37, in turn, reduced the growth of Mycobacterium tuberculosis in culture by 75.7%. These findings suggest that vitamin D contributes to protection against TB by "nonclassical" mechanisms that include the induction of antimicrobial peptides.

The endocrine physiology of the male involves the interplay of the several hormones associated with male sex determination, fetal development, postnatal growth, and sexual maturation as well as the maintenance of the male secondary sex characteristics. Its integrated operation is dependent upon the interaction of signals—both hormonal and neural—between the central nervous system, hypothalamus, pituitary, and testes. The two major functions of the testes are steroid hormone production and spermatogenesis. The hormones responsible for the development and maintenance of the male phenotype are the hypothalamic hormones GnRH and kisspeptin; the pituitary gonadotropins, LH and FSH, the androgenic steroid hormones, testosterone (produced by the gonads) and 5α-dihydrotestosterone (synthesized by certain target tissues; see Chapter 2); anti-Mullerian hormone, produced by the fetal testis; and inhibin, a protein hormone produced by the gonads in both sexes. The female steroid hormone, estradiol, also plays important roles in the male. This chapter discusses the biology and biochemistry of these hormones in the male. Because the male genotype and fetal testicular hormone production play such crucial roles in sexual differentiation, this topic is covered in this chapter as well, along with its final postnatal developmental step, puberty.

This chapter covers hormones of the kidney, heart, and circulatory system, which together play zan indispensable role in the maintenance of life in higher organisms. This includes maintaining the constancy of many components of the extracellular fluid, including the filtering of nitrogenous wastes and maintenance of normal blood pressure, but also as major sources of hormones. The kidney occupies a unique position within the physiological network of the living organism, functioning as the ultimate organ for the monitoring and conservation of body water and all its electrolyte constituents, as well as many small organic molecules. In addition, the kidney plays an important physiological role in maintaining a correct acid–base balance in the circulatory system. Thus it is not surprising to see an array of hormone production sites, as well as sites of hormone action concentrated in the kidney. Many aspects of these physiological structures and processes are also closely linked to the process of blood pressure regulation. Accordingly, a secondary objective of this chapter is to introduce the hormonal aspects of the cardiovascular system. The chapter provides an introduction to 11 hormones that are produced by or have a major action on the cardiovascular system.

A feature essential for life of higher vertebrates is their ability to maintain a relatively constant blood glucose concentration. Superimposed on the requirement for the maintenance of a constant blood glucose level are the perturbations in blood glucose that may occur naturally as a consequence of ongoing physiological and metabolic events. The endocrine gland largely responsible for the maintenance of blood glucose levels and the appropriate cellular uptake and exchange of “fuel metabolites” is the pancreas. The principal hormones secreted by the pancreas are insulin and glucagon; in addition, the pancreas also secretes somatostatin, pancreatic polypeptide (PP), and gastrin. This chapter covers the integrated actions of glucagon and insulin to effect blood glucose homeostasis, and the chemistry, biochemistry, and biological activities of the pancreatic hormones. Clinical aspects are also covered.

Through the secretion of thyroxine, T4, and its active derivative, T3, the thyroid governs myriad physiological events which bring about functional alterations in virtually all metabolic pathways and organs. Thyroid hormone modulates oxygen consumption and basal metabolic rate (BMR), as well as lipid, carbohydrate, and protein metabolism. It has fundamental effects on the nervous system both during development and in childhood and adulthood. Thyroid hormone regulates the synthesis and degradation rates of numerous proteins, including other growth factors and hormones, so that many of its effects emerge as secondary influences on other endocrine pathways. The effects of thyroid hormone fall into two categories of biological responses: (a) effects on cellular differentiation and development, particularly, but not limited to, the nervous system; and (b) effects on metabolic pathways through which the body uses carbohydrates, lipids, and proteins. These two actions are interconnected in that changes in development and growth are both a cause and a consequence of hormonal modulation of metabolism. This chapter covers thyroid hormone secretion, its regulation, and its actions and some of their mechanisms, setting the stage to observe the results of excess or insufficiency of this hormone. The symptoms of the most common disease states of the thyroid gland, covered here, are a direct consequence of an interruption of the known actions of its hormones. Furthermore, treatments of these conditions are based directly on our understanding of the mechanisms of the synthesis of thyroid hormone and its regulation.

The oral cavity, stomach, intestine, colon, liver, gallbladder, and pancreas operate as an integrated physiological unit to effect the digestion and absorption of dietary acquired nutrients. This chapter focuses on the gastrointestinal hormones. There are ~30 peptides known to be expressed within the digestive tract, thus making the stomach, intestine, and colon the largest endocrine organ in the body. The regulatory peptides synthesized by the gut include hormones, growth factors, and peptide neurotransmitters. The gastrointestinal hormones are a family of polypeptides produced by specialized endocrine cells present in the stomach, intestine, and colon which function both as traditional hormonal messengers and as neurotransmitters. These hormones mediate a wide variety of specific biological responses carried out by the stomach, intestine, and colon, as well as the endocrine and exocrine pancreas, liver, and gallbladder, which, when collectively integrated, optimize the physiological conditions necessary to permit the efficient digestion and absorption of protein, carbohydrates, and fat as well as minerals and vitamins from the lumen of the intestine. There is also increasing evidence of neuroendocrine communication between the brain and gut that provides an additional level of integration and complexity to these digestive processes.

