Hypercalcaemia of malignancy: a case of vitamin-D-mediated hypercalcaemia in lymphoma.

Hypercalcaemia of malignancy (HCM) is a paraneoplastic syndrome that often portends a poor prognosis. We present an extremely rare (<1%) case of HCM due to extrarenal calcitriol (1,25-(OH)2D) production in a patient with splenic marginal zone lymphoma. A man in his 80s presented with a 3-week history of fatigue, unsteadiness and abdominal pain, and new findings of anaemia, kidney injury and hypercalcaemia. Laboratory evaluation, bone marrow biopsy and positron emission tomography/computed tomography (PET/CT) confirmed the diagnosis of splenic marginal zone lymphoma which produced calcitriol (1,25-(OH)2D3), causing the patient's hypercalcaemia.

Unusual behaviour of an unusual tumour: calcitriol-induced hypercalcaemia in metastatic oesophageal neuroendocrine carcinoma.

Hypercalcaemia in malignancy is most commonly caused by paraneoplastic secretion of parathyroid hormone-related protein or osteolytic metastases. Very rarely (<1% of cases), the mechanism behind increased serum calcium is increased production of calcitriol (1,25-dihydroxyvitamin D) and even rarer is the occurrence of this phenomenon in solid malignancies, with few such instances reported in the literature. We present a case of a neuroendocrine malignancy originating in the oesophagus associated with calcitriol-induced hypercalcaemia, a phenomenon that has not been previously described. We review the pathophysiology of calcitriol-induced hypercalcaemia and previously reported cases of solid tumours with this presentation.

Vitamin D3-mediated resistance to a multiple sclerosis model disease depends on myeloid cell 1,25-dihydroxyvitamin D3 synthesis and correlates with increased CD4+ T cell CTLA-4 expression.

Microglial cell activation is the earliest biomarker of the inflammatory processes that cause central nervous system (CNS) lesions in multiple sclerosis. We hypothesized that 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) production by activated microglia and macrophages in the CNS inhibits these inflammatory processes. To test this hypothesis, we targeted the Cyp27b1 gene specifically in myeloid cells, then analyzed the influence of disrupted myeloid cell 1,25-(OH)2D3 synthesis on vitamin D3-mediated resistance to experimental autoimmune encephalomyelitis (EAE). Myeloid cell 1,25-(OH)2D3 synthesis was essential for vitamin D3-mediated EAE resistance. Increased CTLA-4 expression in the CNS-infiltrating CD4+ Tconv and Treg cells and decreased splenic B cell CD86 expression correlated with resistance. These new data provide solid support for the view that vitamin D3 reduces MS risk in part through a mechanism involving myeloid cell 1,25-(OH)2D3 production and CTLA-4 upregulation in CNS-infiltrating CD4+ T cells. We suggest that CTLA-4 serves as a vitamin D3-regulated immunological checkpoint in multiple sclerosis prevention.

Off­target effect of imatinib and nilotinib on human vitamin D3 metabolism.

Prolonged treatment with tyrosine kinase inhibitors (TKI) including imatinib (IMA) or nilotinib (NIL), induces severe disturbances of bone metabolism in patients with chronic myeloid leukaemia. As vitamin D3 (VD3) is involved in the complex cycle of bone remodelling, the present study investigated in vitro, the influence of IMA and NIL on VD3 metabolism i) in HaCaT cells and ii) in cultured outer root sheath keratinocytes (ORS­KC) from hair follicles of IMA treated children. Cells were incubated in the presence of IMA or NIL. Concomitantly, specific inhibitors were applied to analyze the inhibition of the VD3 processing cytochrome P450 isoenzyme family by TKIs. In vitro, IMA and NIL significantly impaired the production of calcitriol in HaCaT and cultured ORS­KC cells from hair follicles of IMA treated children. For NIL, this inhibitory effect demonstrated a 4­fold increase. In HaCaT and ORS­KC, application of specific CYP450 inhibitors revealed that CYP27B1 was impaired by IMA and NIL leading to an intracellular accumulation of calcidiol. However, during TKI treatment, KC of IMA treated children revealed no differences in calcidiol and calcitriol levels. In conclusion, IMA and NIL interfere with the vitamin D3 cascade due to their metabolism by CYP27B1.

