Hormonal Form Of Vitamin D3 Research Articles

The Over-Irradiation Metabolite Derivative, 24-Hydroxylumister-ol3, Reduces UV-Induced Damage in Skin.

The hormonal form of vitamin D3, 1,25(OH)2D3, reduces UV-induced DNA damage. UV exposure initiates pre-vitamin D3 production in the skin, and continued UV exposure photoisomerizes pre-vitamin D3 to produce "over-irradiation products" such as lumisterol3 (L3). Cytochrome P450 side-chain cleavage enzyme (CYP11A1) in skin catalyzes the conversion of L3 to produce three main derivatives: 24-hydroxy-L3 [24(OH)L3], 22-hydroxy-L3 [22(OH)L3], and 20,22-dihydroxy-L3 [20,22(OH)L3]. The current study investigated the photoprotective properties of the major over-irradiation metabolite, 24(OH)L3, in human primary keratinocytes and human skin explants. The results indicated that treatment immediately after UV with either 24(OH)L3 or 1,25(OH)2D3 reduced UV-induced cyclobutane pyrimidine dimers and oxidative DNA damage, with similar concentration response curves in keratinocytes, although in skin explants, 1,25(OH)2D3 was more potent. The reductions in DNA damage by both compounds were, at least in part, the result of increased DNA repair through increased energy availability via increased glycolysis, as well as increased DNA damage recognition proteins in the nucleotide excision repair pathway. Reductions in UV-induced DNA photolesions by either compound occurred in the presence of lower reactive oxygen species. The results indicated that under in vitro and ex vivo conditions, 24(OH)L3 provided photoprotection against UV damage similar to that of 1,25(OH)2D3.

Lipopolysaccharide and cAMP modify placental calcitriol biosynthesis reducing antimicrobial peptides gene expression

Calcitriol, the hormonal form of vitamin D3 (VD), stimulates placental antimicrobial peptides expression; nonetheless, the regulation of calcitriol biosynthesis in the presence of bacterial products and its consequence on placental innate immunity have scarcely been addressed. We investigated how some bacterial products modify placental VD metabolism and its ability to induce antimicrobial peptides gene expression. Cultured human trophoblasts biosynthesized calcitriol only in the presence of its precursor calcidiol, a process that was inhibited by cyclic-AMP but stimulated by lipopolysaccharide (LPS). Intracrine calcitriol upregulated cathelicidin, S100A9, and β-defensins (HBDs) gene expression, while LPS further stimulated HBD2 and S100A9. Unexpectedly, LPS significantly repressed cathelicidin basal mRNA levels and drastically diminished calcidiol ability to induce it. Meanwhile, cyclic-AMP, which is used by many microbes to avoid host defenses, suppressed calcitriol biosynthesis, resulting in significant inhibition of most VD-dependent microbicidal peptides gene expression. While LPS stimulated calcitriol biosynthesis, cyclic-AMP inhibited it. LPS downregulated cathelicidin mRNA expression, whereas cyclic-AMP antagonized VD-dependent-upregulation of most antimicrobial peptides. These findings reveal LPS and cyclic-AMP involvement in dampening placental innate immunity, highlighting the importance of cyclic-AMP in the context of placental infection and suggesting its participation to facilitate bacterial survival.