The pituitary gland has often been called “the master gland” because so many important functions are governed by the hormones it secretes. In this chapter readers see that the pituitary hormones are under the control of both the central nervous system, especially the hypothalamus and the secretions of the distal endocrine glands, and other internal signals. The hypothalamic releasing hormones function to control the secretions of the anterior pituitary hormones, either positively or negatively. The hormones secreted by the distal endocrine glands exert, in general, negative feedback effects in the brain and in the anterior pituitary. There are many signals originating either outside or inside the body that are mediated by the central nervous system. Thus, changes in the environment can ultimately stimulate the secretion of hypothalamic releasing hormones, which, through their effects on pituitary hormonal secretion allow the body to adapt to the change. Likewise, signals from within the organism can trigger this cascade system so that secretion of hormones from distant target glands can be affected. The chapter covers control cascades; anatomical relationships; structure, synthesis, secretion, and target cells of the hypothalamic releasing hormones; chemistry of the anterior pituitary hormones; regulation and biological actions of growth hormone and prolactin; involvement of the hypothalamus in appetite regulation; and clinical aspects of the hypothalamus and pituitary.

This chapter focuses on four hormones: parathyroid hormone (PTH), calcitonin (CT), a steroid hormone metabolically produced from vitamin D3, namely 1α,25(OH)2-vitamin D3 [1α,25(OH)2D3] and fibroblast growth factor 23 (FGF23). Historically, the first three have been intimately involved in calcium and phosphate homeostasis. However, as a consequence of major advances in biological research over the past 30 years, these four hormones have been discovered to have many additional assignments. For optimal growth and function, organisms with a skeleton require an adequate supply of calcium and phosphate to meet both their structural and metabolic needs. These higher organisms must be dietarily supplied with calcium and phosphate on a regular basis. The plasma concentrations of these substances are maintained within a surprisingly narrow limit by an endogenous hormonal control mechanism. The maintenance of calcium and phosphate homeostasis involves the delicate and coordinated interrelationships of absorption by the intestine, accretion by bone osteoblasts and reabsorption by bone osteoclasts, and renal tubular reabsorption and/or urinary excretion of calcium and phosphate by the kidney. This chapter discusses the details of related anatomy, processes, and hormonal mechanisms.

With its discovery in 1920, the molecule vitamin D achieved prominence as a nutritionally essential vitamin important for calcium homeostasis, particularly in the intestine and bone. Then in 1932, the elucidation of vitamin D's chemical structure revealed that this vitamin was in fact a steroid. But it was not until the late 1960s that it was appreciated that the steroid vitamin D was a precursor of a new steroid hormone, 1alpha,25(OH)2-vitamin D3 [1alpha,25(OH)2D3], that is produced by the kidney acting as an endocrine gland. The discovery in 1969 of the nuclear vitamin D receptor (VDR) for 1alpha,25(OH)2D3 initiated a two-decade-long proliferation of reports that collectively described the broad sphere of influence of the vitamin D endocrine system that is defined by the presence of the VDR in over 30 tissue/organs of man. The new genomic frontiers defined by the cellular presence of the VDR include the immune system's B and T lymphocytes, hair follicle, muscle, adipose tissue, bone marrow, and cancer cells. Unexpectedly in the mid 1980s, a new world of 1alpha,25(OH)2D3-mediated rapid responses (RR) was discovered. These were responses that occurred too rapidly (minutes to an hour) to be explained as the simple consequence of the nuclear VDR regulating gene transcription. Some RR examples include the rapid intestinal absorption of calcium (transcaltachia), secretion of insulin by pancreatic beta-cells, opening of voltage-gated Ca2+ and Cl- channels in osteoblasts, and the rapid migration of endothelial cells. The question then arose as to whether there was a second receptor, apart from the nuclear VDR, which responded to the presence of 1alpha,25(OH)2D3 to generate RR? After some false starts, it now appears that the classic VDR, long known to reside in the cell nucleus, in some cells is also associated with caveolae present in the plasma membrane. Furthermore, the chemical properties of the conformationally flexible 1alpha,25(OH)2D3 allow it to generate different shaped ligands for the VDR that are selective either for genomic or for RR. This minireview summarizes a proposed conformational ensemble model of the VDR that provides insight into how different ligand shapes of 1alpha,25(OH)2D3 acting through the VDR in different cellular locations can selectively mediate both genomic and RR.

We recently demonstrated that suppressing 1alpha,25-(OH)2-D3 by increasing dietary calcium decreases adipocyte intracellular Ca2+ ([Ca2+]i), stimulates lipolysis, and inhibits lipogenesis. High calcium diets also increase core temperature and white adipose tissue uncoupling protein 2 (UCP2) expression in aP2-agouti transgenic mice. Accordingly, we have evaluated the role of 1alpha,25-(OH)2-D3 in regulating human adipocyte UCP2 expression. Treatment of human adipocytes for 48 h with 1 nM 1alpha,25-(OH)2-D3 inhibited UCP2 mRNA and protein levels by 50% (P

