Serum cholesterol and expression of ApoAI, LXRβ and SREBP2 in vitamin D receptor knock-out mice

Vitamin D insufficiency has been reported to be associated with increased blood cholesterol concentrations. Here we used two strains of VDR knock-out (VDR-KO) mice to study whether a lack of vitamin D action has any effect on cholesterol metabolism. In 129S1 mice, both in male and female VDR-KO mice serum total cholesterol levels were significantly higher than those in wild type (WT) mice (20.7% (P = 0.05) and 22.2% (P = 0.03), respectively). In addition, the serum high-density lipoprotein-bound cholesterol (HDL-C) level was 22% (P = 0.03), respectively higher in male VDR-KO mice than in WT mice. The mRNA expression levels of five cholesterol metabolism related genes in livers of 129S1 mice were studied using quantitative real-time PCR (QRT-PCR): ATP-binding cassette transporter A1 (ABCA1), regulatory element binding protein (SREBP2), apolipoprotein A-I (ApoAI), low-density lipoprotein receptor (LDLR) and liver X receptor beta (LXRβ). In the mutant male mice, the mRNA level of ApoAI and LXRβ were 49.2% (P = 0.005) and 38.8% (P = 0.034) higher than in the WT mice. These changes were not observed in mutant female mice, but the female mutant mice showed 52.5% (P = 0.006) decrease of SREBP2 mRNA expression compared to WT mice. Because the mutant mice were fed with a special rescue diet, we wanted to test whether the increased cholesterol levels in mutant mice were due to the diet. Both the WT and mutant NMRI mice were given the same diet for 3 weeks before the blood sampling. No difference in cholesterol or in HDL-C between WT and mutant mice was found. The results suggest that the food, gender and genetic background have an effect on the cholesterol metabolism. Although VDR seems to regulate some of the genes involved in cholesterol metabolism, its role in the regulation of serum cholesterol seems to be minimal.     

Premature aging in vitamin D receptor mutant mice

Hypervitaminosis vitamin D₃ has been recently implicated in premature aging through the regulation of 1alpha hydroxylase expression by klotho and fibroblast growth factor-23 (Fgf-23). Here we examined whether the lack of hormonal function of vitamin D₃ in mice is linked to aging phenomena. For this, we used vitamin D₃ receptor (VDR) “Tokyo” knockout (KO) mice (fed with a special rescue diet) and analyzed their growth, skin and cerebellar morphology, as well as overall motor performance. We also studied the expression of aging-related genes, such as Fgf-23, nuclear factor kappaB (NF-kappaB), p53, insulin like growth factor 1 (IGF1) and IGF1 receptor (IGF1R), in liver, as well as klotho in liver, kidney and prostate tissues. Overall, VDR KO mice showed several aging related phenotypes, including poorer survival, early alopecia, thickened skin, enlarged sebaceous glands and development of epidermal cysts. There was no difference either in the structure of cerebellum or in the number of Purkinje cells. Unlike the wildtype controls, VDR KO mice lose their ability to swim after 6 months of age. Expression of all the genes was lower in old VDR KO mice, but only NF-kappaB, Fgf-23, p53 and IGF1R were significantly lower. Since the phenotype of aged VDR knockout mice is similar to mouse models with hypervitaminosis D₃, our study suggests that VDR genetic ablation promotes premature aging in mice, and that vitamin D₃ homeostasis regulates physiological aging.     

雌激素改善维生素D受体基因敲除雌性小鼠的骨、钙代谢

以维生素 D 受体基因敲除雌性小鼠为模型,研究雌激素对骨、钙代谢的调节作用。外源性给予雌二醇一个月后,观察小鼠血钙水平的变化,同时测定小鼠骨密度,并利用胫骨非脱钙 von Kossa 染色观察钙化的骨小梁和未钙化的类骨质面积的变化。结果显示,外源性给予雌二醇一个月后,维生素 D 受体基因敲除小鼠的血钙水平,从(2.10±0.37) mmol/L 上升到(2.80±0.41) mmol/L (P<0.05); 骨密度从(0.037±0.006) g/cm2增高到(0.048±0.007) g/cm2,显著改善(P<0.05) ;钙化骨小梁面积显著增加,未钙化的类骨质面积显著缩小。结果提示,外源性雌二醇对骨、钙代谢具有非依赖于维生素 D 的正向调节作用。     

Structure-activity relationship studies on vitamin D lactam derivatives as vitamin D receptor antagonist

Structure–activity relationship studies on 1α,25-dihydroxyvitamin D₃-26,23-lactams (DLAMs), antagonists of vitamin D, were conducted, focusing on the substituents of the phenyl group. One of the derivatives (23S,25S)-DLAM-1P-3,5(OEt)₂, showed potent antagonistic activity with an IC₅₀ of 90 nM.     