In vivo response of the human epigenome to vitamin D: A Proof-of-principle study

In vitro cell culture studies showed that the hormonal form of vitamin D3, 1α,25-dihydroxyvitamin D3, significantly (p < 0.05) affects the human epigenome at thousands of genomic loci. Phase II of the VitDbol vitamin D intervention trial (NCT02063334) involved a proof-of-principle study of one individual, who was exposed three times every 28 days to an oral bolus (2000 μg) of vitamin D3. Blood samples were taken directly before each supplementation as well as one and two days after, chromatin was isolated from peripheral blood mononuclear cells without any further in vitro culture and at all nine time points epigenome-wide chromatin accessibility was assessed by applying FAIRE-seq (formaldehyde-assisted isolation of regulatory elements sequencing). The vitamin D3 bolus resulted in an average raise in 25-hydroxyvitamin D3 (25(OH)D3) serum concentration of 11.9 and 19.4 nM within one and two days, respectively. Consistently accessible chromatin was detected at 5205 genomic loci, the 853 most prominent of which a self-organizing map algorithm classified into early, delayed and non-responding genomic regions: 70 loci showed already after one day and 361 sites after two days significant (p < 0.0001) chromatin opening or closing. Interestingly, more than half of these genomic regions overlap with transcription start sites, but the change of chromatin accessibility at these sites has no direct effect on the transcriptome. Some of the vitamin D responsive chromatin sites cluster at specific loci within the human genome, the most prominent of which is the human leukocyte antigen region in chromosome 6. In conclusion, this study demonstrates that under in vivo conditions a rather minor rise in 25(OH)D3 serum levels is sufficient to result in significant changes at hundreds of sites within the epigenome of human leukocytes.

Calcium and phosphorus metabolism and nutrition of poultry: are current diets formulated in excess?

Calcium (Ca) and phosphorus (P) are important nutrients in poultry diet formulations. In the present review, we discuss recent advances in our understanding of the metabolism of Ca and P in poultry. Recent data are provided in support of the proposition that current poultry diets are formulated in excess for Ca and P. The quantities of Ca and P available for metabolism reflect rates of intestinal absorption, bone accretion and resorption, glomerular filtration, renal tubular reabsorption, and intestinal endogenous losses. Ca and P homeostasis is largely under endocrine control. Parathyroid hormone and the hormonal form of vitamin D3 are the two hormones credited with this role. However, a novel hormonal axis involving Fibroblast Growth Factor 23 and Klotho has been recently delineated, which, in conjunction with parathyroid hormone and vitamin D3, tightly regulates Ca and P homeostasis. Recent studies have suggested that current commercial diets for both broilers and layers contain excess Ca and P, the content of which could be reduced without affecting production or bird welfare. The challenge in reducing Ca and P concentrations in poultry diets is the uncertainty about what concentrations of Ca and P can be fed without compromising bird welfare. This is because there are limited data on the available P and Ca concentrations in poultry feedstuffs determined biologically. This is further complicated by the need for agreement on evaluation systems for evaluation of Ca and P bioavailability. We conclude that direct ileal or pre-caecal digestible Ca and P values are preferred.

The expression of RNA helicase DDX5 is transcriptionally upregulated by calcitriol through a vitamin D response element in the proximal promoter in SiHa cervical cells.

The DEAD box RNA helicase DDX5 is a multifunctional protein involved in the regulatory events of gene expression. Herein, we presented evidence indicating that DDX5 is transcriptionally upregulated by calcitriol, the hormonal form of vitamin D3. In silico analysis revealed the presence of two putative vitamin D response elements (VDREs) in the DDX5 promoter region. Using luciferase reporter assays, we demonstrated that the DDX5 promoter containing these putative VDREs significantly increased the luciferase activity in vitamin D receptor (VDR)-positive SiHa cells upon calcitriol treatment. Electrophoretic mobility shift assays showed the ability of VDR and retinoid X receptor to interact only with the most proximal VDRE, while chromatin immunoprecipitation analysis confirmed the occupancy of this VDRE by the VDR. Finally, we demonstrated that calcitriol significantly increased both DDX5 mRNA and protein in SiHa cells. In summary, this study shows that DDX5 gene is transcriptionally upregulated by calcitriol through a VDRE located in its proximal promoter. Given the importance of DDX5 as a master regulator of differentiation programs, our study suggests that the pro-differentiating properties of calcitriol may be related with the induction of DDX5.

Calcitriol increases Dicer expression and modifies the microRNAs signature in SiHa cervical cancer cells.

