Hormone Derivatives Research Articles

Rapid and Comprehensive Impurity Profiling of Synthetic Thyroxine by Ultrahigh-Performance Liquid Chromatography–High-Resolution Mass Spectrometry

Rapid and efficient quality control according to the public authority regulations is mandatory to guarantee safety of the pharmaceuticals and to save resources in the pharmaceutical industry. In the case of so-called "grandfather products" like the synthetic thyroid hormone thyroxine, strict regulations enforce a detailed chemical analysis in order to characterize potentially toxic or pharmacologically relevant impurities. We report a straightforward workflow for the comprehensive impurity profiling of synthetic thyroid hormones and impurities employing ultrahigh-performance liquid chromatography (UHPLC) hyphenated to high-resolution mass spectrometry (HRMS). Five different batches of synthetic thyroxin were analyzed resulting in the detection of 71 impurities within 3 min total analysis time. Structural elucidation of the compounds was accomplished via a combination of accurate mass measurements, computer based calculations of molecular formulas, multistage high-resolution mass spectrometry (HRMS(n)), and nuclear magnetic resonance spectroscopy, which enabled the identification of 71 impurities, of which 47 have been unknown so far. Thirty of the latter were structurally elucidated, including products of deiodination, aliphatic chain oxidation, as well as dimeric compounds as new class of thyroid hormone derivatives. Limits of detection for the thyroid compounds were in the 6 ng/mL range for negative electrospray ionization mass spectrometric detection in full scan mode. Within day and day-to-day repeatabilities of retention times and peak areas were below 0.5% and 3.5% R.SD. The performance characteristics of the method in terms of robustness and information content clearly show that UHPLC-HRMS is adequate for the rapid and reliable detection, identification, and semiquantitative determination of trace levels of impurities in synthetic pharmaceuticals.

Biosynthesis of 3-Iodothyronamine (T1AM) Is Dependent on the Sodium-Iodide Symporter and Thyroperoxidase but Does Not Involve Extrathyroidal Metabolism of T4

3-Iodothyronamine (T(1)AM) is an endogenous thyroid hormone derivative with unknown biosynthetic origins. Structural similarities have led to the hypothesis that T(1)AM is an extrathyroidal metabolite of T(4). This study uses an isotope-labeled T(4) [heavy-T(4) (H-T(4))] that can be distinguished from endogenous T(4) by mass spectrometry, which allows metabolites to be identified based on the presence of this unique isotope signature. Endogenous T(1)AM levels depend upon thyroid status and decrease upon induction of hypothyroidism. However, in hypothyroid mice replaced with H-T(4), the isotope-labeled H-T(3) metabolite is detected, but no isotope-labeled T(1)AM is detected. These data suggest that T(1)AM is not an extrathyroidal metabolite of T(4), yet is produced by a process that requires the same biosynthetic factors necessary for T(4) synthesis.

P02-18 Pharmacological characterisation of NNC0195–0092, a long-acting growth hormone derivative

P02-18 Pharmacological characterisation of NNC0195–0092, a long-acting growth hormone derivative

Impact of pretreatment factors, biopsy Gleason grade volume indices and post-treatment nadir PSA on overall survival in patients with metastatic prostate cancer treated with step-up hormonal therapy

In the era when various treatment agents for advanced-stage prostate cancer are available, it is important to investigate overall survival for metastatic prostate cancer treated with step-up hormonal treatment as a reference against new treatment regimens, including antitumor agents and/or new hormonal derivatives, and it is desirable to explore pretreatment, biopsy-related and post-treatment prognostic factors to establish tailor-made treatment strategies. Between 1992 and 2002, 94 patients were diagnosed with prostate cancer with distant metastases in our facility. Various pretreatment clinical findings including serum PSA, testosterone, alkaline phosphatase (ALP), digital rectal examination (DRE) and extension of disease score were investigated for predicting outcomes in step-up hormonal treatment. We also investigated the impact of pathological findings including Gleason grading, tumor volume indices, various Gleason grade 5 volume indices in biopsy specimens, and post-treatment PSA nadir following step-up hormonal treatment on overall survival. The 3- and 5-year overall survival was 72.4% and 62.5%, respectively. According to univariate analyses, patients with PSA ≤ 100 ng ml(-1), ALP <440 IU l(-1), T1c to T3 on DRE, extension of disease (EOD) score 0-3, no Gleason grade 5 cancer in biopsy specimens or less such cancer and good response at any stage of hormonal therapy had significantly better overall survival than did patients with alternative status. A multivariate Cox proportional hazard model revealed that PSA nadir after first-line hormonal treatment, PSA nadir after second-line treatment and Gleason grade 5 volume index were independent prognostic factors. Even in very advanced prostate cancer, local pathological indices, a Gleason grade 5 volume index in particular, could differentiate patients with better prognosis from worse prognosis. Step-up hormonal therapy including luteinizing hormone-releasing hormone agonist, estrogen derivatives and steroid hormones may be valuable in patients with metastatic prostate cancer, especially in good responders at any stage of hormonal therapy.

