Contributions of bile acids to gastrointestinal physiology as receptor agonists and modifiers of ion channels.

Bile acids (BAs) are known to be important regulators of intestinal motility and epithelial fluid and electrolyte transport. Over the past two decades, significant advances in identifying and characterizing the receptors, transporters, and ion channels targeted by BAs have led to exciting new insights into the molecular mechanisms involved in these processes. Our appreciation of BAs, their receptors, and BA-modulated ion channels as potential targets for the development of new approaches to treat intestinal motility and transport disorders is increasing. In the current review, we aim to summarize recent advances in our knowledge of the different BA receptors and BA-modulated ion channels present in the gastrointestinal system. We discuss how they regulate motility and epithelial transport, their roles in pathogenesis, and their therapeutic potential in a range of gastrointestinal diseases.

Relationship between BsmI polymorphism and VDR gene methylation profile, gender, metabolic profile, oxidative stress, and inflammation in adolescents / Relación entre polimorfismo BsmI y perfil de metilación del gen VDR, género, perfil metabólico, estrés oxidativo e inflamación en adolescentes

Background: the biological activity of vitamin D depends on the activity of its receptor or VDR. On the other hand, the activity of this receptor is influenced by its state of methylation. The objective of this study was to verify if the BsmI polymorphism of the VDR gene influences its methylation profile in adolescents. Secondly, it was to verify if the status of some metabolic factors (oxidative stress, inflammation, lipid profile, and glycemia) in the serum, and gender-adjusted vitamin D levels are independent factors with an influence on the VDR methylation profile. Methods and results: the study included 198 adolescents of both sexes, aged 15-19 years, who underwent testing for VDR gene methylation polymorphisms, serum vitamin D levels, and metabolic, oxidative stress, and systemic inflammation markers. It was observed that the BB genotype was less methylated than the other groups (26.1 % versus 30.3 %, and 29.3 % for Bb and bb, respectively), although without statistical differences between them. The odds ratio indicated a protection of 13 % (partially methylated) for vitamin D status, while alpha glycols increased the risk ratio (of being partially methylated) by 3 %. MDA was protective at a 28 % chance of risk that adolescents with higher levels of lipid peroxidation would be hypomethylated. Conclusion: we conclude that the methylation profile of the VDR gene is not influenced by the different BsmI polymorphism genotypes, and that serum vitamin D and serum markers of oxidative stress and inflammation can modulate this profile. (AU)

Antecedentes: la actividad biológica de la vitamina D depende de la actividad de su receptor, el VDR. Por otro lado, la actividad de este receptor está influenciada por su estado de metilación. El objetivo de este estudio es verificar si el polimorfismo BsmI del gen VDR influye en el perfil de metilación del mismo en los adolescentes. En segundo lugar, verificar si los factores metabólicos (estrés oxidativo, inflamación, perfil lipídico y glucemia) del suero y la vitamina D ajustada por sexo actúan independientemente de los polimorfismos sobre el perfil de metilación del VDR. Métodos y resultados: el estudio incluyó a 198 adolescentes de ambos sexos, de 15 a 19 años de edad, que se sometieron a análisis de polimorfismos de metilación del gen VDR, niveles de vitamina D, marcadores metabólicos, estrés oxidativo e inflamación sistémica. Se observó que el genotipo BB estaba menos metilado que los otros grupos (26,1 % contra 30,3 % y 29,3 % para Bb y bb respectivamente), aunque sin diferencias estadísticas entre ellos. El odds ratio indicó una protección del 13 % (parcialmente metilado) para el estado de la vitamina D, mientras que los alfa glicoles aumentaron el índice de riesgo (de estar parcialmente metilado) en un 3 %. La MDA fue protectora con un 28 % de probabilidad de riesgo de que los adolescentes con niveles más altos de peroxidación lipídica fueran hipometilados. Conclusión: concluimos que el perfil de metilación del gen VDR no está influenciado por los diferentes genotipos del polimorfismo BsmI y que la vitamina D y los marcadores de estrés oxidativo e inflamación en el suero pueden modular este perfil. (AU)

Relationship between BsmI polymorphism and VDR gene methylation profile, gender, metabolic profile, oxidative stress, and inflammation in adolescents.