Endogenous Calcitriol Synthesis Controls the Humoral IgE Response in Mice.

The vitamin D receptor participates in the control of IgE class-switch recombination in B cells. The physiologic vitamin D receptor agonist, 1,25(OH)2D3 (calcitriol), is synthesized by the essential enzyme 25-hydroxyvitamin D3-1α-hydroxylase (CYP27B1), which can be expressed by activated immune cells. The role of endogenous calcitriol synthesis for the regulation of IgE has not been proven. In this study, we investigated IgE-responses in Cyp27b1-knockout (KO) mice following sensitization to OVA or intestinal infection with Heligmosomoides polygyrus Specific Igs and plasmablasts were determined by ELISA and ELISpot, Cyp27b1 expression was measured by quantitative PCR. The data show elevated specific IgE and IgG1 concentrations in the blood of OVA-sensitized Cyp27b1-KO mice compared with wild-type littermates (+898 and +219%). Accordingly, more OVA-specific IgG1-secreting cells are present in spleen and fewer in the bone marrow of Cyp27b1-KO mice. Ag-specific mechanisms are suggested as the leucopoiesis is in general unchanged and activated murine B and T lymphocytes express Cyp27b1 Accordingly, elevated specific IgE concentrations in the blood of sensitized T cell-specific Cyp27b1-KO mice support a lymphocyte-driven mechanism. In an independent IgE-inducing model, i.e., intestinal infection with H. polygyrus, we validated the increase of total and specific IgE concentrations of Cyp27b1-KO compared with wild-type mice, but not those of IgG1 or IgA. We conclude that endogenous calcitriol has an impact on the regulation of IgE in vivo. Our data provide genetic evidence supporting previous preclinical and clinical findings and suggest that vitamin D deficiency not only promotes bone diseases but also type I sensitization.

Women with Recurrent Miscarriage Have Decreased Expression of 25-Hydroxyvitamin D3-1α-Hydroxylase by the Fetal-Maternal Interface.

BACKGROUND: Effects of vitamin D deficiency in pregnancy have been associated with some adverse pregnancy outcomes. The 25-hydroxyvitamin D3-1α-hydroxylase (CYP27B1) is integral to the vitamin D metabolic pathway. The enzyme catalyzes localized conversion of pro-hormone 25-hydroxyvitamin D3 to active 1,25-dihydroxyvitamin D3. Our aim was to investigate the expression of CYP27B1 at the fetal-maternal interface in the first trimester pregnancy and to determine whether CYP27B1 was associated with recurrent miscarriage (RM). METHODS: Expressions of CYP27B1 mRNA and protein in villi and decidua from 20 women undergoing primary miscarriage, 20 women with RM and 20 women with normal pregnancy were evaluated by western blot, and quantitative real-time PCR. The co-localization of CYP27B1 and certain cytokines including IL-10, IFN-γ, TNF-α, and IL-2 expression were examined using immunohistochemistry and confocal microscopy. RESULTS: Women with RM had a significantly lower expression of CYP27B1 mRNA and protein in villous and decidual tissues compared with the normal pregnant women (P = 0.000 in villus, P = 0.002 in decidua for mRNA; P = 0.036 in villus, P = 0.007 in decidua for protein.). Compared with the normal pregnancy, immunostaining for CYP27B1 was significantly decreased in villous trophoblasts and decidual glandular epithelial cells in RM women. No significant differences in the localization of CYP27B1, IL-10, IFN-γ, TNF-α, and IL-2 expression were identified between the normal pregnant and RM women. CONCLUSIONS: Women with RM have a lower level of CYP27B1 expression in chorionic villi and decidua compared with normal pregnant women, suggesting that reduced CYP27B1 expression may be associated with RM. The consistent localization of CYP27B1 and IL-10, IFN-γ, TNF-α, and IL-2 expression in villous and decidual tissues suggests the importance of the local production of 1,25(OH)2D3 at the fetal-maternal interface to regulate cytokine responses.

Tumoral Vitamin D Synthesis by CYP27B1 1-α-Hydroxylase Delays Mammary Tumor Progression in the PyMT-MMTV Mouse Model and Its Action Involves NF-κB Modulation.