Molecules important for thyroid hormone synthesis and action - known facts and future perspectives

Thyroid hormones are of crucial importance for the functioning of nearly every organ. Remarkably, disturbances of thyroid hormone synthesis and function are among the most common endocrine disorders affecting approximately one third of the working German population. Over the last ten years our understanding of biosynthesis and functioning of these hormones has increased tremendously. This includes the identification of proteins involved in thyroid hormone biosynthesis like Thox2 and Dehal where mutations in these genes are responsible for certain degrees of hypothyroidism. One of the most important findings was the identification of a specific transporter for triiodothyronine (T3), the monocarboxylate transporter 8 (MCT8) responsible for directed transport of T3 into target cells and for export of thyroid hormones out of thyroid epithelial cells. Genetic disturbances of MCT8 in patients result in a biochemical constellation of high T3 levels in combination with low or normal TSH and thyroxine levels leading to a new syndrome of severe X-linked mental retardation. Importantly mice lacking MCT8 presented only with a mild phenotype, indicating that compensatory mechanisms exist in mice. Moreover, it has become clear that not only genomic actions of T3 exist. T3 is also capable to activate adhesion receptors and it signals via activation of PI3K and MAPK pathways. Most recently, thyroid hormone derivatives were identified, the thyronamines which are decarboxylated thyroid hormones initiating physiological actions like lowering body temperature and heart rate, thereby acting in opposite direction to the classical thyroid hormones. So far it is believed that thyronamines function via the activation of a G-protein coupled receptor, TAAR1. The objective of this review is to summarise the recent findings in thyroid hormone synthesis and action and to discuss their implications for diagnosis of thyroid disease and for treatment of patients.

Effects of the thyroid hormone derivatives 3-iodothyronamine and thyronamine on rat liver oxidative capacity

Thyronamines T 0AM and T 1AM are naturally occurring decarboxylated thyroid hormone derivatives. Their in vivo administration induces effects opposite to those induced by thyroid hormone, including lowering of body temperature. Since the mitochondrial energy-transduction apparatus is known to be a potential target of thyroid hormone and its derivatives, we investigated the in vitro effects of T 0AM and T 1AM on the rates of O 2 consumption and H 2O 2 release by rat liver mitochondria. Hypothyroid animals were used because of the low levels of endogenous thyronamines. We found that both compounds are able to reduce mitochondrial O 2 consumption and increase H 2O 2 release. The observed changes could be explained by a partial block, operated by thyronamines, at a site located near the site of action of antimycin A. This hypothesis was confirmed by the observation that thyronamines reduced the activity of Complex III where the site of antimycin action is located. Because thyronamines exerted their effects at concentrations comparable to those found in hepatic tissue, it is conceivable that they can affect in vivo mitochondrial O 2 consumption and H 2O 2 production acting as modulators of thyroid hormone action.