INTRODUCTION: Background: the biological activity of vitamin D depends on the activity of its receptor or VDR. On the other hand, the activity of this receptor is influenced by its state of methylation. The objective of this study was to verify if the BsmI polymorphism of the VDR gene influences its methylation profile in adolescents. Secondly, it was to verify if the status of some metabolic factors (oxidative stress, inflammation, lipid profile, and glycemia) in the serum, and gender-adjusted vitamin D levels are independent factors with an influence on the VDR methylation profile. Methods and results: the study included 198 adolescents of both sexes, aged 15-19 years, who underwent testing for VDR gene methylation polymorphisms, serum vitamin D levels, and metabolic, oxidative stress, and systemic inflammation markers. It was observed that the BB genotype was less methylated than the other groups (26.1 % versus 30.3 %, and 29.3 % for Bb and bb, respectively), although without statistical differences between them. The odds ratio indicated a protection of 13 % (partially methylated) for vitamin D status, while alpha glycols increased the risk ratio (of being partially methylated) by 3 %. MDA was protective at a 28 % chance of risk that adolescents with higher levels of lipid peroxidation would be hypomethylated. Conclusion: we conclude that the methylation profile of the VDR gene is not influenced by the different BsmI polymorphism genotypes, and that serum vitamin D and serum markers of oxidative stress and inflammation can modulate this profile.

INTRODUCCIÓN: Antecedentes: la actividad biológica de la vitamina D depende de la actividad de su receptor, el VDR. Por otro lado, la actividad de este receptor está influenciada por su estado de metilación. El objetivo de este estudio es verificar si el polimorfismo BsmI del gen VDR influye en el perfil de metilación del mismo en los adolescentes. En segundo lugar, verificar si los factores metabólicos (estrés oxidativo, inflamación, perfil lipídico y glucemia) del suero y la vitamina D ajustada por sexo actúan independientemente de los polimorfismos sobre el perfil de metilación del VDR. Métodos y resultados: el estudio incluyó a 198 adolescentes de ambos sexos, de 15 a 19 años de edad, que se sometieron a análisis de polimorfismos de metilación del gen VDR, niveles de vitamina D, marcadores metabólicos, estrés oxidativo e inflamación sistémica. Se observó que el genotipo BB estaba menos metilado que los otros grupos (26,1 % contra 30,3 % y 29,3 % para Bb y bb respectivamente), aunque sin diferencias estadísticas entre ellos. El odds ratio indicó una protección del 13 % (parcialmente metilado) para el estado de la vitamina D, mientras que los alfa glicoles aumentaron el índice de riesgo (de estar parcialmente metilado) en un 3 %. La MDA fue protectora con un 28 % de probabilidad de riesgo de que los adolescentes con niveles más altos de peroxidación lipídica fueran hipometilados. Conclusión: concluimos que el perfil de metilación del gen VDR no está influenciado por los diferentes genotipos del polimorfismo BsmI y que la vitamina D y los marcadores de estrés oxidativo e inflamación en el suero pueden modular este perfil.

Yes-associated protein regulates the hepatoprotective effect of vitamin D receptor activation through promoting adaptive bile duct remodeling in cholestatic mice.

Mounting clinical evidence has revealed that the vitamin D receptor (VDR) is associated with cholestatic liver injury, although the functions of VDR in this condition remain largely unexplored. Here, we investigated the effects of VDR activation on bile duct ligation (BDL) mice, and the underlying mechanisms were further investigated. A low-calcemic VDR agonist, paricalcitol (PAL, 200 ng/kg), was intraperitoneally injected into BDL mice every other day for 5 days or 28 days. Liver histology, liver function indicators, cholangiocyte proliferation, fibrosis scores, and inflammation were evaluated. Mice treated with PAL were rescued from the decreased survival rate induced by BDL and liver damage was reduced. Mechanistically, PAL promoted cholangiocyte proliferation, which was likely conducive to proliferating bile duct maturation and increased branching of bile ducts. PAL treatment also increased the expression of Yes-associated protein (YAP) and its target protein epithelial cell adhesion molecule (EpCam) and decreased the level of inactive cytoplasmic phosphorylated YAP. YAP knockdown abrogated PAL-induced primary bile duct epithelial cell proliferation, confirmed with YAP inhibitor administration. In addition, BDL-induced liver fibrosis and inflammatory cell infiltration were reduced by PAL treatment at both day 5 and day 28 post-BDL. In conclusion, VDR activation mitigates cholestatic liver injury by promoting adaptive bile duct remodeling through cholangiocytic YAP upregulation. Because PAL is an approved clinical drug, it may be useful for treatment of cholestatic liver disease. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Current treatment options for secondary hyperparathyroidism in patients with stage 3 to 4 chronic kidney disease and vitamin D deficiency.