Biologically active vitamin D (1,25-dihydroxycholecalciferol or 1,25(OH)2D) is synthetized from inactive prohormone 25-hydroxycholecalciferol (25(OH)D) by the enzyme CYP27B1 1-α-hydroxylase in kidney and several extrarenal tissues including breast. Although the development of breast cancer has been linked to inadequate vitamin D status, the importance of bioactive vitamin D production within tumors themselves is not fully understood. To investigate the role of tumoral vitamin D production in mammary epithelial cell progression to breast cancer, we conducted a Cre-loxP-mediated Cyp27b1 gene ablation in the mammary epithelium of the polyoma middle T antigen-mouse mammary tumor virus (PyMT-MMTV) mouse breast cancer model. Targeted ablation of Cyp27b1 was accompanied by significant acceleration in initiation of spontaneous mammary tumorigenesis. In vivo, cell proliferation, angiogenesis, cell cycle progression, and survival markers were up-regulated in tumors by Cyp27b1 ablation, and apoptosis was decreased. AK thymoma (AKT) phosphorylation and expression of several components of nuclear factor κB (NF-κB), integrin, and signal transducer and activator of transcription 3 (STAT3) signaling pathways were increased in Cyp27b1-ablated tumors compared with nonablated controls. In vitro, 1,25(OH)2D treatment induced a strong antiproliferative action on tumor cells from both ablated and nonablated mice, accompanied by rapid disappearance of NF-κB p65 from the nucleus and segregation in the cytoplasm. In contrast, treatment with the metabolic precursor 25(OH)D was only effective against cells from nonablated mice. 25(OH)D did not inhibit growth of Cyp27b1-ablated cells, and their nuclear NF-κB p65 remained abundant. Our findings demonstrate that in-tumor CYP27B1 1-α-hydroxylase activity plays a crucial role in controlling early oncogene-mediated mammary carcinogenesis events, at least in part by modulating tumoral cell NF-κB p65 nuclear translocation.

[Phosphate metabolism and iron deficiency].

Autosomal dominant hypophosphatemic rickets(ADHR)is caused by gain-of-function mutations in FGF23 that prevent its proteolytic cleavage. Fibroblast growth factor 23(FGF23)is a hormone that inhibits renal phosphate reabsorption and 1,25-dihydroxyvitamin D biosynthesis. Low iron status plays a role in the pathophysiology of ADHR. Iron deficiency is an environmental trigger that stimulates FGF23 expression and hypophosphatemia in ADHR. It was reported that FGF23 elevation in patients with CKD, who are often iron deficient. In patients with nondialysis-dependent CKD, treatment with ferric citrate hydrate resulted in significant reductions in serum phosphate and FGF23.

Inhibitory effects of imatinib on vitamin D3 synthesis in human keratinocytes.

Chronic myeloid leukemia (CML) is a myeloproliferative disease characterized by the presence of the BCR­ABL1 fusion gene, a constitutively active, oncogenic tyrosine kinase that is responsible for the clinical features of CML. Tyrosine kinase inhibitors, such as imatinib, have markedly altered the treatment of CML. However, tyrosine kinase inhibitors are associated with side effects on bone metabolism, in adult and pediatric patients. Vitamin D3 is involved in the complex cycle of bone remodeling, therefore the present study aimed to investigate the influence of imatinib on vitamin D3 metabolism in the HaCaT human keratinocyte cell line, using commercially available enzyme assays. Imatinib was shown to significantly reduce the production of calcidiol and calcitriol. Based on interaction studies of imatinib with the cytochrome P450 (CYP450) inhibitors VID400 and ketoconazole, it is proposed that imatinib may interfere with the vitamin D3 cascade due to its metabolism by CYP27B1, which is involved in vitamin D3 metabolism.

Vitamin D receptor agonists' anti-inflammatory properties.

One century after its discovery, vitamin D has been shown to be, in fact, a pleiotropic steroid hormone, which, besides regulation of calcium homeostasis and bone turnover, has antiproliferative, prodifferentiation, antibacterial, immunomodulatory, and anti-inflammatory properties in various cells and tissues. D hormone (1α,25(OH)2 D), regulated in an endocrine, autocrine, and paracrine manner, must be bound to the specific nuclear vitamin D receptor (VDR) to exert epigenetic and genetic effects, acting as a connection between extracellular stimuli and genomic responses of the cells. Since only high doses of hormone, provoking hypercalcemia, can achieve immunomodulatory effects, more than 3000 VDR agonists have been synthesized. Numerous experimental trials have been performed in animal models, evidencing the preventive and therapeutic potential of VDR agonists for chronic inflammatory diseases and cancer. Considering the selective anti-inflammatory effects of VDR agonists compared to glucocorticoids, sparing microbicidal functions, the fear of hypercalcemia as their only frequent side effect becomes a questionable reason for the lack of clinical studies.