Molecular Basis for Certain Neuroprotective Effects of Thyroid Hormone

The pathophysiology of brain damage that is common to ischemia–reperfusion injury and brain trauma include disodered neuronal and glial cell energetics, intracellular acidosis, calcium toxicity, extracellular excitotoxic glutamate accumulation, and dysfunction of the cytoskeleton and endoplasmic reticulum. The principal thyroid hormones, 3,5,3′-triiodo-l-thyronine (T3) and l-thyroxine (T4), have non-genomic and genomic actions that are relevant to repair of certain features of the pathophysiology of brain damage. The hormone can non-genomically repair intracellular H+ accumulation by stimulation of the Na+/H+ exchanger and can support desirably low [Ca2+]i.c. by activation of plasma membrane Ca2+–ATPase. Thyroid hormone non-genomically stimulates astrocyte glutamate uptake, an action that protects both glial cells and neurons. The hormone supports the integrity of the microfilament cytoskeleton by its effect on actin. Several proteins linked to thyroid hormone action are also neuroprotective. For example, the hormone stimulates expression of the seladin-1 gene whose gene product is anti-apoptotic and is potentially protective in the setting of neurodegeneration. Transthyretin (TTR) is a serum transport protein for T4 that is important to blood–brain barrier transfer of the hormone and TTR also has been found to be neuroprotective in the setting of ischemia. Finally, the interesting thyronamine derivatives of T4 have been shown to protect against ischemic brain damage through their ability to induce hypothermia in the intact organism. Thus, thyroid hormone or hormone derivatives have experimental promise as neuroprotective agents.

Neuroprotective Actions of Neurosteroids

Neurosteroids were initially defined as steroid hormones locally synthesized within the nervous tissue. Subsequently, they were described as steroid hormone derivatives that devoid hormonal action but still affect neuronal excitability through modulation of ionotropic receptors. Neurosteroids are further subdivided into natural (produced in the brain) and synthetic. Some authors distinguish between hormonal and regular neurosteroids in the group of natural ones. The latter group, including hormone metabolites like allopregnanolone or tetrahydrodeoxycorticosterone, is devoid of hormonal activity. Both hormones and their derivatives share, however, most of the physiological functions. It is usually very difficult to distinguish the effects of hormones and their metabolites. All these substances may influence seizure phenomena and exhibit neuroprotective effects. Neuroprotection offered by steroid hormones may be realized in both genomic and non-genomic mechanisms and involve regulation of the pro- and anti-apoptotic factors expression, intracellular signaling pathways, neurotransmission, oxidative, and inflammatory processes. Since regular neurosteroids show no affinity for steroid receptors, they may act only in a non-genomic mode. Multiple studies have been conducted so far to show efficacy of neurosteroids in the treatment of the central and peripheral nervous system injury, ischemia, neurodegenerative diseases, or seizures. In this review we focused primarily on neurosteroid mechanisms of action and their role in the process of neurodegeneration. Most of the data refers to results obtained in experimental studies. However, it should be realized that knowledge about neuroactive steroids remains still incomplete and requires confirmation in clinical conditions.

Polymorphisms in genes involved in the estrogen pathway and mammographic density

BackgroundSingle nucleotide polymorphisms (SNPs) in genes involved in the estrogen pathway appear to be associated with breast cancer risk and possibly with mammographic density (MD), but little is known of these associations among premenopausal women. This study examines the association of 11 polymorphisms in five estrogen-related genes (estrogen receptors alpha and beta (ERα, ERβ), 17β-hydroxysteroid dehydrogenase 1 (HSD17B1), catechol-O-methyltransferase (COMT), cytochrome P450 1B1 (CYP1B1)) with premenopausal MD. Effect modification of four estrogen-related factors (parity, age at menarche, hormonal derivatives use and body mass index (BMI)) on this relation is also assessed.MethodsPolymorphisms were genotyped in 741 premenopausal Caucasian women whose MD was measured in absolute density (AD, cm2) and percent density using a computer-assisted method. Multivariate linear models were used to examine the associations (Ptrend) and interactions (Pi).ResultsNone of the SNPs showed a statistically significant association with AD. However, each additional rare allele of rs1056836 CYP1B1 was associated with a reduction in AD among nulliparous women (Ptrend = 0.004), while no association was observed among parous women (Ptrend = 0.62; Pi = 0.02). An increase in the number of rare alleles of the HSD17B1 SNP (rs598126 and rs2010750) was associated with an increase in AD among women who never used hormonal derivatives (Ptrend = 0.06 and Ptrend = 0.04, respectively), but with a decrease in AD among past hormonal derivatives users (Ptrend = 0.04; Pi = 0.02 and Ptrend = 0.08; Pi = 0.01, respectively). Moreover, a negative association of rs598126 HSD17B1 SNP with AD was observed among women with higher BMI (>median) (Ptrend = 0.01; Pi = 0.02). A negative association between an increased number of rare alleles of COMT rs4680 SNP and AD was limited to women who never used hormonal derivatives (Ptrend = 0.02; Pi = 0.03) or with late age at menarche (>median) (Ptrend = 0.03; Pi = 0.02). No significant association was observed between polymorphisms in the ERα or ERβ genes and AD. Similar results, although less significant, were observed when MD was assessed in percent density.ConclusionSNPs located in CYP1B1, COMT or HSD17B1 genes seem to be associated with MD in some strata of estrogen-related factors. Our findings suggest that modifying effects of estrogen-related factors should be considered when evaluating associations of polymorphisms in estrogen-related genes with premenopausal mammographic density.