The steroid hormone 1α,25-dihydroxyvitamin D 3 [1α, 25-(OH) 2 D 3 ] promotes vascular smooth muscle cell (VSMC) growth and calcification, but the precise mechanism by which 1α, 25-(OH) 2 D 3 regulates VSMC migration is unknown. In rat aortic SMCs, we found that 1α, 25-(OH) 2 D 3 (0.1 to 100 nmol/L) induced a dose-dependent increase in VSMC migration. This response required the activation of phosphatidylinositol 3-kinase (PI3 kinase) because 1α, 25-(OH) 2 D 3 -induced migration was completely abolished by the PI3 kinase inhibitors, LY294002 (10 μmol/L) or wortmannin (30 nmol/L). Furthermore, the RNA polymerase inhibitor, 5,6-dichlorobenzimidazole riboside (50 μmol/L), did not affect 1α, 25-(OH) 2 D 3 -induced VSMC migration, suggesting that gene transcription is not involved in this rapid response. Using analogs of 1α, 25-(OH) 2 D 3 , which have been characterized for their abilities to induce either transcriptional or nontranscriptional responses of 1α, 25-(OH) 2 D 3 , we found that 1α,25-dihydroxylumisterol, which is a potent agonist of the rapid, nongenomic responses, was equipotent with 1α, 25-(OH) 2 D 3 in inducing PI3 kinase activity and VSMC migration. Moreover, 1β, 25-(OH) 2 D 3 , which specifically antagonizes the nongenomic actions of 1α, 25-(OH) 2 D 3 , abolished 1α, 25-(OH) 2 D 3 -induced PI3 kinase activity and VSMC migration, whereas the inhibitor of the genomic actions of vitamin D, (23S)-25-dehydro-1α-OH-D 3 -26,23-lactone, did not affect these responses. These results indicate that 1α, 25-(OH) 2 D 3 induces VSMC migration independent of gene transcription via PI3 kinase pathway, and suggest a possible mechanism by which 1α, 25-(OH) 2 D 3 may contribute to neointima formation in atherosclerosis and vascular remodeling.

We reported recently that suppression of the renal 1alpha,25-dihyroxyvitamin D3 (1lpha,25-(OH)2-D3) production in aP2-agouti transgenic mice by increasing dietary calcium decreases adipocyte intracellular Ca2+ ([Ca2+]i), stimulates lipolysis, inhibits lipogenesis, and reduces adiposity. However, it was not clear whether this modulation of adipocyte metabolism by dietary calcium is a direct effect of inhibition of 1alpha,25-(OH)2-D3-induced [Ca2+]i. Accordingly, we have now evaluated the direct role of 1alpha,25-(OH)2-D3. Human adipocytes exhibited a 1alpha,25-(OH)2-D3 dose-responsive (1-50 nM) increase in [Ca2+]i (P0.01). This action was mimicked by 1alpha,25-dihyroxylumisterol3 (1alpha,25-(OH)2-lumisterol3) (P0.001), a specific agonist for a putative membrane vitamin D receptor (mVDR), and completely prevented by 1b,25-dihydroxyvitamin D3 (1beta,25-(OH)2-D3), a specific antagonist for the mVDR. Similarly, 1alpha,25-(OH)2-D3 (5 nM) caused 50%-100% increases in adipocyte fatty acid synthase (FAS) expression and activity (P0.02), a 61% increase in glycerol-3-phosphate dehydrogenase (GPDH) activity (P0.01), and an 80% inhibition of isoproterenol-stimulated lipolysis (P0.001), whereas 1beta,25-(OH)2-D3 completely blocked all these effects. Notably, 1alpha,25-(OH)2-lumisterol3 exerted more potent effects in modulating adipocyte lipid metabolism, with 2.5- to 3.0-fold increases in FAS expression and activity (P0.001) and a threefold increase in GPDH activity (P0.001). Also 1alpha,25-(OH)2-lumisterol3 was approximately twice as potent in inhibiting basal lipolysis (P0.025), whereas 1beta,25-(OH)2-D3 completely blocked all these effects. These data suggest that 1alpha,25-(OH)2-D3 modulates adipocyte Ca2+ signaling and, consequently, exerts a coordinated control over lipogenesis and lipolysis. Thus, a direct inhibition of 1alpha,25-(OH)2-D3-induced [Ca2+]i may contribute to an anti-obesity effect of dietary calcium, and the mVDR may represent an important target for obesity.

Vitamin D 3 produces biologic responses as a consequence of its metabolism into 1α,25(OH) 2 -vitamin D 3 [1α,25(OH) 2 D 3 ] and 24R,25(OH) 2 -vitamin D 3 . The metabolic production of these two seco steroids and their generation of the plethora of biologic actions that are attributable to the parent vitamin D 3 are orchestrated via the integrated operation of the vitamin D endocrine system. This system is very similar in its organization to that of classic endocrine systems and is characterized by an endocrine gland (the kidney, the source of the two steroid hormones), target cells which possess receptors for the steroid hormones, and a feed-back loop involving changes in serum Ca 2+ that alter the secretion of parathyroid hormone (a stimulator of the renal 1-hydroxylase) which modulates the output by the kidney of the steroid hormones. There are, however, at least two unique aspects to the vitamin D endocrine system. (a) The chemical structures of vitamin D and its steroid hormones dictate that these be highly conformationally flexible molecules present a wide variety of shapes to their biologic environments. (b) It is now believed that 1α,25(OH) 2 D 3 produces biologic responses through two distinct receptors which recognize totally different shapes of the conformationally flexible 1α,25(OH) 2 D 3 . Thus, the classic actions of 1α,25(OH) 2 D 3 to regulate gene transcription occur as a consequence of the stereospecific interaction of a modified 6-s- trans bowl-shape of 1α,25(OH) 2 D 3 with its nuclear receptor (VDR nuc ). The ability of 1α,25(OH) 2 D 3 to generate a variety of rapid (seconds to minutes) biologic responses (opening of chloride channels, activation of PKC and MAP kinases) requires a planar 6-s- cis ligand shape which is recognized by a putative plasma membrane receptor (VDR mem ) to initiate appropriate signal transduction pathways. This report summarizes the evidence for the specificity of different ligand shapes and the operation of the two receptor families for 1α,25(OH) 2 D 3 .