Introduction: Secondary hyperparathyroidism (SHPT) represents a complication of chronic kidney disease (CKD). Vitamin D system is altered since early CKD, and vitamin D deficiency is an established trigger of SHPT. Although untreated SHPT may degenerate into tertiary hyperparathyroidism with detrimental consequences in advanced CKD, best treatments for counteracting SHPT from stage 3 CKD are still debated. Enthusiasm on prescription of vitamin D receptor activators (VDRA) in non-dialysis renal patients, has been mitigated by the risk of low bone turnover and positive calcium-phosphate balance. Nutritional vitamin D is now suggested as first-line therapy to treat SHPT with low 25(OH)D insufficiency. However, no high-grade evidence supports the best choice between ergocalciferol, cholecalciferol, and calcifediol (in its immediate or extended-release formulation).Areas covered: The review discusses available data on safety and efficacy of nutritional vitamin D, VDRA and nutritional therapy in replenishing 25(OH)D deficiency and counteracting SHPT in non-dialysis CKD patients.Expert opinion: Best treatment for low 25(OH)D and SHPT remains unknown, due to incomplete understanding of the best homeostatic, as mutable, adaptation of mineral metabolism to CKD progression. Nutritional vitamin D and nutritional therapy appear safest interventions, whenever contextualized with single-patient characteristics. VDRA should be restricted to uncontrolled SHPT by first-line therapy.

Local NF-κB Activation Promotes Parathyroid Hormone Synthesis and Secretion in Uremic Patients.

Secondary hyperparathyroidism (SHPT) in uremic patients is characterized by parathyroid gland (PTG) hyperplasia and parathyroid hormone (PTH) elevation. Previously, we demonstrated that NF-κB activation contributed to parathyroid cell proliferation in rats with chronic kidney disease. Although vitamin D inhibits inflammation and ameliorates SHPT, the contribution of vitamin D deficiency to SHPT via local NF-κB activation remains to be clarified. PTGs collected from 10 uremic patients with advanced SHPT were used to test the expressions of vitamin D receptor (VDR), NF-κB, and proliferating cell nuclear antigen (PCNA). Freshly excised PTG tissues were incubated for 24 hours in vitro with VDR activator (VDRA) calcitriol or NF-κB inhibitor pyrrolidine thiocarbamate (PDTC). Chromatin immunoprecipitation (ChIP) and luciferase reporter assays were performed to investigate the regulation of PTH transcription by NF-κB. We found higher levels of activated NF-κB and lower expression of VDR in nodular hyperplastic PTGs than in diffuse hyperplasia. In cultured PTG tissues, treatment with VDRA or PDTC inhibited NF-κB activation and PCNA expression, and downregulated preproPTH mRNA and intact PTH levels. ChIP assays demonstrated the presence of NF-κB binding sites in PTH promoter. Furthermore, in luciferase reporter assays, addition of exogenous p65 significantly increased PTH luciferase activity by 2.4-fold (P < 0.01), while mutation of NF-κB binding site at position -908 of the PTH promoter suppressed p65-induced PTH reporter activity (P < 0.01). In summary, local NF-κB activation contributes to SHPT and mediates the transcriptional activation of PTH directly in uremic patients. Vitamin D deficiency may be involved in SHPT via the activation of NF-κB pathway.

Influence of VDR and HFE polymorphisms on blood lead levels of occupationally exposed workers.

Lead is a ubiquitous heavy metal toxin of significant public health concern. Every individual varies in their response to lead's toxic effects due to underlying genetic variations in lead metabolizing enzymes or proteins distributed in the population. Earlier studies, including our lab, have attributed the influence of ALAD (δ-Aminolevulinate dehydratase) polymorphism on blood lead retention and ALAD activity. The present study aimed to investigate the influence of VDR (Vitamin D receptor) and HFE (Hemochromatosis) polymorphisms in modulating blood lead levels (BLLs) of occupationally exposed workers. 164 lead-exposed subjects involved in lead alloy manufacturing and battery breaking and recycling processes and 160 unexposed controls with BLLs below 10 µg/dL recruited in the study. Blood lead levels, along with a battery of biochemical assays and genotyping, were performed. Regression analysis revealed a negative influence of BLLs on ALAD activity (p < 0.0001) and a positive influence on smokeless tobacco use (p < 0.001) in lead-exposed subjects. A predicted haplotype of the three VDR polymorphisms computed from genotyping data revealed that T-A-A haplotype increased the BLLs by 0.93 units (p ≤ 0.05) and C-C-A haplotype decreased the BLLs by 7.25 units (p ≤ 0.05). Further analysis revealed that the wild-type CC genotype of HFE H63D presented a higher median BLL, indicating that variant C allele may have a role in increasing the concentration of lead. Hence, the polymorphism of genes associated with lead metabolism might aid in predicting genetic predisposition to lead and its associated effects.