Thyronamines—Past, Present, and Future

Thyronamines (TAMs) are a newly identified class of endogenous signaling compounds. Their structure is identical to that of thyroid hormone and deiodinated thyroid hormone derivatives, except that TAMs do not possess a carboxylate group. Despite some initial publications dating back to the 1950s, TAMs did not develop into an independent area of research until 2004, when they were rediscovered as potential ligands to a class of G protein-coupled receptors called trace-amine associated receptors. Since this discovery, two representatives of TAMs, namely 3-iodothyronamine (3-T(1)AM) and thyronamine (T(0)AM), have been detected in vivo. Intraperitoneal or central injection of 3-T(1)AM or T(0)AM into mice, rats, or Djungarian hamsters caused various prompt effects, such as metabolic depression, hypothermia, negative chronotropy, negative inotropy, hyperglycemia, reduction of the respiratory quotient, ketonuria, and reduction of fat mass. Although their physiological function remains elusive, 3-T(1)AM and T(0)AM have already revealed promising therapeutic potential because they represent the only endogenous compounds inducing hypothermia as a prophylactic or acute treatment of stroke and might thus be expected to cause fewer side effects than synthetic compounds. This review article summarizes the still somewhat scattered data on TAMs obtained both recently and more than 20 yr ago to yield a complete and updated picture of the current state of TAM research.

Vitamin D homeostasis is compromised due to increased urinary excretion of the 25-hydroxycholecalciferol-vitamin D-binding protein complex in the Zucker diabetic fatty rat

Altered serum concentrations of the major circulating form of vitamin D [25-hydroxycholecalciferol (25D(3))] and its active hormone derivative [1,25-dihydroxycholecalciferol (1,25D(3))] have been linked to non-insulin-dependent diabetes mellitus (NIDDM). However, a mechanistic basis for this occurrence has not been fully elucidated. Normally, renal reabsorption of vitamin D-binding protein-bound 25D(3) absolutely requires receptor-mediated endocytosis via a receptor complex containing megalin, cubilin, and disabled-2 (Dab2), whereas an absence of megalin or its endocytic partners can lead to a marked urinary loss of 25D and severe vitamin D deficiency. Therefore, we hypothesized that reduced serum vitamin D status in NIDDM may be due to reduced expression of megalin and/or its endocytic partners and increased urinary excretion of protein-complexed 25D(3). In the present study, we utilized Zucker diabetic fatty Rats (ZDF) to demonstrate that renal reuptake of the 25D(3)-DBP complex was compromised in ZDF animals, which was reflected by a reduction in expression of megalin and Dab2. Moreover, serum levels of both 25D(3) and 1,25D(3) were reduced, and urinary 25D(3), 1,25D(3), and DBP excretion were elevated in the ZDF animals compared with their lean controls regardless of vitamin D levels in the diet. Taken together, these are the first reports to our knowledge that associate compromised renal reabsorption of the 25D(3)-DBP complex with expression of megalin and its endocytic partners in NIDDM, which in turn can lead to compromised vitamin D status.