The effect of 1α,25-dihydroxylumisterol3 (1α,25(OH)2lumisterol3) on insulin release from rat pancreatic β-cells was measured to investigate the nongenomic action of vitamin D via the putative membrane vitamin D receptor (mVDR). 1α,25(OH)2lumisterol3, a specific agonist of mVDR, dose-dependently augmented 16.7 mm glucose-induced insulin release from rat pancreatic islets and increased the intracellular Ca2+ concentration ([Ca2+]i), though not increasing Ca2+ efficacy in the exocytotic system. These effects were completely abolished by an antagonist of mVDR, 1β,25-dihydroxyvitamin D3 (1β,25(OH)2D3), or by a blocker of voltage-dependent Ca2+ channels, nitrendipine. Moreover, both [Ca2+]i elevation, caused by membrane depolarization, and sufficient intracellular glucose metabolism are required for the expression of these effects. 1α,25(OH)2lumisterol3, therefore, has a rapid insulinotropic effect, through nongenomic signal transduction via mVDR, that would be dependent on the augmentation of Ca2+ influx throu...

Recent studies have shown that 1alpha,25-dihydroxyvitamin D3 [1alpha,25-(OH)2D3] actions in cell growth and differentiation are mediated by both its nuclear receptor (VDRnuc) and its rapid membrane-related effects. In the present study, we investigated the effect of 1alpha,25-(OH)2D3 on p42mapk phosphorylation using human acute promyelocytic leukemia cells (NB4). 1Alpha,25-(OH)2D3 (10[-8] M) significantly increased p42mapk phosphorylation in a time- and dose-dependent manner, with the earliest response detectable at 30 sec. Because 1alpha,25-(OH)2D3 is a conformationally flexible molecule, we have used a series of conformationally locked (6-s-cis vs. 6-s-trans) analogs to evaluate which shape is optimal for activation. Four 6-s-cis-locked analogs (HF, JM, JN, and JP) and two 6-s-trans-locked analog (JB and JD) were studied. HF, JM, JN, and JP all increased p42mapk phosphorylation at 1 and 5 min (10[-8] M), but JB and JD had little effect. Analog HL [1beta,25-(OH)2D3], a specific antagonist for only the rapid effects of 1alpha,25-(OH)2D3, attenuated 1alpha,25-(OH)2D3-induced p42mapk phosphorylation 65-90%. To assess the potential involvement of the VDRnuc in mediating the analog's action, the relative abilities of the analogs to compete with [3H]1alpha,25-(OH)2D3 for binding in vitro to the VDRnuc of NB4 cells was measured. All 6-s-cis analogs bound poorly to VDRnuc (relative competitive index, 0.5-2%) compared with 1alpha,25-(OH)2D3 (relative competitive index, 100%). The present studies demonstrate for the first time that in NB4 cells 1alpha,25-(OH)2D3 rapidly activates the p42mapk pathway, and that this effect can be selectively mediated by analogs that can assume a 6-s-cis conformation.

Both 1alpha,25-dihydroxyvitamin D3 [1alpha,25(OH)2D3] and phorbol 12-myristate 13-acetate [PMA] are cytodifferentiation agents regulating myeloid cell line differentiation along the monocyte/macrophage pathway. Alkaline phosphatase (ALP) activity has been reported to be absent or present at low levels in monocytes/macrophages. In the present study, the acute promyelocytic leukemia cell line (NB4) was found to undergo monocyte/macrophage differentiation and strongly express ALP activity after exposure to the combination of 1alpha,25(OH)2D3 and PMA treatment. The ALP stimulation was both dose- and time-dependent with the ED50 doses for 1alpha,25(OH)2D3 and PMA at 2 x 10(-10) M and 4 x 10(-11) M, respectively. Assessment of the cell morphology via cell adherence, phagocytosis and ALP staining confirmed that NB4 cells treated by 1alpha,25(OH)2D3 plus PMA showed macrophage characteristics, but were also strongly stained with ALP. The present study is the first report that NB4 cells express ALP activity as a consequence of combination treatment with 1alpha,25(OH)2D3 and PMA.

金沢大学環日本海域環境研究センター生物多様性研究部門, The effects of vitamin D3, 24,25(OH)2 vitamin D3, 25(OH) vitamin D3 and 1,25(OH)2 vitamin D3 were investigated on the serum calcium and phosphate levels of freshwater catfish, Heteropneustes fossilis. The fish were injected daily intraperitoneally with these secosteroids for 10 days. Blood samples were collected at day 1, 3, 5 and 10. Serum calcium and inorganic phosphate levels were elevated by all of the treatments except for 24,25(OH)2 vitamin D3.

Six A-ring analogs of 1alpha,25-dihydroxyvitamin D3 (1, 1alpha,25-(OH)2-D3) 3-deoxy-3-thia-1alpha,25-(OH)2-D3 (3), 3-deoxy-3-thia-1alpha,25-(OH)2-D3-3alpha-oxide (6), 3-deoxy-3-thia-1alpha,25-(OH)2-D3-3beta-oxide (7) and the 5,6-trans counterparts 5, 8, and 9, respectively--were tested for their ability to inhibit 25-hydroxy-D3-1alpha-hydroxylase (1-OH-ase) in vitro in mitochondria isolated from kidneys of vitamin D deficient chicks. The six analogs were also evaluated in terms of their ability to bind to the chicken intestinal nuclear receptor (VDR) in comparison to the natural hormone 1alpha,25-(OH)2-D3. Analog 7 is not only the best inhibitor of the 1-OH-ase but it also binds effectively to the chick intestinal receptor. It is established that vitamin D analogs must have a 1alpha oxygen group for effective inhibition of the 1-OH-ase. This functional group is also needed for effective binding to the chick intestinal VDR.