Exons and functional regions of the human vitamin D receptor gene around and within the main 1a promoter are well conserved among mammals

The human vitamin D receptor (hVDR) gene encompasses eight exons (2–9) in the so-called coding region and six more exons (1a–1f) in the so-called regulatory region, which contains several reported promoters. Evolutionary comparison performed on the VDR promoter sequences of a dozen of mammalian species shows a very high conservation of numerous regions around and in the 1a promoter, including exons 1e, 1a and 1d, and the Sp1 site region. This suggests that the so-called 1a promoter is well conserved among mammals. Homology among mammals also concerns three functional SNP site regions of the hVDR 1a promoter, the 1e-G-1739A SNP region (a Cdx-2 binding site), and both 1a-G-1521C and 1a-A-1012G sites, the 1a-1012A being located within a GATA site. Interestingly, the 1521G and 1012A nucleotides are being evolutionary conserved, suggesting that the 1521C/1012G haplotype, which is found in human chromosomes (43% of Caucasians), is a human specificity.     

Aberrant nest building and prolactin secretion in vitamin D receptor mutant mice

1,25(OH)₂D₃, the hormonal form of vitamin D, is a neuroactive seco-steroid hormone with multiple functions in the brain. Most of these effects are mediated through the nuclear vitamin D receptor (VDR), widely distributed in the central nervous system. Our earlier studies showed that mutant mice lacking functional VDR have specific behavioural abnormalities, including anxiety and aberrant maternal behaviour, which may be hormonally regulated. Here we describe impaired nest building behaviour in VDR mutant mice. Since prolactin plays a key role in the regulation of nest building in both sexes, we also examine whether VDR mutant mice have altered prolactin levels. Overall, serum prolactin levels were increased in VDR mutant mice, accompanied by marked impairments in their nest building activity. In contrast, there were no differences in prolactin mRNA expression levels between wildtype control mice and VDR mutant mice. Collectively, these data suggest that partial genetic ablation of VDR affects prolactin system in mice, and that altered serum prolactin levels in VDR mutants may underlie some of their behavioural abnormalities, such as impaired nest building.     

Vestibular dysfunction in vitamin D receptor mutant mice

The vitamin D endocrine system is essential for calcium and bone homeostasis. Vitamin D deficits are associated with muscle weakness and osteoporosis, whereas vitamin D supplementation may improve muscle function, body sway and frequency of falls, growth and mineral homeostasis of bones. The loss of muscle strength and mass, as well as deficits in bone formation, lead to poor balance. Poor balance is one of the main causes of falls, and may lead to dangerous injuries. Here we examine balance functions in vitamin D receptor deficient (VDR−/−) mice, an animal model of vitamin D-dependent rickets type II, and in 1α-hydroxylase deficient (1α-OHase−/−) mice, an animal model of pseudovitamin D-deficiency rickets. Recently developed methods (tilting box, rotating tube test), swim test, and modified accelerating rotarod protocol were used to examine whether the absence of functional VDR, or the lack of a key vitamin D-activating enzyme, could lead to mouse vestibular dysfunctions. Overall, VDR−/− mice, but not 1α-OHase−/− mice, showed shorter latency to fall from the rotarod, smaller fall angle in the tilting box test, and aberrant poor swimming. These data suggest that VDR deficiency in mice is associated with decreased balance function, and may be relevant to poorer balance/posture control in humans with low levels of vitamin D.     

Targeted expression of human vitamin D receptor in adipocytes decreases energy expenditure and induces obesity in mice

Our previous studies demonstrated a high fat diet-resistant lean phenotype of vitamin D receptor (VDR)-null mutant mice mainly due to increased energy expenditure, suggesting an involvement of the VDR in energy metabolism. Here, we took a transgenic approach to further define the role of VDR in adipocyte biology. We used the aP2 gene promoter to target the expression of the human (h) VDR in adipocytes in mice. In contrast to the VDR-null mice, the aP2-hVDR Tg mice developed obesity compared with the wild-type counterparts without changes in food intake. The increase in fat mass was mainly due to markedly reduced energy expenditure, which was correlated with decreased locomotive activity and reduced fatty acid β-oxidation and lipolysis in the adipose tissue in the transgenic mice. Consistently, the expression of genes involved in the regulation of fatty acid transport, thermogenesis, and lipolysis were suppressed in the transgenic mice. Taken together, these data confirm an important role of the VDR in the regulation of energy metabolism.     