A CONSTRAINT-BASED STOICHIOMETRIC MODEL OF THE STEROIDOGENIC NETWORK OF ZEBRAFISH (DANIO RERIO)

The metabolic process of steroidogenesis exhibits a complex biochemical network topology as the activity of various steroidogenic enzymes control cholesterol metabolism to steroid hormone derivatives. In this paper, a stoichiometric reconstruction of a sub-set of 65 reactions from the zebrafish (Danio rerio) steroidogenic network is presented and simulated using uniform reaction constraints. The reconstruction defined a set of 65 enzyme catalyzed reactions and 37 exchange or transport reactions for steroid metabolites. The reconstructed reactions were inclusive of cholesterol and androgen/estrogen metabolism. Biased (statement of network objective function) and un-biased (no statement of objective function) analyses were applied to identify network properties dependent on reaction stoichiometry. Random sampling of flux distributions through the network identified highly-correlated reaction sets that corresponded to the catalysis of steroid metabolites of physiological relevance. Subsequently, optimal flux distributions through network pathways were determined for the production of the three steroidogenic metabolites of: 11-deoxycorticosterone, testosterone and 17β-estradiol. Furthermore, flux variability analyses revealed and confirmed optimal network fluxes through physiologically feasible pathways. The stoichiometric dependence of reactions was also confirmed by conducting deletions of reactions utilized for the optimal production of 17β-estradiol. This paper demonstrates the potential application of constraint-based reconstruction and simulation techniques in enabling the construction of deterministic and predictive physiological models. This acknowledgement is poignant considering the susceptibility of the steroidogenic network to environmental and anthropogenic stressors.

Vitamin D homeostasis is disrupted in a type II diabetic rat model due to aberrant reuptake in kidney

Alterations of serum levels of the major circulating form of vitamin D (25‐hydroxycholecalciferol, 25D3) and its active hormone derivative (1,25‐dihydroxycholecalciferol, 1,25D3) have been associated with type II diabetes, but a mechanism for this occurrence has not been fully elucidated. In this study, we used the Zucker Diabetic Fatty (ZDF) rat model to characterize the role of renal reabsorption in vitamin D homeostasis in diabetic and non‐diabetic rats fed vitamin D sufficient (1000 IUs/kg), deficient (0 IUs/kg), and supplemented diets (10000 IUs/kg). Using ELISA, we analyzed serum and urinary 25D3, 1,25D3 and vitamin D binding protein (DBP) levels in 16 wk old ZDF and lean rats. Serum levels of both 25D3 and 1,25D3 were significantly lower in diabetic animals than their non‐diabetic controls fed either vitamin D deficient or sufficient diets. Furthermore, diabetic rats excreted greater proportions of DBP and 25D3 in urine than their respective controls. Vitamin D supplementation markedly increased serum 25D3 in diabetic rats, but had no effect on serum 1,25D3 or urinary excretion ratios. Taken together, these data suggest that compromised renal reabsorption of 25D3 and DBP may be a significant contributing mechanism leading to reduced vitamin D status in diabetes. Supported by USDA (Iowa State University Nutrition and Wellness Center SRPG to MR) and the Iowa State University Experiment Station.

A claudin‐4 modulator enhances the mucosal absorption of peptide

ObjectiveParacellular pathway is a principal route for absorption of biologics (peptides, proteins and nucleic acids); however, tight junctions (TJs) restrict passing of biologics across epithelium. Claudin is known as a key component in TJs‐barrier. We previously found that a claudin‐4 modulator, C‐terminal fragment of Clostridium perfringens enterotoxin (C‐CPE), enhanced jejunal absorption of dextran. In this study, we investigated whether a claudin‐4 modulator is a potent mucosal absorption‐enhancer of a peptide drug.MethodsMucosal absorption was evaluated by rat jejunal, pulmonary and nasal administration of human parathyroid hormone derivative, hPTH(1–34) and C‐CPEs. Plasma hPTH(1–34) levels were measured by RIA. Binding of C‐CPEs to claudin‐4 was assayed by ELISA and BIAcore analysis.ResultsPre‐treatment with C‐CPE enhanced mucosal absorption of hPTH(1–34). However, the jejunal and pulmonary absorption‐enhancing effects were not observed by co‐treatment with C‐CPE. The deletion of 10 amino acids at the N‐terminal of C‐CPE increased its solubility by 30‐fold. Moreover, the truncated C‐CPE bound to claudin‐4, modulated the TJ‐barrier and enhanced jejunal absorption of dextran. Co‐treatment of the truncated C‐CPE also enhanced jejunal and pulmonary absorption of hPTH(1–34).ConclusionClaudin‐4 modulator may be a promising absorption‐enhancer of a peptide drug.