Muscle has long been recognized as a target tissue for 1,25-dihydroxy-vitamin D3 (1,25[OH]2D3). Evidence of the presence of VDR is provided here, thus supporting the existence of a receptor-mediated mechanism of action of 1,25(OH)2D3. Vitamin D receptor (VDR) expression is evidenced by detection of VDR-mRNA, through reverse transcription and polymerase chain reaction (RT/PCR), in chicken muscle and muscle cells (myoblasts) as well as in a variety of tissues such as intestine, kidney, heart and brain. VDR presence is also demonstrated by Southern blot of PCR products with a specific VDR-cDNA probe and by immunocytochemistry carried out on myoblasts and cardiac myocytes. Localization of VDR is mainly nuclear and more faintly detected in the cytosol.

Vitamin D is responsible, through the actions of its metabolite, 1 alpha,25-dihydroxyvitamin D3 [1 alpha,25-(OH)2D3], for the generation of a wide array of biological responses, particularly in the intestine, kidney, and bone. 1 alpha,25-(OH)2D3 is known to interact with its nuclear receptor to mediate the regulation of gene transcription. Although many genes and gene products have been shown to be regulated by 1 alpha,25-(OH)2D3 (e.g. calbindin-D28K in the intestine and kidney; collagen, osteocalcin,and osteopontin in bone), their recognition has been largely the result of empirical testing. In this report we have used subtractive hybridization analysis of complementary DNA libraries prepared from messenger RNA (mRNA) isolated from the intestine and kidney of vitamin D-replete or vitamin D-deficient chicks to identify genes for novel proteins whose steady state mRNA levels are regulated by dietary vitamin D status. In the kidney we observed the down-regulated expression of at least seven mitochondrially encoded transcripts and the up-regulated expression of five nuclear encoded genes, two of which are metallothionein and the beta-subunit of aldolase. In the intestine, six mitochondrially encoded transcripts are up-regulated, and seven nuclear encoded transcripts were either up- or down-regulated. Thus, in addition to identifying new nuclear encoded genes whose mRNAs are regulated by vitamin D status, our approach has demonstrated the tissue-specific regulation of mitochondrial gene expression in the intestine and kidney.

Synopsis. Calcium is one ofthe key elements required for maintenance and survival oflife in animals with both endo- and exoskeletons. Because there is a wide variation in dietary calcium (dependent upon the local habitat) and dietary practices, and because there is a changing physiolog? ical need throughout life (e.g., for mammals, during growth, puberty, pregnancy, lactation, and menopause), it is essential that the process of intestinal calcium absorption be adaptable and responsive to both the dietary and physiological circumstances. This article reviews the evidence that transcaltachia, or the rapid stimulation of intestinal Ca2+ transport by the steroid hormone, 1 a,25(OH)2-vitamin D3, as studied in the chicken, meets many ofthe objectives of an adaptive intestinal calcium transport process. Transcaltachia is studied in a perfused chick duodenum, where 45Ca2+ is placed in the lumen and potential agonists are perfused into the celiac artery; the transcaltachic reponse represents the stimulation within 4-8 min ofthe transfer of 45Ca2+ from the lumen to the vascular perfusate. la,25(OH)2-vitamin D3 stimulation of transcaltachia occurs via nongenomic mechanisms which involve a plasma membrane receptor for the secosteroid and the coupled opening of voltage-gated Ca2+ channels on the basal lateral membrane of the intestinal epithelial cell and the acti? vation of the second messengers, protein kinase C and cAMP.

1 alpha,25-dihydroxyvitamin D3 [1 alpha,25(OH)2D3] has been shown to generate biological responses via both genomic and nongenomic pathways. In previous studies concerning the mechanism of 1 alpha,25(OH)2D3-stimulated rapid intestinal transport of Ca2+, a process termed transcaltachia, we assessed the ability of seven analogs of 1 alpha,25(OH)2D3 to initiate transcaltachia in the vitamin D-replete chick and bind in vitro to the classical nuclear chick intestinal receptor for 1 alpha,25(OH)2D3 (VDR). Two groups of analogs were found, one able to stimulate transcaltachia and the other able to bind to the VDR. In the present study, we have examined both the genomic effect of these analogs in the intestine of vitamin D-deficient chicks and their in vitro binding to vitamin D binding protein. It was found that analogs known to bind effectively to the nuclear receptor in vitro could achieve a significant occupancy of the VDR in vivo and stimulated calbindin-D28K messenger RNA and protein synthesis. In contrast, those analogs that were good agonists of transcaltachia were ineffective either in occupying the VDR in vitro or in stimulating in vivo calbindin-D28K messenger RNA and protein synthesis. These differences are consistent with our hypothesis that there may be two receptors for 1,25-(OH)2D3 with differing ligand binding domains, one being the classical nuclear VDR and the second a membrane receptor associated with transcaltachia.