Vitamin D inhibits Tissue Factor and CAMs expression in oxidized low-density lipoproteins-treated human endothelial cells by modulating NF-κB pathway.

Epidemiologic studies have clearly demonstrated the correlation existing between Vitamin D (Vit. D) deficiency and increased risk of developing cardiovascular disease, suggesting that it might have a protective role in this clinical setting. Although many experimental studies have investigated the molecular mechanisms by which Vit. D might exert these effects, its potential role in protecting against athero-thrombosis is still partially unknown. We have investigated whether Vit. D might exert anti athero-thombotic effects by preventing expression of adhesion molecules (CAMs) and Tissue Factor (TF), molecules involved in atherothrombotic pathophysiology, in oxLDL stimulated endothelial cells (HUVEC). Moreover, we have investigated whether Vit. D effects might be due to the NF-kB modulation. HUVEC cultivated in medium enriched with Vit. D (10 nM) were stimulated with oxLDL (50 µg/ml). TF gene (RT-PCR), protein (Western blot), surface expression (FACS) and procoagulant activity (FXa generation assay) were measured. Similarly, CAMs gene (RT-PCR), surface expression (FACS) and soluble values (ELISA) were measured. NF-kB translocation was also investigated. Vit. D significantly reduced TF gene as well protein expression and procoagulant activity in oxLDL-treated HUVEC. Similar effects were observed for CAMs. These effects were associated with Vit. D modulation of NF-κB pathway. This study, although in vitro, indicate that Vit. D has protective effect on endothelial cells by inhibiting expression of TF and CAMs, proteins involved in atherothrombotic pathophysiology. Further studies will be necessary to translate these findings to a clinical scenario to better define the potential therapeutical role of Vit. D supplementation in the management of cardiovascular disease in patients with Vit. D deficiency.

Design, synthesis and biological evaluation of novel 2-alkylidene 19-norcalcitriol analogs.

Continuing our studies aimed at A-ring modified vitamin D compounds, we designed novel 19-norcalcitriol derivatives bearing at C-2 pegylated chains of different lengths. The terminal fragments of these substituents contain hydroxyls or moieties possessing nitrogen and/or sulfur atoms capable of transition metal ions complexation. Also, two conjugate-type platinum(II) complexes of 19-norcalcitriol were obtained in which l-methionine served as chelating moiety. The convergent synthesis of the target 19-norcalcitriol analogs involved several steps with the crucial one being condensation of A-ring phosphine oxide and the known Grundmann ketone by Wittig-Horner reaction. Further elaboration of the 2-alkylidene substituent provided all final compounds which were then tested to determine their affinity for the vitamin D receptor and cytotoxic activity.

Paclitaxel, Carboplatin and 1,25-D3 Inhibit Proliferation of Ovarian Cancer Cells In Vitro.

BACKGROUND/AIM: The combination of paclitaxel and carboplatin is the standard chemotherapy for ovarian cancer. Previous studies have implied that vitamin D (1,25-D3) may have growth inhibitory effects in ovarian cancer. This study aimed to investigate the effect of paclitaxel, carboplatin and 1,25-D3 on the growth of ovarian cancer cells in vitro, based on the hypothesis that 1,25-D3 might potentiate the effect of paclitaxel and/or carboplatin. MATERIALS AND METHODS: Three non-commercial ovarian carcinoma cell lines UT-OV-1(mucinous), UT-OV-3B (serous) and UT-OV-4 (endometrioid) were exposed to different concentrations of 1,25-D3, paclitaxel and carboplatin, respectively. The cell viability was measured using a Crystal violet assay kit. The cellular vitamin D receptor (VDR) mRNA levels were measured by qRT-PCR using the LightCycler equipment. RESULTS: The growth-inhibitory effect of the combination of paclitaxel and carboplatin was 56% in UT-OV-1, 33% in UT-OV-3B and 47% in UT-OV-4 cells. Single 1,25-D3 (10 µM) inhibited the growth of UT-OV-3B and UT-OV-4 by 23% and 28%, respectively, whereas no effect was seen in UT-OV-1 cells. These results are in line with the finding that the expression of VDR was high in UT-OV-3B and UT-OV-4, but very low in UT-OV-1. The combination of 1,25-D3, paclitaxel and carboplatin resulted in 61%, 46% and 58% growth reduction in UT-OV-1, UT-OV-3B and UT-OV-4 cells, respectively. The additive effect of 1,25-D3 was 21% in UT-OV-4, 20% in UT-OV-3B and 12% in UT-OV-1 cell line. CONCLUSION: The results imply that combining 1,25-D3 with paclitaxel and carboplatin may potentiate their growth inhibitory effect on ovarian cancer cells with high VDR expression.