MicroRNAs play important roles in cancer biology. Calcitriol, the hormonal form of vitamin D3, regulates microRNAs expression in tumor cells. In the present study we asked if calcitriol would modify some of the components of the microRNA processing machinery, namely, Drosha and Dicer, in calcitriol-responsive cervical cancer cells. We found that calcitriol treatment did not affect Drosha mRNA; however, it significantly increased Dicer mRNA and protein expression in VDR-positive SiHa and HeLa cells. In VDR-negative C33-A cells, calcitriol had no effect on Dicer mRNA. We also found a vitamin D response element in Dicer promoter that interacts in vitro to vitamin D and retinoid X receptors. To explore the biological plausibility of these results, we asked if calcitriol alters the microRNA expression profile in SiHa cells. Our results revealed that calcitriol regulates the expression of a subset of microRNAs with potential regulatory functions in cancer pathways, such as miR-22, miR-296-3p, and miR-498, which exert tumor-suppressive effects. In summary, the data indicate that in SiHa cells, calcitriol stimulates the expression of Dicer possibly through the vitamin D response element located in its promoter. This may explain the calcitriol-dependent modulation of microRNAs whose target mRNAs are related to anticancer pathways, further adding to the various anticancer mechanisms of calcitriol.

Vitamin D3 regulates the formation and degradation of gap junctions in androgen-responsive human prostate cancer cells.

1α-25(OH)2 vitamin D3 (1-25D), an active hormonal form of Vitamin D3, is a well-known chemopreventive and pro-differentiating agent. It has been shown to inhibit the growth of several prostate cancer cell lines. Gap junctions, formed of proteins called connexins (Cx), are ensembles of cell-cell channels, which permit the exchange of small growth regulatory molecules between adjoining cells. Cell-cell communication mediated by gap junctional channels is an important homeostatic control mechanism for regulating cell growth and differentiation. We have investigated the effect of 1-25D on the formation and degradation of gap junctions in an androgen-responsive prostate cancer cell line, LNCaP, which expresses retrovirally-introduced Cx32. Connexin32 is expressed by the luminal and well-differentiated cells of normal prostate and prostate tumors. Our results document that 1-25D enhances the expression of Cx32 and its subsequent assembly into gap junctions. Our results further show that 1-25D prevents androgen-regulated degradation of Cx32, post-translationally, independent of androgen receptor (AR)-mediated signaling. Finally, our findings document that formation of gap junctions sensitizes Cx32-expressing LNCaP cells to the growth inhibitory effects of 1-25D and alters their morphology. These findings suggest that the growth-inhibitory effects of 1-25D in LNCaP cells may be related to its ability to modulate the assembly of Cx32 into gap junctions.

Vitamin D Metabolites and/or Analogs&amp;#58; Which D for Which Patient?

Numerous drugs with vitamin D activity are available for clinical use and it may not be easy for the nonspecialist to select the most suitable for the individual patient. In this paper we review the main characteristics of the available drugs and provide evidence about any potential specific clinical indications, with special emphasis on renal patients, in order to facilitate the optimal choice. Natural vitamin D products (i.e. those identical to natural metabolites) are first examined, followed by the most frequently used synthetic molecules (i.e. bioengineered molecules not-existing in nature), which are generally indicated as " analogs". Either cholecalciferol, ergocalciferol or calcifediol can be employed in subjects with normal renal function and in CKD stage 3-5 patients to correct vitamin D deficiency and improve, respectively, age- or growth-related bone disease and secondary hyperparathyroidism. Calcifediol can be considered more rapid and effective. In all cases, especially with increasing doses, the risk of hypercalcemia must be taken into account. Calcitriol, which can be regarded as the active hormonal form of vitamin D, has the most potent hypercalcemic effect in both normal and renal failure patients. In renal patients calcitriol is a potent inhibitor of parathyroid activity, but the risk of hypercalcemia, now regarded as harmful, is evident whenever pharmacologic doses are used. Alfacalcidol, requiring 25-hydroxylation to become the active hormonal form of vitamin D3, is prescribed in normal subjects to treat osteoporosis and in renal patients to cure hyperparathyroidism and renal bone disease. Doxercalciferol, transformed into the active hormonal form of vitamin D2 following 25-hydroxylation, is mostly studied in renal patients in whom it cures secondary hyperparathyroidism, possibly with a lower calcemic effect than calcitriol. Paricalcitol, a vitamin D2 analog not requiring activation, has been specifically developed to suppress PTH in renal patients with a limited calcemic effect. As such it is now regarded as a powerful drug useful to treat even severe cases of secondary hyperparathyroidism. Importantly, reno-protective and cardio-protective effects of this analog have been recently evaluated by means of randomized clinical trials in renal patients with partially positive renal effects and negative cardiac results, thus additional studies are needed for confirmation. 22-oxacalcitriol, a vitamin D3 analog with a limited calcemic effect available in Japan, is mostly used in renal patients affected by secondary hyperparathyroidism. The clinical activity of some vitamin D analogs is such that they can be employed in diseases like cancer and autoimmunity. The clinical activity of some vitamin D analogs is such that they can be employed in diseases like cancer and autoimmunity. In summary, available drugs with vitamin D like activity are not all the same either in terms of pharmacological actions, and side-effects. They have specific characteristics that may be useful to know in order to operate the best choice in the individual patient.