Biological responses mediated by vitamin D occur as a consequence of the integrated actions of the vitamin D endocrine system. The vitamin D endocrine system is characterized by the sequential two-step metabolism of vitamin D to 1α,25(OH)2D3 by the liver and kidney, and by the ability to generate biological responses in over 30 target tissues through nuclear receptor (nVDR) regulation of gene transcription and nongenomic pathways. It is now clear that the vitamin D endocrine system embraces many more target tissues than simply the intestine, bone and kidney. Notable additions to this list of tissues containing the nVDR include pancreatic B cells, pituitary gland, breast tissue, placenta, lymphocytes, keratinocytes, colon, and prostate, as well as many cancer cell lines. In addition to the classical actions of 1α,25(OH)2D3 on mediating calcium homeostasis, this seco steroid has been identified as a potent stimulator of cell differentiation as well as an inhibitor of proliferation. Over the past decade at least 400 analogs of 1α,25(OH)2D3 have been chemically synthesized and their biological properties systematically explored in a variety of assays which quantified both their calcemic effects and cell differentiating potential. The objective has been to identify new analogs devoid of the classical calcemic consequences of high does of 1α,25(OH)2D3, namely hypercalcemia, soft tissue calcification and nephrocalcinosis. As a consequence, several analogs of 1α,25(OH)2D3 have recently been identified and are discussed in this paper for consideration as possible chemotherapeutic agents for acute promyelocytic leukemia, breast, colon, and prostate cancer, or as immunosuppressive agents with possible beneficial structure-activity profiles for use in cardiac allografts, autommune graft rejection, lupus erthematosus and psoriasis.

Computer simulation of calcium homeostasis in chicks predicted an oscillatory behavior of bone calcium flow and kidney 25-hydroxyvitamin D3-1-hydroxylase with a periodicity of 56 h and a 9 h phase difference between the two signals. In growing chickens subjected to a light: dark cycle of 22:2 h, and intravenously dosed with 45Ca, the temporal changes in plasma 45Ca could be described by an exponential decline with superimposed diurnal oscillations. The activity of the renal 25-hydroxyvitamin D3-1-hydroxylase in chicks subjected to a 12:12 h light: dark cycle ALSO followed diurnal oscillations, with a nadir at the beginning of the light period and a peak 12 h later. The production of 1,25-dihydroxyvitamin D3 by primary cultures of chicken kidney cells oscillated with a periodicity of 5.6 h or shorter. It is suggested that despite the differences in phase and periodicity between the simulation predictions and actual results, the oscillations in both 1-hydroxylase and bone calcium flow could be coupled through the hormonal systems involved in regulation of plasma calcium.

The steroid hormone 1 alpha,25-dihydroxyvitamin D3 (1 alpha,25-(OH)2D3) elicits biological responses by both genomic and nongenomic pathways. This report describes purification of a receptor for 1 alpha,25-(OH)2D3 (VDR) located in the basal-lateral membrane (BLM) of vitamin D-replete chick intestinal epithelium, which is implicated in the nongenomic stimulation of calcium transport (transcaltachia). The BLM-VDR exhibited saturable binding for [3H]1,25-(OH)2D3 (KD = 0.72 x 10(-9)M, Bmax = 0.24 pmol/mg of protein). A 4500-fold purification of the BLM-VDR receptor was achieved. In addition, saturable binding was observed for [3H]24R,25-(OH)2D3 at physiologically relevant levels (KD = 19 x 10(-9) M, Bmax = 2.4 pmol/mg of protein) to a component apparently distinct from the 1 alpha,25-(OH)2D3 BLM-VDR. A functional correlation between the BLM-VDR and transcaltachia was observed in three experimental situations: (i) vitamin D deficiency, which suppresses transcaltachia, resulted in reduced specific [3H]1 alpha,25-(OH)2D3 binding in the BLM-VDR, relative to corresponding fractions from vitamin D-replete chicks; (ii) the BLM-VDR exhibited down-regulation of specific [3H]1 alpha,25-(OH)2D3 binding following exposure to nonradioactive 1 alpha,25-(OH)2D3; and (iii) the relative potencies of two "6-s-cis" analogs of 1 alpha,25-(OH)2D3 to initiate transcaltachia and their ability to compete with [3H]1 alpha,25-(OH)2D3 for binding to the BLM-VDR were parallel. The combined results support the existence of a plasmalemal 1 alpha,25-(OH)2D3 receptor which is a prime candidate for signal transduction in transcaltachia.

The process of involvement of the classical nuclear receptor with hormone-dependent nongenomic, extranuclear cell signaling contributes to cell biology at multiple levels. The classical view/assumption that the binding of 1,25(OH) 2 D 3 to the VDR simply activates its translocation to the nucleus of the cell where it cis- regulates the genome has been proven to be an oversimplification for the VDR and many other NRs. We have described and highlighted the vitamin D/VDR structure–function studies that differentiate nuclear and extranuclear signaling. Specifically, the presence of two overlapping ligand-binding sites within the VDR provides a physicochemical explanation for the analog-specific functions known for 1α,25(OH) 2 -lumisterol D 3 (JN) and 1β,25(OH) 2 -vitamin D 3 (HL) and the ability of 1,25(OH) 2 D 3 to send signals dynamically by functioning as both a nongenomic and genomic agonist ligand. 1,25(OH) 2 D 3 extranuclear, nongenomic, rapid responses include the regulation of phospholipases, phosphatases, kinases, ion channels, and second messengers. These signaling factors can themselves regulate the genome and transcribed/translated products via mechanisms that integrate with 1,25(OH) 2 D 3 -VDR cis regulation of genes. In this way, extranuclear signaling can alter cell physiology in a manner independent of nuclear VDR transregulation. Thus by expanding our understanding of nongenomic structure–function and combining this knowledge with what is known regarding genomic structure–function, we hope to elucidate the mechanistic nuances that underpin the adverse responses of hypercalcemia and soft tissue calcification.