Artesunate interacts with the vitamin D receptor to reverse sepsis-induced immunosuppression in a mouse model via enhancing autophagy.

BACKGROUND AND PURPOSE: Immunosuppression is the predominant cause of mortality for sepsis because of failure to eradicate pathogens. No effective and specific drugs capable of reversing immunosuppression are clinically available. Evidences implicate the involvement of the vitamin D receptor (NR1I1) in sepsis-induced immunosuppression. The anti-malarial artesunate was investigated to determine action on sepsis-induced immunosuppression. EXPERIMENTAL APPROACH: The effect of artesunate on sepsis-induced immunosuppression was investigated in mice and human and mice cell lines. Bioinformatics predicted vitamin D receptor as a candidate target for artesunate, which was then identified using PCR and immunoblotting. Vdr, Atg16l1 and NF-κB p65 were modified to investigate artesunate 's effect on pro-inflammatory cytokines release, bacterial clearance and autophagy activities in sepsis-induced immunosuppression. KEY RESULTS: Artesunate significantly reduced the mortality of caecal ligation and puncture (CLP)-induced sepsis immunosuppression mice challenged with Pseudomonas aeruginosa and enhanced pro-inflammatory cytokine release and bacterial clearance to reverse sepsis-induced immunosuppression in vivo and in vitro. Mechanistically, artesunate interacted with vitamin D receptor, inhibiting its nuclear translocation, which influenced ATG16L1 transcription and subsequent autophagy activity. Artesunate inhibited the physical interaction between vitamin D receptor and NF-κB p65 in LPS-tolerant macrophages and then promoted the nuclear translocation of NF-κB p65, which activated the transcription of NF-κB p65 target genes such as pro-inflammatory cytokines. CONCLUSION AND IMPLICATIONS: Our findings provide evidence that artesunate interacted with vitamin D receptor to reverse sepsis-induced immunosuppression in an autophagy and NF-κB-dependent manner, highlighting a novel approach for sepsis treatment and drug repurposing of artesunate has a bidirectional immunomodulator.

Lymphocyte and monocyte vitamin D receptor expression during paricalcitol or calcitriol treatments in patients with stage 5 chronic kidney disease.

PURPOSE: In this study, we aimed to investigate the effect of paricalcitol and calcitriol usage on vitamin D receptor (VDR) contents of CD8+ , CD4+ lymphocytes and monocytes in stage 5d chronic kidney disease (CKD) patients. METHODS: Thirty-six hemodialysis patients older than 18 years of age and 19 healthy controls (group HC) without any known acute or chronic diseases were included in the study. The group of patients undergoing scheduled hemodialysis comprised three subgroups: group CL: patients on calcitriol (n: 10), group PC: patients on paricalcitol (n: 13), and group NT: patients not taking any vitamin D or VDR activating medications (n: 13). CD8+/VDR, CD4+/VDR and MONO/VDR values were representing the ratio of VDR representing cells among related cell group. On the other hand, values of CD8+/MFI, CD4+/MFI and MONO/MFI have shown the total amount of cellular VDR content per cell which has been given as of mean fluorescence intensity in the flow cytometric process. Main CKD mineral bone disorder parameters such as a hemogram, serum BUN, creatinine, albumin, Ca, iP, iPTH, 25(OH)D3 levels were also measured. RESULTS: Average VDR contents in CD8+, CD4+ and monocytes were not different among three patient groups on hemodialysis. But in all hemodialysis subgroups, CD8+/VDR, CD4+/VDR, MONO/VDR, CD8+/MFI, CD4+/MFI and MONO/MFI levels were found to be higher compared with the healthy control subjects (p < 0.001). Among hemodialysis groups, no significant CD8+/VDR, CD4+/VDR, and MONO/VDR content differences were found with regard to the type of VDR activator agent used. There was no difference in serum levels of 25(OH)D3 and CRP among groups participating in the study. CONCLUSION: There was no difference between CD8+/VDR, CD4+/VDR, and MONO/VDR levels in hemodialysis patients using calcitriol or paricalcitol, suggesting that both treatment agents may have a similar effect on VDR contents in lymphocytes and monocytes in that patient population. But in all hemodialysis subgroups, CD8+/VDR, CD4+/VDR, and MONO/VDR levels were found to be higher compared with the healthy control subjects, suggesting an overexpression of VDR through a non CRP and/or 25(OH)D3 dependent mechanism.

Oncomir MicroRNA-346 Is Upregulated in Colons of Patients With Primary Sclerosing Cholangitis.

