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Abstract: The renin-angiotensin system (RAS) plays a central role in the pathogenesis of hypertension. Recently, we discovered that 1α,25-dihydroxyvitamin D3 [1,25(OH)2D3] functions as a negative endocrine regulator of renin biosynthesis, which provides a molecular basis to explore the potential of Vitamin D analogs as renin inhibitors to control the RAS. To search for renin-inhibiting Vitamin D analogs, we screened 20 Vitamin D analog compounds using As4.1-hVDR cell (a juxtaglomerular cell line) culture by Northern blot and luciferase reporter assays. We found that the Gemini compounds, which have two side-chains at carbon-20 position, were particularly active in suppressing renin expression. Eight Gemini compounds were identified that were equally or 10- to 100-times more potent than 1,25(OH)2D3 in renin inhibition. These Gemini compounds also potently stimulate 25-hydroxyvitamin D 24-hydroxylase expression in As4.1-hVDR cells. Administration of compound RO-27-5646 [1,25-dihydroxy-21-(3-methyl-3-hydroxy-butyl)-19-nor-cholecalciferol] in mice caused a marked reduction in renal renin mRNA expression without invoking severe hypercalcemia as seen in 1,25(OH)2D3 treatment. These data establish in principle that Vitamin D analogs can indeed inhibit renin expression in vitro and in vivo, and support the notion that low calcemic Vitamin D analogs can potentially be used as therapeutic agents to control the RAS. [Copyright &y& Elsevier]

Abstract: We have used mouse embryonic fibroblasts (MEFs) derived from VDR(+/−) and VDR(−/−) mice to determine whether the nuclear vitamin D receptor (VDR) is directly involved in the regulation of NF-κB activation. We found that the basal IκBα protein level was markedly decreased in VDR(−/−) MEFs compared to VDR(+/−) MEFs; however, degradation of IκBα and its phosphorylation were not altered in VDR(−/−) cells, neither were the levels of IKKα and IKKβ proteins. Consistently, p65 nuclear translocation was increased in unstimulated VDR(−/−) cells. The physical interaction between VDR and p65 was absent in VDR(−/−) MEFs, which may free p65 and increase its activity. Consequently, these alterations combined led to a marked increase in NF-κB transcriptional activity. Consistently, induction of IL-6 by TNFα or IL-1β was much more robust in VDR(−/−) than in VDR(+/−) cells, indicating that VDR(−/−) cells are more susceptible to inflammatory stimulation. Therefore, fibroblasts lacking VDR appear to be more pro-inflammatory due to the intrinsic high NF-κB activity. The reduction of IκBα in VDR(−/−) MEFs may be partially explained by the lack of VDR-mediated stabilization of IκBα by 1,25(OH)2D3. These data suggest that VDR plays an inhibitory role in the regulation of NF-κB activation. [Copyright &y& Elsevier]

Homocysteine (Hcy) is a synthetic amino acid containing sulfhydryl group, which is an intermediate product of the deep metabolic pathway of methionine and cysteine. The abnormal increase in fasting plasma total Hcy concentration caused by various factors is called hyperhomocysteine (HHcy). HHcy is closely relevant to the occurrence and progression of diverse cardiovascular and cerebrovascular diseases, such as coronary heart disease, hypertension and diabetes, etc. Vitamin D/vitamin D receptor (VDR) pathway is pointed out that prevent cardiovascular disease by reducing serum homocysteine levels. Our research is designed to explore the potential mechanism of vitamin D in the prevention and treatment of HHcy. The Hcy and 25(OH)D 3 levels in mouse myocardial tissue, serum or myocardial cells were detected using ELISA kits. The expression levels of VDR, Nrf2 and methionine synthase (MTR) were observed using Western blotting, immunohistochemistry and real time polymerase chain reaction (PCR). General information of the mice, including diet, water intake and body weight, was recorded. Vitamin D up-regulated the mRNA and protein expression of Nrf2 and MTR in mouse myocardial tissue and cells. CHIP assay determined that the combination of Nrf2 binding to the S1 site of the MTR promoter in cardiomyocytes using traditional PCR and real time PCR. Dual Luciferase Assay was applied to detect the transcriptional control of Nrf2 on MTR. The up-regulation effect of Nrf2 on MTR was verified by Nrf2 knockout and overexpression in cardiomyocytes. The role of Nrf2 in vitamin D inhibition of Hcy was revealed using Nrf2-knockdown HL-1 cells and Nrf2 heterozygous mice. Western blotting, real time PCR, IHC staining and ELISA showed that Nrf2 deficiency could restrain the increase in MTR expression and the decrease in Hcy level induced by vitamin D. The transcriptional activities of Nrf2/MTR were activated by vitamin D/VDR with a decrease in Hcy. Vitamin D/VDR upregulates MTR in an Nrf2-dependent manner, thereby reducing the risk of HHcy. [Display omitted] • Vitamin D relieved hyperhomocysteinemia. • Vitamin D could up-regulate Nrf2 and MTR. • MTR was a direct target gene of Nrf2. • Vitamin D/VDR could reduce myocardial homocysteine levels through activation of Nrf2 mediated MTR. [ABSTRACT FROM AUTHOR]

The renin–angiotensin system (RAS) plays a central role in the regulation of blood pressure, volume and electrolyte homeostasis. Inappropriate activation of the RAS may lead to hypertension. Clinical and epidemiological studies have suggested a correlation between Vitamin D-deficiency and high blood pressure. Our recent studies demonstrate that Vitamin D is a potent endocrine suppressor of renin biosynthesis to regulate the RAS. Mice lacking the Vitamin D receptor (VDR) have elevated production of renin and angiotensin (Ang) II, leading to hypertension, cardiac hypertrophy and increased water intake. These abnormalities can be prevented by treatment with an ACE inhibitor or AT1 receptor antagonist. Vitamin D repression of renin expression is independent of calcium metabolism, the volume- and salt-sensing mechanisms and the Ang II feedback regulation. In normal mice, Vitamin D-deficiency stimulates renin expression, whereas injection of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] reduces renin synthesis. In cell cultures, 1,25(OH)2D3 directly suppresses renin gene transcription by a VDR-dependent mechanism. Furthermore, we have found that Gemini compounds have more potent renin-suppressing activity than 1,25(OH)2D3. Collectively, our studies reveal a critical role of the Vitamin D endocrine system in the regulation of blood pressure and volume homeostasis, and suggest that low calcemic Vitamin D analogs may potentially be developed into a new class of anti-hypertensive agents to control renin production and blood pressure. [Copyright &y& Elsevier]