The steroid hormone 1 alpha,25-dihydroxyvitamin D3 (1 alpha,25-(OH)2D3) generates biological responses via both genomic and nongenomic mechanisms. This article reports the biological profile of four A-ring diastereomers of this secosteroid (results are expressed as percentage of the response of 1 alpha,25-(OH)2D3. The activity of the compounds, 1 alpha,25-(OH)2D3, 1 alpha,25-(OH)2-3-epivitamin D3, 1 beta,25-(OH)2D3, 1 beta,25-(OH)2-3-epivitamin D3, for in vivo intestinal Ca2+ absorption and bone Ca2+ mobilization and in vitro binding to the nuclear receptor (genomic responses) was, respectively, 100, 2.8, 1 alpha,25-(OH)2-3-epivitamin D3 > 1 beta,25-(OH)2D3 > 1 beta,25-(OH)2-3-epivitamin D3. 1 beta,25-(OH)2D3 was a potent antagonist of 1 alpha,25-(OH)2D3-mediated transcaltachia and 45Ca2+ uptake in ROS 17/2.8 cells but was unable to block the genomic 1 alpha,25-(OH)2D3 induction of chick calbindin-D28k (in vivo), induction of MG-63 cell osteocalcin, and HL-60 cell differentiation. These results suggest that analogs of 1 alpha,25-(OH)2D3 may be synthesized which are selective agonists or antagonists of genomic or nongenomic responses in the vitamin D endocrine system.

HL-60 cells have been shown to differentiate into monocyte-like cells as a consequence of the interaction of 1,25-dihydroxyvitamin D 3 (1,25(OH) 2 D 3 ) with a specific intracellular receptor (VDR). Of c-myc it has been reported that it plays a role in cell proliferation and differentiation. We have investigated the expression of VDR and c-myc mRNA of three subclones of HL-60 cells (N, R and MR) after exposure to 1,25(OH) 2 D 3 . The N cell is a normal clone in which 1,25(OH) 2 D 3 inhibited cell proliferation and also induced cell differentiation. The R cell is a resistant clone whose proliferation and differentiation are not affected by 1,25(OH) 2 D 3 . The MR cell is a mixed response type clone whose proliferation was not inhibited by 1,25(OH) 2 D 3 , while its differentiation was actually induced by 1,25(OH) 2 D 3 . The VDR mRNA expression of the N and MR cells reached a peak at 2 h (2.3- and 2.6-fold induction, respectively) and returned to the control level after 24 h treatment of 1,25(OH) 2 D 3 . The c-myc mRNA expression was significantly inhibited in the l,25(OH) 2 D 3 -induced N cells, but not in the MR cells. In contrast, 1,25(OH) 2 D 3 did not induce any changes of the VDR and c-myc mRNA levels in the R cells. In separate experiments, the level of the VDR in the three clones was determined via Scatchard analysis; the VDR of the N, MR and R cells were found to be 4500/cell, 3570/cell and less than 600/cell, respectively. These results are consistent with an involvement of 1,25(OH) 2 D 3 and VDR in differentiation events, and support the notion that c-myc is related to cell proliferation rather than cell differentiation.

Vitamin D(3) is metabolized in vivo through 25-(OH)D(3) (25D) to both 1α,25-(OH)(2)D(3) (1,25D) and 24R,25-(OH)(2)D(3) (24,25D). Whereas it is assumed that this metabolism occurs primarily in the kidney, recent studies show that there are extrarenal 1α-and 24R-hydroxylase activities as well, and in chondrocytes, these enzymes are regulated by hormones and growth factors. Furthermore, chondrocytes from the resting zone of growth plate cartilage are a target cell population for 24,25D action, suggesting that this vitamin D metabolite may be targeted to this tissue in vivo. To test this hypothesis, 30 normal male Sprague Dawley rats (120 ±20 g) were divided into three groups of eight animals each, and a control group of six animals, and fed ad libitum for 2 wk, a standard rat chow (Teklad LM-485), which contained 3 IU vitamin D(3)/g. The rats were then injected im daily at 9:00AM: , for 4 consecutive d, with 0.1 mL of either [(3)H]-25D, [(3)H]-1,25D or [(3)H]-24,25D. Each dose contained 13 pmol of hormone (0.36 μCi/dose). The distribution of these metabolites was assessed in tibial bone (B) following ablation of the bone marrow, articular cartilage from the tibia (AC), costochondral growth plate cartilage (GC), serum (S), small intestine (I), and kidney (K). The use of high specific activity tritiated vitamin D metabolites facilitated determining tissue localization and further metabolism without perturbation of the body pools of each major metabolite. Accumulation of [(3)H]-1,25D or [(3)H]-24,25D in each tissue was compared to circulating serum levels. In rats dosed with [(3)H]-25D, the tissue:serum ratios for 1,25D were 4.1 (AC), 35.4 (GC), 1.3 (B), 0.7 (K), and 3.0 (I); and tissue:serum ratios for 24,25D were 1.6 (AC), 9.9 (GC), 0.04 (B), 0.2 (K), and 0.4 (I). In rats dosed with [(3)H]-24,25D alone, GC was the only tissue to accumulate the administered metabolite at a concentration significantly higher than that of serum. Similarly, in rats dosed with [(3)H]-1,25D alone, GC was the only tissue to accumulate 1,25D at a concentration higher than that of serum. These results demonstrate, for the first time, that under in vivo conditions, GC specifically accumulates 24,25D and 1,25D. This suggests that growth plate may be a target organ for these two hormones.