Research Resource: Whole Transcriptome RNA Sequencing Detects Multiple 1α,25-Dihydroxyvitamin D3-Sensitive Metabolic Pathways in Developing Zebrafish

The biological role of vitamin D receptors (VDR), which are abundantly expressed in developing zebrafish (Danio rerio) as early as 48 h after fertilization, and before the development of a mineralized skeleton and mature intestine and kidney, is unknown. We probed the role of VDR in developing zebrafish biology by examining changes in expression of RNA by whole transcriptome shotgun sequencing (RNA-seq) in fish treated with picomolar concentrations of the VDR ligand and hormonal form of vitamin D(3), 1α,25-dihydroxyvitamin D(3) [1α,25(OH)(2)D(3))].We observed significant changes in RNAs of transcription factors, leptin, peptide hormones, and RNAs encoding proteins of fatty acid, amino acid, xenobiotic metabolism, receptor-activator of NFκB ligand (RANKL), and calcitonin-like ligand receptor pathways. Early highly restricted, and subsequent massive changes in more than 10% of expressed cellular RNA were observed. At days post fertilization (dpf) 2 [24 h 1α,25(OH)(2)D(3)-treatment], only four RNAs were differentially expressed (hormone vs. vehicle). On dpf 4 (72 h treatment), 77 RNAs; on dpf 6 (120 h treatment) 1039 RNAs; and on dpf 7 (144 h treatment), 2407 RNAs were differentially expressed in response to 1α,25(OH)(2)D(3). Fewer RNAs (n = 481) were altered in dpf 7 larvae treated for 24 h with 1α,25(OH)(2)D(3) vs. those treated with hormone for 144 h. At dpf 7, in 1α,25(OH)(2)D(3)-treated larvae, pharyngeal cartilage was larger and mineralization was greater. Changes in expression of RNAs for transcription factors, peptide hormones, and RNAs encoding proteins integral to fatty acid, amino acid, leptin, calcitonin-like ligand receptor, RANKL, and xenobiotic metabolism pathways, demonstrate heretofore unrecognized mechanisms by which 1α,25(OH)(2)D(3) functions in vivo in developing eukaryotes.

Cell-Type-Specific Effects of Silibinin on Vitamin D-Induced Differentiation of Acute Myeloid Leukemia Cells Are Associated with Differential Modulation of RXRα Levels