A claudin-4 modulator enhances the mucosal absorption of a biologically active peptide

Biologics, such as peptides, proteins and nucleic acids, are emerging pharmaceuticals. Passage across the epithelium is the first step in the absorption of biologics. Tight junctions (TJ) function as seals between adjacent epithelial cells, preventing free movement of solutes across the epithelium. We previously found that modulation of a key TJ component, claudin-4, is a potent method to enhance jejunal absorption when we used dextran as a model drug and the C-terminal fragment of Clostridium perfringens enterotoxin (C-CPE) as a claudin-4 modulator. Here, we investigated whether the claudin-4 modulator enhances jejunal, nasal and pulmonary absorption of a biologics human parathyroid hormone derivative, hPTH(1-34). The claudin-4 modulator enhanced nasal but not jejunal and pulmonary absorption of hPTH(1-34). C-CPE is hydrophobic with low solubility of less than 0.3 mg/ml, but deletion of 10 amino acids at the N-terminal of C-CPE increased its solubility by 30-fold. Moreover, the N-terminal truncated C-CPE bound to claudin-4, modulated the TJ-barrier and enhanced jejunal absorption of dextran. The N-terminal-truncated C-CPE also enhanced jejunal and pulmonary absorption of hPTH(1-34). This report is the first to indicate that a claudin-4 modulator may be a promising enhancer of the jejunal, pulmonary and nasal absorption of a peptide drug.

Transport of thyroid hormones is selectively inhibited by 3-iodothyronamine

Thyroid hormone transporters are responsible for the cellular uptake of thyroid hormones, which is a prerequisite for their subsequent metabolism and action at nuclear thyroid hormone receptors. A recently discovered thyroid hormone derivative, 3-iodothyronamine (T(1)AM), has distinct biological effects that are opposite those of thyroid hormone. Here we investigate the effects of T(1)AM on thyroid hormone transporters using COS-1 cells transfected with the multispecific organic anion transporting polypeptides (OATPs) 1A2, 1B3, and 1C1, as well as the specific thyroid hormone transporters MCT8 and MCT10, and show that T(1)AM displays differential inhibition of T(3) and T(4) cellular uptake by these transporters. T(1)AM inhibits T(3) and T(4) transport by OATP1A2 with IC(50) values of 0.27 and 2.1 microM, respectively. T(4) transport by OATP1C1, which is thought to play a key role in thyroid hormone transport across the blood-brain barrier, is inhibited by T(1)AM with an IC(50) of 4.8 microM. T(1)AM also inhibits both T(3) and T(4) uptake via MCT8, the most specific thyroid hormone transporter identified to date, with IC(50) values of 95 and 31 microM, respectively. By contrast, T(1)AM has no effect on thyroid hormone transport by OATP1B3 and MCT10. Given that OATP1A2, OATP1C1, and MCT8 are all present in the brain, T(1)AM may play an important role in modulating thyroid hormone delivery and activity in specific target regions in the central nervous system.

3-Iodothyronamine (T1AM): a new chapter of thyroid hormone endocrinology?

3-Iodothyronamine (T(1)AM) is an endogenous thyroid hormone derivative with distinct biological effects that are largely opposite those of thyroid hormone. Administration of T(1)AM to rodents results in rapid and profound reduction in body temperature, heart rate, and metabolism. The structural similarities between thyroxine, T(1)AM, and monoamine neurotransmitters suggest an intriguing role for T(1)AM as both a neuromodulator and a hormone-like molecule that may constitute a part of thyroid hormone action. Several recent studies into its molecular mechanisms of action have shown that T(1)AM can target extracellular receptors such as the trace amine-associated receptors and the alpha(2A) adrenergic receptor, modulate the membrane transport of neurotransmitters, and serve as a substrate of specific membrane transport cellular uptake machinery. This review discusses recent T(1)AM studies, focusing on both the observed in vivo effects of T(1)AM administration and its actions at the molecular level.