Vitamin D in liver diseases: from mechanisms to clinical trials.

Traditionally regarded as a typical vitamin regulating calcium and phosphorus homeostasis, vitamin D is now discovered as a highly versatile molecule with emerging roles in immunity, cancer, infectious diseases, fibrosis, fatty liver diseases, and alcoholic liver diseases. A large body of clinical evidence has demonstrated the prevalence and risks of vitamin D deficiency in various chronic diseases. Biologically active vitamin D, 1,25-dihydroxylvitamin D3, is synthesized in two distinct systems. In addition to the classic two-step hydroxylation in the liver and kidneys, 1,25-dihydroxylvitamin D3 can also be produced locally by immune cells in response to infection. The bioactive vitamin D generated in these two pools apparently functions differently: while the former facilitates calcium adsorption and homeostasis, the latter confers immune regulation. The immune regulatory functions of vitamin D are demonstrated by induction of antimicrobial peptides, suppression of innate immune response, induction of Th2 cytokines, and stimulation of T-regulatory T cells. Vitamin D deficiency or insufficiency is overwhelmingly associated with viral hepatitis, cirrhosis, and fatty liver diseases. Recent clinical trials have shown that vitamin D supplements significantly enhance the efficacy of interferon plus ribavirin therapy through sustained virological response. A recent study showed that 25-dihydroxyvitamin D rather than 1,25-dihydroxyvitamin D could directly suppress hepatitis C virus assembly. Moreover, clinical evidence has shown that vitamin D deficiency is associated with alcoholic and non-alcoholic fatty liver diseases. In this review, we highlight some recent advances in vitamin D researches and clinical trails.

Antibiotic-like actions of vitamin D.

Vitamin D is a secosteroid hormone that has expanding importance for a healthy lifestyle and disease prevention. A multitude of studies have highlighted that vitamin D acts not only in bone and calcium homeostasis but is critically important for human immunity. The discovery that the storage form of vitamin D (25-hydroxyvitamin D3) can be locally converted to the active form (1,25-hydroxyvitamin D3) in immune cells, epithelial cells and numerous other non-renal tissues highlights the importance of maintaining sufficient stores. When responding to a specific external stimulus, like bacterial invasion, intracrine synthesis of active vitamin D has the ability to regulate gene expression providing a specific response and directing cellular actions. These responses include the generation of antimicrobial peptides with production of these peptides dependent on vitamin D status. Vitamin D deficiency is associated with an increased rate of infection. This paper highlights the antibiotic like actions of vitamin D and importance of vitamin D sufficiency.

[Vitamin D for prevention of diseases?]. / Vitamin D zur Prävention von Erkrankungen?

Vitamin D3 shows a multitude of possible preventive effects in various diseases. Calcitriol, the biologically active form of vitamin D3, affects not only bone metabolism but also acts on the renal renin secretion, the pancreatic insulin production in the beta cells, growth and proliferation of smooth and cardiac muscle cells and the function of lymphocytes and macrophages. Although the human body can synthesise vitamin D3 itself, vitamin D deficiency is common in the German population. Numerous trials studied the association between vitamin D deficiency and different diseases. It is known that even mild forms of vitamin D deficiency increase the risk for cardiovascular diseases or diabetes mellitus. Furthermore, an association with cancer such as pancreatic or colorectal cancer was observed. This is attributed to the influence of vitamin D on cell differentiation, angiogenesis, DNA repair mechanisms and the transcription of numerous genes. In addition, effects of vitamin D deficiency in diseases such as Parkinson's disease, multiple sclerosis and autoimmune diseases are discussed. However, up to now the level of evidence of all these observations is low. There are missing confirmatory randomized controlled trials. Noting the possible preventive effects of vitamin D, a moderate exposure to sunlight to increase vitamin D synthesis can be recommended. Even a controlled supplementation of vitamin D in patients with vitamin D deficiency is considered as reasonable. However, an uncritical substitution of high-dose vitamin D should be avoided because of the risk of hypercalcaemia.