As background information, the reader should appreciate that at he 13th Vitamin DWorkshop in 2006 it was agreed that about half f elderly North Americans and Western Europeans and probably lso two thirds of the rest of the world are vitamin D deficient as udgedby their inability tomaintain a healthy bonedensity [1–3]. It as also generally agreed that the serum concentration of 25(OH)D n normal subjects is the best indicator for judging the vitamin D tatus in patients with vitamin D-related disease states [4]. The14thWorkshoponVitaminD,held inBrugge, Belgium,Octoer 4–8, 2009wasattendedby419 scientists from35countrieswho ere privileged to listen and participate in a Vitamin D Roundtable hat was held in order to allow presentation and broad discusion of two distinct views of and approaches to worldwide vitamin nutritional status. One Roundtable position is that an absolute inimum 25(OH)D level of 20ng/ml (50nmol/l) is necessary in all ndividuals in order to support and maintain all the classic actions f vitamin D on bone andmineral health and that, according to this riterion, a large proportion of the world’s population is vitamin D eficient. Thosewhohold this position further believe that thehuge ffort needed to ameliorate this deficiency must be undertaken as oon and actively as possible and that the target 25(OH)D levels of 20ng/ml should be obtained in the majority of the target populaion. The second Roundtable position is that newer data showing ssociations between vitamin D status and prevalence of several iseases such as cardiovascular disease, hypertension, colon and reast cancer, multiple sclerosis as well as the involvement of vitain D in muscle strength and immune functions [5], indicates that arget levels of 25(OH)D should be 30–40ng/ml (75–100nmol/l) t the minimum. As a basis for policy decision making, these two ositions are incompatible with one another. However, through onsideration of the aspects of vitamin D nutrition upon which the roponents of the two views agree, as well as acknowledging diferences in opinion, consensus on how to proceed in the near term an emerge. This was the goal of the Vitamin D Roundtable. TheRoundtable, chaired byAnthonyNorman (USA) andChristoher Gallagher (USA), began with 15-min presentations from obert Heaney (USA) and Reinhold Vieth (Canada), both propoents of 25(OH)D>40ng/ml, followed by presentations by Roger ouillon (Belgium) and Paul Lips (Netherlands),who advocateminmum25(OH)D levels of 20ng/ml. A selected group of experts from round theworld, Bess Dawson-Hughes (USA), John Pettifor (South frica), Peter Ebeling (Australia), and Christine Lamberg-Allardt

The steroid hormone 1α,25-dihydroxyvitamin-D 3 [1α,25(OH) 2 D 3 ] stimulates biological responses via both genomic mechanisms and nongenomic mechanisms (opening of voltage-gated Ca 2+ channels). We report here that 1β,25(OH) 2 -vitamin-D 3 (a) is devoid of activity as an agonist for transcaltachia, (b) is a potent stereospecific antagonist of 1α,25(OH) 2 D 3 stimulation of the nongenomic transcaltachia response and also (c) has less than 1% the ability of 1α,25(OH) 2 D 3 to bind to the chick intestinal nuclear 1β,25(OH) 2 D 3 receptor. We conclude that the membrane response element(s) which generates the nongenomic response of transcaltachia has a different ligand specificity than the classic nuclear 1α,25(OH) 2 D 3 receptor.

Physiological studies of the Atlantic cod, Gadus morhua, have suggested a role for the vitamin D3 system in this marine teleost similar to that in other vertebrates. Accordingly, the present study was carried out to assess the plasma concentrations of vitamin D3, 25-hydroxyvitamin D3 (25OHD3), and 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] in this fish. Additionally, the presence of binding proteins in plasma and target-specific tissue receptors for these vitamin D3 metabolites was studied in organs normally associated with calcium regulation. Plasma levels of 25-OHD3 (undetectable to 148 pg/ml; n = 5) were comparatively low (20-30 ng/ml), whereas the levels of vitamin D3 (approximately 30 ng/ml) and 1,25-(OH)2D3 (approximately 50 pg/ml) were comparable to levels reported in higher vertebrates. Cod plasma contained a protein that bound both 25OHD3 and 1,25-(OH)2D3. This plasma binding protein revealed low affinity for 25OHD3, did not bind G-actin, and had a sedimentation coefficient of 3.4S. High affinity 1,25-(OH)2D3 receptors [Kd, 1.02 +/- 0.36 (n = 6), 1.02 +/- 0.3 (n = 5), and 0.95 +/- 0.51 (n = 5) nM; mean +/- SEM] were found in high salt cytosols from intestine, liver, and gills, respectively, and had sedimentation coefficients (3.6-3.8S in 0.3 M KCl sucrose gradients) similar to those in higher vertebrates. No specific 1,25-(OH)2D3 binding was found in kidney, ultimobranchial glands, corpuscles of Stannius, or bone. The finding of significant quantities of 1,25-(OH)2D3 in the plasma, the presence of plasma binding proteins that bind this seco-steroid, and the localization of specific high affinity receptors for this metabolite in calcium regulatory tissues in teleosts are all consistent with a physiological role for the vitamin D3 system in the calcium regulation of the cod.