INTRODUCTION: Primary sclerosing cholangitis (PSC) is a cholestatic liver disorder that is frequently associated with ulcerative colitis (UC). Patients with PSC and UC (PSC-UC) have a higher risk of colorectal neoplasia compared with patients with UC. The oncogenic properties of microRNA-346 (miR-346) have been recently reported. We investigated the expression of miR-346 and its 2 target genes, the receptor of vitamin D (VDR), and the tumor necrosis factor-α (TNF-α), which are known to modulate carcinogenesis. METHODS: Ascending and sigmoid colon biopsies were obtained from patients with PSC, PSC and UC (PSC-UC), UC, and healthy controls (n = 10 in each group). Expressions of VDR, TNF-α, 18S RNA, p27, miR-346, and reference microRNA, miR-191, were evaluated by real-time PCR using human TaqMan Gene Expression and TaqMan MicroRNA Assays. Functional studies with miR-346 mimic and inhibitor were conducted in HepG2 and Caco-2 cells. The effect of ursodeoxycholic acid on miR-346 expression was examined in Caco-2 cells. RESULTS: An increased expression of miR-346 in the ascending colon of PSC-UC was observed (P < 0.001 vs all groups). In patients with UC, an exceptionally low colonic expression of miRNA-346 was accompanied by the extensive upregulation of VDR and TNF-α genes. A functional in vitro analysis demonstrated that inhibition of miR-346 resulted in the upregulation of VDR and TNF-α, whereas the induction of miR-346 activity suppressed VDR, TNF-α, and p27. DISCUSSION: The upregulation of miRNA-346 in the colon of patients with PSC may be responsible for the disturbance of VDR and TNF-α signaling pathway, which could result in an inadequate suppression of neoplasia.

The Effect of Caffeine on Calcitriol-Inducible Vitamin D Receptor-Controlled Gene Expression in Intestinal and Osteoblastic Cells.

Some epidemiological studies suggested caffeine consumption as the cause for bone mineral density loss. Certain genes involved in this process are regulated by vitamin D receptor (VDR). Therefore, we investigated if caffeine can affect inducible expression of VDR-regulated genes, some of them being involved in bone mineralization process. By employing reporter gene assay, polymerase chain reaction, and western blotting, we monitored the VDR activity and expression in cell cultures of intestinal (LS180), osteosarcoma (HOS), and normal human osteoblasts in vitro. While caffeine stimulated calcitriol-inducible VDR-dependent nanoluciferase activity in stable reporter cell line IZ-VDRE (derived from LS180), it rather modulated mRNA levels of target genes, like CYP24A1, BGLAP, SPP1, and TNSF11 in LS180 and HOS cells. However, caffeine significantly decreased calcitriol-inducible CYP24A1, TNSF11, and SPP1 transcripts in osteoblasts. This decrease had non-linear U-shaped profile. Our in vitro data demonstrate biphasic action of caffeine on the expression of certain calcitriol-inducible VDR-regulated genes in normal human osteoblasts.

Genetic Polymorphisms of Pharmacogenomic VIP Variants in the Circassian Subpopulation from Jordan.

BACKGROUND: It has been suggested that genetic variation within candidate pharmacogenes contributes to the differences in drug safety and efficacy as well as risk of adverse drug reactions among different ethnic groups. Illustrating the polymorphic distribution of Very Important Pharmacogenes (VIPs) in various ethnic groups will contribute to the development of personalized medicine for those populations. OBJECTIVE: The present study aimed to identify the polymorphic distribution of VIPs in the Circassian subpopulation of Jordan and compare their allele frequencies with those of other populations. METHODS: A total of 130 healthy and unrelated Circassian adults from Jordan were randomly recruited and genotyped for eleven VIP variants within the thiopurine S-methyltransferase (TPMT), ATP-binding cassette, sub-family B, member 1 (ABCB1), and vitamin D receptor (VDR) genes via Sequenom's MassARRAY® genotyping platform (iPLEX GOLD). RESULTS: Our data on the allelic frequencies of the investigated VIP variants were compared to those of 18 other populations, comprising 11 HapMap populations, 6 Exome Aggregation Consortium populations, and the Chechen- Jordanian population from Jordan. Circassian-Jordanians were found to most resemble the African, Chechen- Jordanian, European (Finnish), European (non-Finnish), and South-Asian populations. CONCLUSION: Circassians from Jordan significantly differ from other populations in terms of the allelic frequencies of selected VIP variants. The present findings constitute the first set of pharmacogenetic data for Circassian population from Jordan, providing a basis for safe drug administration that may be useful in diagnosing and treating diseases in this ethnic group.