Colonic vitamin D metabolism: implications for the pathogenesis of inflammatory bowel disease and colorectal cancer.

In epidemiological studies serum levels below 30 nM of 25-OHD(3), the precursor of the active vitamin D metabolite 1,25-(OH)(2)D(3), were consistently associated with incidence of colorectal cancer. The active vitamin D metabolite possesses antimitotic, prodifferentiating and proapoptotic capacity in vivo and in vitro. The intestinal autocrine/paracrine vitamin D system, which is the main source of local 1,25-(OH)(2)D(3) plays a critical role in maintaining both mucosal immunity and normal growth of epithelial cells. It has been hypothesized that the VDR-mediated signaling antagonizing TNF-α and IL-6 receptor-activated pro-inflammatory and proliferative intracellular pathways, may prevent development of IBD and colitis-associated colorectal cancer. Conversely, any situation that impairs the efficiency of the 1,25-(OH)(2)D(3)/VDR signaling system at the level of the gut mucosa, e.g. vitamin D insufficiency, may increase risk for the development of IBD and colorectal cancer. Therefore, not only adequate serum levels of the precursor 25-OHD(3) are essential, but also optimal expression of the 1α-hydroxylating enzyme CYP27B1. The 1,25-(OH)(2)D(3) catabolizing hydroxylase CYP24A1 is increasingly expressed during colon cancer progression, indicating that colonocytes are released from normal growth control by the steroid hormone. Securing adequate levels of calcitriol by inhibition of catabolism and support of 1α-hydroxylation by calcium, phytoestrogens and folate could be a valid approach to control, at least in part, IBD and CRC pathogenesis.

Expression profiling of vitamin D receptor in placenta, decidua and ovary of pregnant mice.

OBJECTIVES: The presence of vitamin D receptor (VDR) and the identification of localized vitamin D3 synthesis in placenta and decidua implicate the importance of vitamin D3 in reproductive function. There is, however, no data on the expression profile of VDR in the mouse placenta and endometrium throughout the pregnancy period. STUDY DESIGN: In the present work expression of VDR in reproductive tissues of pregnant mice at different gestational phases has been addressed. Expression of VDR was determined by semi-quantitative RT-PCR, Western blotting and immunohistochemistry. RESULTS: The results showed that VDR mRNA and protein were expressed in decidua, placenta and ovary throughout the pregnancy. VDR gene expression in placenta was significantly elevated in late pregnancy when compared to that of mid pregnancy. Additionally, VDR expression level in decidua rose significantly as pregnancy progressed from early to mid stages. VDR expression in decidua of pregnant mice was higher in comparison to endometrium of non-pregnant mice. Immunohistochemical analysis revealed that VDR protein is consistently expressed by luminal and glandular epithelial cells of decidua, giant cells, glycogen rich cells and labyrinth cells of placenta and by almost all follicular cell types of ovary. Surveying the expression of VDR at the protein level by Western blotting confirmed PCR results. CONCLUSION: It seems that expression of VDR in reproductive organs is finely tuned during pregnancy indicating its eminent role in reproductive biology.

Vitamin D: an innate antiviral agent suppressing hepatitis C virus in human hepatocytes.

UNLABELLED: Vitamin D supplementation was reported to improve the probability of achieving a sustained virological response when combined with antiviral treatment against hepatitis C virus (HCV). Our aim was to determine the in vitro potential of vitamin D to inhibit HCV infectious virus production and explore the mechanism(s) of inhibition. Here we show that vitamin D(3) remarkably inhibits HCV production in Huh7.5 hepatoma cells. These cells express CYP27B1, the gene encoding for the enzyme responsible for the synthesis of the vitamin D hormonally active metabolite, calcitriol. Treatment with vitamin D(3) resulted in calcitriol production and induction of calcitriol target gene CYP24A1, indicating that these cells contain the full machinery for vitamin D metabolism and activity. Notably, treatment with calcitriol resulted in HCV inhibition. Collectively, these findings suggest that vitamin D(3) has an antiviral activity which is mediated by its active metabolite. This antiviral activity involves the induction of the interferon signaling pathway, resulting in expression of interferon-ß and the interferon-stimulated gene, MxA. Intriguingly, HCV infection increased calcitriol production by inhibiting CYP24A1 induction, the enzyme responsible for the first step in calcitriol catabolism. Importantly, the combination of vitamin D(3) or calcitriol and interferon-α synergistically inhibited viral production. CONCLUSION: This study demonstrates for the first time a direct antiviral effect of vitamin D in an in vitro infectious virus production system. It proposes an interplay between the hepatic vitamin D endocrine system and HCV, suggesting that vitamin D has a role as a natural antiviral mediator. Importantly, our study implies that vitamin D might have an interferon-sparing effect, thus improving antiviral treatment of HCV-infected patients.