Plant polyphenols have been shown to enhance the differentiation of acute myeloid leukemia (AML) cells induced by the hormonal form of vitamin D3 (1α,25-dihydroxyvitamin D3; 1,25D). However, how these agents modulate 1,25D effects in different subtypes of AML cells remains poorly understood. Here, we show that both carnosic acid (CA) and silibinin (SIL) synergistically enhancd 1,25D-induced differentiation of myeloblastic HL60 cells. However, in promonocytic U937 cells, only CA caused potentiation while SIL attenuated 1,25D effect. The enhanced effect of 1,25D+CA was accompanied by increases in both the vitamin D receptor (VDR) and retinoid X receptor alpha (RXRα) protein levels and vitamin D response element (VDRE) transactivation in both cell lines. Similar increases were observed in HL60 cells treated with 1,25D + SIL. In U937 cells, however, SIL inhibited 1,25D-induced VDRE transactivation concomitant with downregulation of RXRα at both transcriptional and posttranscriptional levels. These inhibitory effects correlated with the inability of SIL, with or without 1,25D, to activate the Nrf2/antioxidant response element signaling pathway in U937 cells. These results suggest that opposite effects of SIL on 1,25D-induced differentiation of HL60 and U937 cells may be determined by cell-type-specific signaling and transcriptional responses to this polyphenol resulting in differential modulation of RXRα expression.

Calcitriol stimulates prolactin expression in non-activated human peripheral blood mononuclear cells: Breaking paradigms

Calcitriol, the hormonal form of vitamin D3, exerts immunomodulatory effects through the vitamin D3 receptor (VDR) and increases prolactin (PRL) expression in the pituitary and decidua. Nevertheless, the effects of calcitriol upon lymphocyte PRL have not been evaluated. Therefore, we investigated calcitriol effects upon PRL in resting and phytohemagglutinin-activated human peripheral blood mononuclear cells (PBMNC) and Jurkat T lymphoma cells. Immunoblots showed constitutive expression of the 50-kDa VDR species in activated PBMNC and Jurkat cells, while a 75-kDa species was recognized in both resting and activated-PBMNC. Only in resting PBMNC calcitriol significantly stimulated PRL expression in a dose-dependent manner. The positive control CYP24A1, a highly VDR-responsive gene, was stimulated by calcitriol, effect that was stronger in resting than in activated-PBMNC (P<0.05), and without effect in Jurkat cells. Calcitriol upregulation of PRL and CYP24A1 was significantly inhibited by the VDR antagonist TEI-9647. EMSA showed that resting PBMNC contain a protein that binds to DR3-type VDRE. Cell activation reduced basal CYP24A1 while induced CYP27B1, VDR and pregnane X receptor (PXR) expression. In summary, calcitriol stimulated PRL and CYP24A1 gene expression in quiescent lymphocytes through a VDR-mediated mechanism. Our results suggest that the 75-kDa VDR species could be participating in calcitriol-mediated effects, and that activation induces factors such as PXR that restrain VDR transcriptional processes. This study supports the presence of a functional VDR in quiescent lymphocytes, providing evidence to reevaluate the VDR paradigm that assumes that lymphocytes respond to calcitriol only after activation. Altogether, our results offer new insights into the mechanisms whereby PRL is regulated in immune cells.

The vitamin D3 transcriptomic response in skin cells derived from the Atlantic bottlenose dolphin

The Atlantic bottlenose dolphin has attracted attention due to the evident impact that environmental stressors have taken on its health. In order to better understand the mechanisms linking environmental health with dolphin health, we have established cell cultures from dolphin skin as in vitro tools for molecular evaluations. The vitamin D3 pathway is one mechanism of interest because of its well established chemopreventative and immunomodulatory properties in terrestrial mammals. On the other hand, little is known of the physiological role of this molecule in aquatic animals. 1,25-dihydroxyvitamin D3 (1,25D3), the bioactive and hormonal form of vitamin D3, exerts its biological function by binding to the vitamin D receptor (VDR), a ligand-activated regulator of gene transcription. Therefore, we investigated the transcriptomic changes induced by 1,25D3 administration in dolphin skin cells. Identification of specific genes activated by 1,25D3 has provided clues to the physiological function of the vitamin D3 pathway in the dolphin. We found that exposure of the cells to 1,25D3 upregulated transactivation of a vitamin D-sensitive promoter. cDNA microarray analysis, using a novel dolphin array, identified specific gene targets within this pathway, and real-time PCR (qPCR) confirmed the enhanced expression of select genes of interest. These transcriptional changes correlated with an increase in VDR levels. This is the first report of the presence and activation of the vitamin D3 pathway in a marine mammal, and our experimental results demonstrate a number of similarities to terrestrial animals. Conservation of this pathway in the Atlantic bottlenose dolphin is consistent with the importance of nonclassic functions of vitamin D3, such as its role in innate immunity, similar to what has been demonstrated in other mammals.