Changes in microparticle profiles by vitamin D receptor activation in chronic kidney disease - a randomized trial.

BACKGROUND: Microparticles (MPs) are biomarkers and mediators of disease through their expression of surface receptors, reflecting activation or stress in their parent cells. Endothelial markers, ICAM-1 and VCAM-1, are implicated in atherosclerosis and associated with cardiovascular risk. Chronic kidney disease (CKD) patients have endothelial dysfunction and high levels of endothelial derived MPs. Vitamin D treatment has been reported to ameliorate endothelial function in CKD patients. We aimed to examine cell specific MP profiles and concentrations of MPs expressing the atherosclerotic markers ICAM-1 and VCAM-1 after treatment with paricalcitol in patients with CKD stage 3-4. METHODS: Sub-study of the previously reported SOLID trial where 36 patients were randomly assigned to placebo, 1 or 2 µg paricalcitol, for 12 weeks. MPs were measured by flow cytometry after labelling with antibodies against endothelial (CD62E), platelet (CD62P, CD41, CD154) leukocyte (CD45) and vascular (CD54, CD106) markers. RESULTS: Patients had a mean age of 65 years with a mean eGFR of 40 mL/min/1.73m2. Concentrations of ICAM-1 positive MPs were significantly reduced by treatment (repeated measures ANOVA p = 0.04). Repeated measures MANOVA of concentrations of endothelial, platelet and leukocyte MPs showed sustained levels in the 2 µg treatment group (p = 0.85) but a decline in the 1 µg (p = 0.04) and placebo groups (p = 0.005). CONCLUSIONS: Treatment with paricalcitol reduces concentrations of ICAM-1 positive MPs. This is accompanied by sustained concentrations of all cell specific MPs in the 2 µg group, and decreasing concentrations in the other groups, possibly due to a more healthy and reactive endothelium with paricalcitol treatment.

From epidemiology and neurometabolism to treatment: Vitamin D in pathogenesis of glioblastoma Multiforme (GBM) and a proposal for Vitamin D + all-trans retinoic acid + Temozolomide combination in treatment of GBM.

Here we review tumoricidal efficacy of Vitamin D analogues in glioblastoma multiforme (GBM) and potential synergisms with retinoic acid and temozolomide based on epidemiological and cellular studies. Epidemiological data suggest that winter birth is associated with higher risk of GBM, and GBM debulking in the winter enhanced mortality, which may relate with lower exposure to sunlight essential to convert cholecalciferol to Vitamin D. Comparative studies on blood bank specimens revealed that higher prediagnosis levels of calcidiol are associated with lower risk of GBM in elderly men. Supplemental Vitamin D reduced mortality in GBM patients in comparison to nonusers. Expression of Vitamin D Receptor is associated with a good prognosis in GBM. Conversely, Vitamin D increases glial tumor synthesis of neutrophins NGF and NT-3, the low affinity neurotrophin receptor p75NTR, IL-6 and VEGF, which may enhance glioma growth. Antitumor synergisms between temozolomide and Vitamin D and Vitamin D with Vitamin A derivatives were observed. Hence, we hypothesize that Calcitriol + ATRA (All-Trans Retinoic Acid) + Temozolomide - CAT combination might be a safer approach to benefit from Vitamin D in the management of high-grade glial tumors. Adding acetazolomide to this protocol may reduce the risk of pseudotumor cerebri, as both Vitamin D and Vitamin A excess may cause intracranial hypertension; this approach may provide further benefit as acetazolomide also exhibits anticancer activity.

Modulation of VDR and Cell Cycle-Related Proteins by Vitamin D in Normal Pancreatic Cells and Poorly Differentiated Metastatic Pancreatic Cancer Cells.

Many in vitro studies support the general idea that vitamin D plays a protective role against cancer. Increased doses of vitamin D dietary supplements have been widely used for the potential anticancer benefits of vitamin D. However, despite substantial epidemiological research, there are no clear conclusive data to support the use of vitamin D as a cancer preventive or treatment agent. In the, herein, reported study, we checked the effects of 1,25-dihydroxyvitamin D3 concentrations on the expression level of the vitamin D receptor (VDR) and cell cycle-related proteins CDKN1A (p21) and CDK1 in pancreatic cells and Panc-1 pancreatic cancer (PC) cells. We found that VDR, CDKN1A, and CDK1 were upregulated by an increase in 1,25-dihydroxyvitamin D3 concentration in normal pancreatic cells but not in the advanced cancer cell line Panc-1 from poorly differentiated metastatic PC cells. A further increase in 1,25-dihydroxyvitamin D3 concentration above the physiological range significantly downregulated the expression of VDR, indicating that VDR is modulated by VDR levels to maintain normal functioning during dramatic variations in vitamin D concentration. By increasing the level of cell cycle inhibitory and promoting proteins p21 and CDK1, vitamin D theoretically has both preventive and promoting effects on pancreatic cell division.