D-Hormone Derivatives for the Treatment of Osteoporosis: From Alfacalcidol to Eldecalcitol

Many readers may have only a vague idea about vitamin D. This is made complicated, in part, because it is also expressed with suffixes such as vitamin D(2) or vitamin D(3). Otherwise the prefix of "active" is also occasionally used. Vitamin D is often referred to as an important nutrient for calcium intake, especially for growing children and the elderly. On the other hand, it serves as a therapeutic drug for osteoporosis and psoriasis. Recent studies have suggested an association with a number of diseases such as cancer, diabetes and Alzheimer's disease. The author has been involved in the research and development of vitamin D applications for over 25 years, and has often witnessed even world-class experts confuse the two roles - vitamin and hormone - of "substance" D. Assuming some readers are not familiar with vitamin D, D hormone or osteoporosis, an outline of vitamin D and D hormone is delineated with a particular focus on the treatment of osteoporosis. Furthermore, development of alfacalcidol as the first prodrug of D hormone (calcitriol) and eldecalcitol as a characteristic new D hormone derivative and basic relationship between calcemic activity and effect on bone in vitamin D (cholecalciferol), D hormone (calcitriol/alfacalcidol), and a new D hormone derivative (eldecalcitol) are introduced.

A native steroid hormone derivative triggers the resolution of inflammation.

Inflamed tissues produce both prostaglandins (PGs) and 7α-hydroxylated derivatives of native circulating 3β-hydroxysteroids. These 7α-hydroxysteroids are in turn transformed into 7β-hydroxylated epimers by 11β-hydroxysteroid dehydrogenase type 1 in the tissue. 7β-Hydroxy-epiandrosterone (7β-hydroxy-EpiA) affects PG production in two models of inflammation, dextran sodium sulfate-induced colitis in the rat and TNF-α-induced activation of PG production and PG synthase expression in cultured human peripheral blood monocytes (hPBMC). Treatment with 7β-hydroxy-EpiA led to a shift from high to low colonic PGE2 levels and from low to high 15-deoxy-Δ12-14-PGJ2 (15d-PGJ2) levels, together with changes in the expression of the respective PG synthases and resolution of colonic inflammation. Addition of 7β-hydroxy-EpiA to hPBMC also changed the expression of PG synthases and decreased PGE2 while increasing 15d-PGJ2 production. These effects were only observed with 7β-hydroxy-EpiA and not with 7α-hydroxy- or 7β-hydroxy-dehydroepiandrosterone (7α-hydroxy-DHEA and 7β-hydroxy-DHEA). 15d-PGJ2, which is the native ligand for peroxisome proliferator-activated receptor subtype γ, contributes to cell protection and to the resolution of inflammation. Our results therefore suggest that 7β-hydroxy-EpiA may facilitate inflammatory resolution by shifting PG production from PGE2 to PGD2 and 15d-PGJ2. The finding that 7β-hydroxy-EpiA was effective at nM concentrations, whereas the two structurally closely related hydroxysteroids 7α-hydroxy-DHEA and 7β-hydroxy-DHEA were inactive suggests that the effects of 7β-hydroxy-EpiA are specific to this steroid and may be mediated by a specific receptor.

Highly Sensitive and High-Throughput Analysis of Plant Hormones Using MS-Probe Modification and Liquid Chromatography–Tandem Mass Spectrometry: An Application for Hormone Profiling in Oryza sativa

We have developed a highly sensitive and high-throughput method for the simultaneous analysis of 43 molecular species of cytokinins, auxins, ABA and gibberellins. This method consists of an automatic liquid handling system for solid phase extraction and ultra-performance liquid chromatography (UPLC) coupled with a tandem quadrupole mass spectrometer (qMS/MS) equipped with an electrospray interface (ESI; UPLC-ESI-qMS/MS). In order to improve the detection limit of negatively charged compounds, such as gibberellins, we chemically derivatized fractions containing auxin, ABA and gibberellins with bromocholine that has a quaternary ammonium functional group. This modification, that we call 'MS-probe', makes these hormone derivatives have a positive ion charge and permits all compounds to be measured in the positive ion mode with UPLC-ESI-qMS/MS in a single run. Consequently, quantification limits of gibberellins increased up to 50-fold. Our current method needs <100 mg (FW) of plant tissues to determine phytohormone profiles and enables us to analyze >180 plant samples simultaneously. Application of this method to plant hormone profiling enabled us to draw organ distribution maps of hormone species in rice and also to identify interactions among the four major hormones in the rice gibberellin signaling mutants, gid1-3, gid2-1 and slr1. Combining the results of hormone profiling data with transcriptome data in the gibberellin signaling mutants allows us to analyze relationships between changes in gene expression and hormone metabolism.