Vitamin D3 induces pro-LL-37 expression in myeloid precursors from patients with severe congenital neutropenia

The innate immune system produces a number of effector molecules that are important for protection against bacterial infections. Neutrophils and antimicrobial peptides are major components of innate defense with the capacity of rapid bacterial killing. Patients with severe congenital neutropenia (SCN) experience recurrent and chronic infections despite recombinant G-CSF-mobilized neutrophils. We have shown previously that these neutrophils are deficient in that they lack the antimicrobial peptide LL-37. Here, we show that pro-LL-37 mRNA is not expressed in neutrophil precursors from patients with SCN, although the gene and promoter region for pro-LL-37, CAMP, does not display any mutations. The hormonal form of vitamin D3 [1,25(OH)2D3] induced the expression of pro-LL-37 in isolated neutrophil progenitors and in EBV-transformed B cells from patients with SCN, whereas all-trans retinoic acid only induced expression in transformed B cells. These results demonstrate that myeloid cells of patients with SCN can produce pro-LL-37, suggesting that other pathways are impaired.

Selective inhibition of cyclooxygenase‐2 (COX‐2) by 1α,25‐dihydroxy‐16‐ene‐23‐yne‐vitamin D3, a less calcemic vitamin D analog

Inducible cyclooxygenase-2 (COX-2) has been implicated to play a role in inflammation and carcinogenesis and selective COX-2 inhibitors have been considered as anti-inflammatory and cancer chemopreventive agents. 1alpha,25-dihydroxyvitamin D3 (1alpha,25(OH)2D3), the active hormonal form of vitamin D3 also has been considered to be a cancer chemopreventive agent in addition to its important role in maintaining calcium homeostasis. Based on these observations, we studied the direct effect of 1alpha,25(OH)2D3 and one of its less calcemic synthetic analogs, 1alpha,25(OH)2-16-ene-23-yne-D3 on the activity of both COX-1 and COX-2 in an in vitro enzyme assay. Preliminary data indicated that both 1alpha,25(OH)2D3 and 1alpha,25(OH)2-16-ene-23-yne-D3 inhibited selectively the activity of COX-2 with no effect on the activity of COX-1. Out of the two compounds, 1alpha,25(OH)2-16-ene-23-yne-D3 was found to be more effective with an IC50 of 5.8 nM. Therefore, the rest of the experiments were performed using 1alpha,25(OH)2-16-ene-23-yne-D3 only. 1alpha,25(OH)2-16-ene-23-yne-D3 inhibited the proliferation of lipopolysaccharide (LPS) stimulated mouse macrophage cells (RAW 264.7) with a reduction in the expression of COX-2 along with other inflammatory mediators like inducible nitric oxide synthase (iNOS) and interleukin-2 (IL-2). Furthermore, 1alpha,25(OH)2-16-ene-23-yne-D3 also inhibited carrageenan induced inflammation in an air pouch of a rat and effectively reduced the expression of COX-2, iNOS, and IL-2 in the tissues of the same air pouch. In both cases, 1alpha,25(OH)2-16-ene-23-yne-D3 did not show any effect on the expression of COX-1. In summary, our results indicate that 1alpha,25(OH)2-16-ene-23-yne-D3, a less calcemic vitamin D analog, exhibits potent anti-inflammatory effects and is a selective COX-2 inhibitor.