Stanozolol promotes osteogenic gene expression and apposition of bone mineral in vitro

Abstract Stanozolol (ST) is a synthetic androgen with high anabolic potential. Although it is known that androgens play a positive role in bone metabolism, ST action on bone cells has not been sufficiently tested to support its clinical use for bone augmentation procedures. Objective: This study aimed to assess the effects of ST on osteogenic activity and gene expression in SaOS-2 cells. Material and Methods: SaOS-2 deposition of mineralizing matrix in response to increasing doses of ST (0-1000 nM) was evaluated through Alizarin Red S and Calcein Green staining techniques at 6, 12 and 24 days. Gene expression of runt-related transcription factor 2 (RUNX2), vitamin D receptor (VDR), osteopontin (SPP1) and osteonectin (ON) was analyzed by RT-PCR. Results: ST significantly influenced SaOS-2 osteogenic activity: stainings showed the presence of rounded calcified nodules, which increased both in number and in size over time and depending on ST dose. RT-PCR highlighted ST modulation of genes related to osteogenic differentiation. Conclusions: This study provided encouraging results, showing ST promoted the osteogenic commitment of SaOS-2 cells. Further studies are required to validate these data in primary osteoblasts and to investigate ST molecular pathway of action.

Effect of Oral Alfacalcidol on Clinical Outcomes in Patients Without Secondary Hyperparathyroidism Receiving Maintenance Hemodialysis: The J-DAVID Randomized Clinical Trial.

Importance: Patients with chronic kidney disease have impaired vitamin D activation and elevated cardiovascular risk. Observational studies in patients treated with hemodialysis showed that the use of active vitamin D sterols was associated with lower risk of all-cause mortality, regardless of parathyroid hormone levels. Objective: To determine whether vitamin D receptor activators reduce cardiovascular events and mortality in patients without secondary hyperparathyroidism undergoing hemodialysis. Design, Setting, and Participants: Randomized, open-label, blinded end point multicenter study of 1289 patients in 207 dialysis centers in Japan. The study included 976 patients receiving maintenance hemodialysis with serum intact parathyroid hormone levels less than or equal to 180 pg/mL. The first and last participants were enrolled on August 18, 2008, and January 26, 2011, respectively. The final date of follow-up was April 4, 2015. Interventions: Treatment with 0.5 µg of oral alfacalcidol per day (intervention group; n = 495) vs treatment without vitamin D receptor activators (control group; n = 481). Main Outcomes and Measures: The primary outcome was a composite measure of fatal and nonfatal cardiovascular events, including myocardial infarctions, hospitalizations for congestive heart failure, stroke, aortic dissection/rupture, amputation of lower limb due to ischemia, and cardiac sudden death; coronary revascularization; and leg artery revascularization during 48 months of follow-up. The secondary outcome was all-cause death. Results: Among 976 patients who were randomized from 108 dialysis centers, 964 patients were included in the intention-to-treat analysis (median age, 65 years; 386 women [40.0%]), and 944 (97.9%) completed the trial. During follow-up (median, 4.0 years), the primary composite outcome of cardiovascular events occurred in 103 of 488 patients (21.1%) in the intervention group and 85 of 476 patients (17.9%) in the control group (absolute difference, 3.25% [95% CI, -1.75% to 8.24%]; hazard ratio, 1.25 [95% CI, 0.94-1.67]; P = .13). There was no significant difference in the secondary outcome of all-cause mortality between the groups (18.2% vs 16.8%, respectively; hazard ratio, 1.12 [95% CI, 0.83-1.52]; P = .46). Of the 488 participants in the intervention group, 199 (40.8%) experienced serious adverse events that were classified as cardiovascular, 64 (13.1%) experienced adverse events classified as infection, and 22 (4.5%) experienced malignancy-related serious adverse events. Of 476 participants in the control group, 191 (40.1%) experienced cardiovascular-related serious adverse events, 63 (13.2%) experienced infection-related serious adverse events, and 21 (4.4%) experienced malignancy-related adverse events. Conclusions and Relevance: Among patients without secondary hyperparathyroidism undergoing maintenance hemodialysis, oral alfacalcidol compared with usual care did not reduce the risk of a composite measure of select cardiovascular events. These findings do not support the use of vitamin D receptor activators for patients such as these. Trial Registration: UMIN-CTR Identifier: UMIN000001194.