Enriched uranium affects the expression of vitamin D receptor and retinoid X receptor in rat kidney

An increasing awareness of the radiological impact of the nuclear power industry and other nuclear technologies is observed nowadays on general population. This led to renew interest to assess the health impact of the use of enriched uranium (EU). The aim of this work was to investigate in vivo the effects of a chronic exposure to EU on vitamin D(3) metabolism, a hormone essential in mineral and bone homeostasis. Rats were exposed to EU in their drinking water for 9 months at a concentration of 40 mg l(-1) (1mg/rat day). The contamination did not change vitamin D plasma level. Vitamin D receptor (vdr) and retinoid X receptor alpha (rxralpha), encoding nuclear receptors involved in the biological activities of vitamin D, showed a lower expression in kidney, while their protein levels were paradoxically increased. Gene expression of vitamin D target genes, epithelial Ca(2+) channel 1 (ecac1) and Calbindin-D28k (cabp-d28k), involved in renal calcium transport were decreased. Among the vitamin D target organs examined, these molecular modifications occurred exclusively in the kidney, which confirms that this organ is highly sensitive to uranium exposure. In conclusion, this study showed that a chronic exposure to EU affects both mRNA and protein expressions of renal nuclear receptors involved in vitamin D metabolism, without any modification of the circulating vitamin D.     

Characterization of heart size and blood pressure in the vitamin D receptor knockout mouse

Our previous studies showed vitamin D deficiency results in increased cardiac contractility, hypertrophy and fibrosis and has profound effects on heart proteomics, structure and function in rat. In this study we found that the heart in vitamin D receptor knockout (VDR-KO) mice is hypertrophied. Six homozygous VDR knockout (−/−), six wild type (+/+) and six heterozygous (+/−) mice were fed a diet containing 2% Ca, 1.25% P and 20% lactose to maintain normal blood calcium and phosphate levels for 12 months. Tail-cuff blood pressure was performed on all mice. Blood pressure determinations showed no differences in systolic or mean blood pressure in WT (+/+), KO (−/−) or HETERO (+/−) mice at 3 and 6 months. However, decreased systolic BP in the KO mice relative to WT at 9 months of age was observed. ECG analysis showed no significant differences in the intact KO, HETERO or WT mice. The mice were killed at 12 months. Heart weight/body weight ratio was 41% (P < .003) greater in the KO mice versus WT and HETERO was 19% (P < .05) increased versus WT. Other VDR-KO tissues did not display hypertrophy. Cross sectional and longitudinal analysis of the heart myofibrils showed highly significant cellular hypertrophy in VDR-KO mice. Trichrome staining of heart tissue showed marked increase in fibrotic lesions in the KO mice. Analysis of plasma renin activity, angiotensin II (AII) and aldosterone levels showed elevated but not significantly different renin activity in KO versus WT and no significant differences in AII or aldosterone levels. Our data do not support the concept that the renin-angiotensin system or hypertension are the factors that elicit these changes. Data presented here reveal that ablation of the VDR signaling system results in profound changes in heart structure. We propose that calcitriol acts directly on the heart as a tranquilizer by blunting cardiomyocyte hypertrophy.     

Association of vitamin D receptor gene polymorphism with the risk of systemic lupus erythematosus

Abstract

Relationship between vitamin D receptor (VDR) gene polymorphism and the risk of systemic lupus erythematosus (SLE) from the published reports are still conflicting. This study was conducted to evaluate the relationship between VDR BsmI (rs1544410), Fok1 (rs2228570), ApaI (rs7975232) and TaqI (rs731236) gene polymorphism and the risk of SLE using meta-analysis method. The association studies were identified from PubMed and Cochrane Library on 1 March 2014, and eligible investigations were included and synthesized using meta-analysis method. Thirteen reports were recruited into this meta-analysis for the association of VDR gene polymorphism with SLE susceptibility. In this meta-analysis for overall populations, the BsmI B allele and bb genotype, Fok1 f allele and ff genotype, and ApaI aa genotype, were associated with the risk of SLE. In Asians, the BsmI B allele, BB genotype and bb genotype, Fok1 f allele and ff genotype were associated with the risk of SLE. In Africans, the BsmI B allele, BB genotype and bb genotype, Fok1 f allele and ff genotype, ApaI A allele, AA genotype and aa genotype were associated with the risk of SLE. However, VDR BsmI, Fok1, ApaI and TaqI gene polymorphism were not associated with the risk of SLE in Caucasians. In conclusion, the BsmI B allele and bb genotype, Fok1 f allele and ff genotype were associated with the risk of SLE in overall populations, and in Asians, but these associations were not found in Caucasians. However, more studies should be conducted to confirm it.     