Comparative therapeutic effects of orally administered 1,25-dihydroxyvitamin D3 and 1alpha-hydroxyvitamin D3 on type-1 diabetes in non-obese diabetic mice fed a normal-calcaemic diet

Frequent injections of the hormonal form of vitamin D(3), 1,25 dihydroxyvitamin D(3) (1,25D3) reportedly inhibits autoimmune type 1 diabetes (T1D) in non-obese diabetic (NOD) mice by correcting some of the abnormalities in antigen-presenting cells which contribute the development of pathogenic T cell responses. This route of administration greatly elevates the levels of these compounds in the bloodstream for hours after treatment, which requires mice to be fed diets formulated to contain much reduced levels of Ca to avoid the toxic effects of hypercalcaemia. In the current work, we demonstrate that feeding 1,25D3 or its synthetic precursor, 1alpha(OH) vitamin D(3) (1alphaD3), as part of a T1D supportive chow diet containing normal levels of Ca, is an effective means of reducing the incidence of disease in NOD mice, but the doses required for protection elicited hypercalcaemia. However, T1D protection elicited by D3 analogue feeding appears, at least partially, to have an immunological basis, as splenic T cells from treated mice had a decreased capacity to adoptively transfer disease. Protection is associated with an increased proportion of T cells with CD4+ forkhead box P3+ regulatory phenotype within the islet infiltrate of treated animals. The 1alphaD3 precursor is converted rapidly to the active 1,25D3 isoform in vivo. However, feeding the 1alphaD3 analogue elicited stronger T1D protection than the 1,25D3 compound, but also induced more severe hypercalcaemia. In future, the dietary supplementation of novel low-calcaemic D3 analogues may enable their continuous delivery at levels that inhibit T1D development in susceptible humans consuming normal levels of Ca.

Overview of general physiologic features and functions of vitamin D

Vitamin D3 is a prohormone produced in skin through ultraviolet irradiation of 7-dehydrocholesterol. It is biologically inert and must be metabolized to 25-hydroxyvitamin D3 in the liver and then to 1alpha,25-dihydroxyvitamin D3 in the kidney before function. The hormonal form of vitamin D3, ie, 1alpha,25-dihydroxyvitamin D3, acts through a nuclear receptor to carry out its many functions, including calcium absorption, phosphate absorption in the intestine, calcium mobilization in bone, and calcium reabsorption in the kidney. It also has several noncalcemic functions in the body. This overview provides a brief description of the physiologic, endocrinologic, and molecular biologic characteristics of vitamin D. It also provides information on new selective analogs of 1alpha,25-dihydroyvitamin D3 for therapy.

Cutting edge: 1,25-dihydroxyvitamin D3 is a direct inducer of antimicrobial peptide gene expression.

The hormonal form of vitamin D(3), 1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)), is an immune system modulator and induces expression of the TLR coreceptor CD14. 1,25(OH)(2)D(3) signals through the vitamin D receptor, a ligand-stimulated transcription factor that recognizes specific DNA sequences called vitamin D response elements. In this study, we show that 1,25(OH)(2)D(3) is a direct regulator of antimicrobial innate immune responses. The promoters of the human cathelicidin antimicrobial peptide (camp) and defensin beta2 (defB2) genes contain consensus vitamin D response elements that mediate 1,25(OH)(2)D(3)-dependent gene expression. 1,25(OH)(2)D(3) induces antimicrobial peptide gene expression in isolated human keratinocytes, monocytes and neutrophils, and human cell lines, and 1,25(OH)(2)D(3) along with LPS synergistically induce camp expression in neutrophils. Moreover, 1,25(OH)(2)D(3) induces corresponding increases in antimicrobial proteins and secretion of antimicrobial activity against pathogens including Pseudomonas aeruginosa. 1,25(OH)(2)D(3) thus directly regulates antimicrobial peptide gene expression, revealing the potential of its analogues in treatment of opportunistic infections.

The pleiotropic actions of vitamin D.