Vitamin D metabolism in mammary gland and breast cancer.

1α,25-Dihydroxycholecalciferol (1,25D) mediates growth inhibition and terminal differentiation in mammary epithelial cells via interaction with the vitamin D receptor (VDR). This review focuses on the concept that cells in the mammary gland express the vitamin D metabolizing enzyme CYP27B1 which converts the circulating vitamin D metabolite 25D to the active metabolite 1,25D. In support of this concept, CYP27B1 is developmentally regulated in mouse mammary gland, with highest levels found during pregnancy and lactation. In addition, human mammary cells cultured from normal breast tissue express VDR, CYP27B1 and the megalin-cubilin complex that facilitates internalization of 25D complexed with the vitamin D binding protein. When incubated with physiological concentrations of 25D, human mammary cells synthesize 1,25D in sufficient quantities to mediate growth inhibition. However, emerging evidence suggests that deregulation of VDR and CYP27B1 occurs during cancer development and contributes to abrogation of the tumor suppressive effects triggered by 25D.

[Non phosphocalcic actions of vitamin D]. / Effets non osseux de la vitamine D.

Vitamin D cannot any more be considered as exclusively necessary to prevent rickets or osteomalacia. Calcitriol produced in the kidney is known to have classical endocrine phosphocalcic properties. However, many other tissues express both vitamin D receptor and 1a-hydroxylase and can convert 25-hydroxy vitamin D into calcitriol. Calcitriol produced locally is considered to have autocrine and paracrine actions on cellular proliferation and differentiation, apoptosis, insulin and renin secretion, interleukin and bactericidal proteins production. Epidemiologic and experimental data argue in favour of a protective role of vitamin D against cancers, type 2 diabetes, cardiovascular, auto-immune and infectious diseases, chronic kidney disease and loss of muscular strength. A few interventional studies confirm the protective effect of vitamin D against cancers, intermediate markers of cardiovascular risk, epidemic influenza, albuminuria and risk of fall. We present here the non phosphocalcic actions of vitamin D.

Evidence that vitamin D(3) promotes mast cell-dependent reduction of chronic UVB-induced skin pathology in mice.

Mast cell production of interleukin-10 (IL-10) can limit the skin pathology induced by chronic low-dose ultraviolet (UV)-B irradiation. Although the mechanism that promotes mast cell IL-10 production in this setting is unknown, exposure of the skin to UVB irradiation induces increased production of the immune modifying agent 1alpha,25-dihydroxyvitamin D(3) (1alpha,25[OH](2)D(3)). We now show that 1alpha,25(OH)(2)D(3) can up-regulate IL-10 mRNA expression and induce IL-10 secretion in mouse mast cells in vitro. To investigate the roles of 1alpha,25(OH)(2)D(3) and mast cell vitamin D receptor (VDR) expression in chronically UVB-irradiated skin in vivo, we engrafted the skin of genetically mast cell-deficient WBB6F(1)-Kit(W/W-v) mice with bone marrow-derived cultured mast cells derived from C57BL/6 wild-type or VDR(-/-) mice. Optimal mast cell-dependent suppression of the inflammation, local production of proinflammatory cytokines, epidermal hyperplasia, and epidermal ulceration associated with chronic UVB irradiation of the skin in Kit(W/W-v) mice required expression of VDR by the adoptively transferred mast cells. Our findings suggest that 1alpha,25(OH)(2)D(3)/VDR-dependent induction of IL-10 production by cutaneous mast cells can contribute to the mast cell's ability to suppress inflammation and skin pathology at sites of chronic UVB irradiation.