Association of vitamin D receptor gene polymorphism with the urine calcium level in nephrolithiasis patients

Abstract

Association of vitamin D receptor (VDR) gene polymorphism with the urine calcium level in nephrolithiasis patients from the published reports are still conflicting. This study was conducted to evaluate the relationship between VDR BsmI (rs1544410), Fok1 (rs2228570), TaqI (rs731236) and ApaI (rs7975232) gene polymorphism and urine calcium level in nephrolithiasis patients using meta-analysis method. The association studies were identified from PubMed, and Cochrane Library on 1 April 2014, and eligible investigations were included and synthesized using meta-analysis method. Four reports were recruited into this meta-analysis for the association of VDR BsmI, Fok1, TaqI and ApaI gene polymorphism with urine calcium level in nephrolithiasis patients. In this meta-analysis, VDR BsmI B allele and BB genotype, Fok1 f allele and ff genotype, TaqI, and ApaI gene polymorphism were not associated with urine calcium level in nephrolithiasis patients. However, the BsmI bb genotype and Fok1 FF genotype were associated with the urine calcium level in nephrolithiasis patients. In conclusion, VDR BsmI bb genotype and Fok1 FF genotype were associated with the urine calcium level in nephrolithiasis patients. However, more studies should be conducted to confirm it.     

Increased nuclear expression and transactivation of vitamin D receptor by the cardiotonic steroid bufalin in human myeloid leukemia cells

The active form of vitamin D₃, 1α,25-dihydroxyvitamin D₃ [1,25(OH)₂D₃], is a potent ligand for the nuclear receptor vitamin D receptor (VDR) and induces myeloid leukemia cell differentiation. The cardiotonic steroid bufalin enhances vitamin D-induced differentiation of leukemia cells and VDR transactivation activity. In this study, we examined the combined effects of 1,25(OH)₂D₃ and bufalin on differentiation and VDR target gene expression in human leukemia cells. Bufalin in combination with 1,25(OH)₂D₃ enhanced the expression of VDR target genes, such as CYP24A1 and cathelicidin antimicrobial peptide, and effectively induced differentiation phenotypes. An inhibitor of the Erk mitogen-activated protein (MAP) kinase pathway partially inhibited bufalin induction of VDR target gene expression. 1,25(OH)₂D₃ treatment induced transient nuclear expression of VDR in HL60 cells. Interestingly, bufalin enhanced 1,25(OH)₂D₃-induced nuclear VDR expression. The MAP kinase pathway inhibitor increased nuclear VDR expression induced by 1,25(OH)₂D₃ and did not change that by 1,25(OH)₂D₃ plus bufalin. A proteasome inhibitor also enhanced 1,25(OH)₂D₃-induced CYP24A1 expression and nuclear VDR expression. Bufalin-induced nuclear VDR expression was associated with histone acetylation and VDR recruitment to the CYP24A1 promoter in HL60 cells. Thus, the Na⁺,K⁺-ATPase inhibitor bufalin modulates VDR function through several mechanisms, including Erk MAP kinase activation and increased nuclear VDR expression.     

Association of vitamin D receptor gene polymorphism with the risk of lung cancer: a meta-analysis

Abstract

Relationship between vitamin D receptor (VDR) gene polymorphism and the risk of lung cancer from the published reports are still conflicting. This study was conducted to evaluate the relationship between VDR TaqI (rs731236), BsmI (rs1544410) and ApaI (rs7975232) gene polymorphism and the risk of lung cancer using meta-analysis method. The association studies were identified from PubMed and Cochrane Library on 1 December 2013, and eligible investigations were included and synthesized using meta-analysis method. Six reports were recruited into this meta-analysis for the association of VDR gene polymorphism with lung cancer susceptibility. In the meta-analysis for ApaI gene polymorphism, AA genotype was associated with the risk of lung cancer in Asians. In the meta-analysis for BsmI gene polymorphism, B allele, BB genotype and bb genotype were associated with lung cancer in Asians, and B allele bb genotype were associated with lung cancer risk in overall populations; furthermore, bb genotype was associated with lung cancer risk in Caucasians. In the meta-analysis for TaqI gene polymorphism, t allele and TT genotype were associated with lung cancer in overall populations and in Caucasians. In conclusion, B allele bb genotype t allele and TT genotype were associated with lung cancer risk in overall populations. AA genotype, B allele, BB genotype and bb genotype were associated with the risk of lung cancer in Asians. Furthermore, bb genotype t allele and TT genotype was associated with lung cancer risk in Caucasians. However, more studies should be conducted to confirm it.