General knowledge of the role of vitamin D3 in human physiology has been shaped by its discovery as a preventive agent of nutritional rickets, a defect in bone development due to inadequate uptake of dietary calcium. Studies on the function of the hormonal form of vitamin D3, 1alpha,25-dihydroxyvitamin D3, have been greatly accelerated by the molecular cloning and structural analysis of the vitamin D3 receptor, which is a ligand-activated regulator of gene transcription. Molecular genetic techniques including genomics have helped reveal that 1alpha,25-dihydroxyvitamin D3 can control more than calcium homeostasis. It has widespread effects on cellular differentiation and proliferation, and can modulate immune responsiveness, and central nervous system function. Moreover, accumulating epidemiological and molecular evidence suggests that 1alpha,25-dihydroxyvitamin D3 acts as a chemopreventive agent against several malignancies including cancers of the prostate and colon. Here, we survey the most-recent findings and discuss their implications for the potential therapeutic uses of vitamin D analogues.

92. Calcitriol and its analog down-regulates αvβ3 integrin expression and suppresses the tumorigenesis of lewis lung carcinoma (LLC) cell line

Aim Calcitriol (CAL) [1,25-(OH)203], active hormonal form of vitamin D3, reveals antitumor activity both in vitro and in vivo. CAL may activate the β3-integrin subunit gene and promote the plasma membrane appearance of αvβ3 on osteociast precursors and on HL-60 leukemia cells. In this study we evaluated the effect of calcitriol and its new analogue (PRI-2191), synthesized in order to avoid hypercalcemic effect of calcitriol, on αvβ3 integrin-dependent mouse lung cancer invasiveness. Material and Methods The influence of CAL or PRI-2191 on the proliferation, adhesion properties and celi cycle of mouse LLC cells were tested by FACS analysis, adhesion and antiproliferative assays. Monoclonal antibodies and flow cytometry were applied for evaluation of integrins expression. Antitumor activity in vivo was examined in the model of subcutaneously growing tumors in C57BI/6 mice. Results We have shown, that LLC cells express a high level of αvβ3 (but not of αllbβ3) integrin and this expression was signiticantly reduced by the in vitro treatment with CAL or PRI-2191. Both compounds inhibited (in about 60%) LLC cells proliferation after 72 h of in vitro treatment. Moreover, both agents augmented adhesion of LLC cells to collagen (ligand tor allb integrin subunit) and diminished adhesion to fibrinogen (ligand for β3 integrin subunits). Further, in vitro incubation of LLC cells with both agents retarded the growth of subcutaneous tumors in mice. n all experiments, biological activity of PRI-2191 was higher than this of CAL, what is in accordance with our previous experiments, showing higher antiproliterative, lower calcemic activities and lower lethal toxicity of new analog as compared to parental drug. Conclusions Our data demonstrate that new analog of calcitriol, PRI-2191, is a good candidate for an antitumor drug, especially in the treatment of cancers in which the invasive potential is, atleast in part, dependent on expression of β3 integrins.

Effects of elicitors and Ca2+ deprivation on the levels of sterols and 1α,25-dihydroxy vitamin D3 in cell cultures of Solanum malacoxylon.

1α,25-dihydroxy vitamin D3 [1α,25(OH)2D3], the hormonal form of vitamin D3 that is essential for the maintenance of calcium-phosphorus homeostasis in birds and mammals, has been found in several plants. In order to study the metabolic role of 1α,25(OH)2D3 in plant cells, we monitored the changes of cellular levels of 1α,25(OH)2D3 in cell suspension cultures of Solanum malacoxylon Sendt. under different experimental conditions. We also measured the levels of cholesterol, the assumed precursor of vitamin D3, and other end-product sterols such as β-sitosterol, stigmasterol and campesterol. It was found that when cells are incubated in a calcium-deprived medium, the level of 1α,25(OH)2D3 increases markedly within a few hours. Treatment with methyl jasmonate (MeJ), a linolenic acid-derived compound that accumulates during wounding stress, was found to reduce the cellular level of 1α,25(OH)2D3 and to promote its secretion into the culture medium. After treatment with cellulysin, a cellulase of fungal origin, the metabolite was no longer detectable, either in the medium or in the cells. Calcium deprivation in the medium and treatment with the elicitors MeJ and cellulase lowered the cellular level of all the sterols tested. The possible role of 1α,25(OH)2D3 is discussed in light of these results.