Vitamin D Hormone Research Articles

Post-ischemic stroke systemic inflammation: Immunomodulation by progesterone and vitamin D hormone

Post-stroke systemic inflammation, due to the injury itself and exacerbated by in-hospital infections, can increase morbidity and mortality in stroke patients. In this study, we examined the immunomodulatory effects of progesterone (P4) alone and in combination with vitamin D hormone (VDH) on acute phase post-stroke peripheral immune dysfunction and functional/behavioral deficits. Adult rats underwent transient middle cerebral artery occlusion/reperfusion (tMCAO) and delayed systemic inflammation was induced by injections of lipopolysaccharide (LPS) beginning 24 h post-stroke. Animals were tested for behavioral outcomes and immune function at day 4 post-stroke. We also measured infarction volume and markers of neuronal inflammation (GFAP, IL-6) and apoptosis (cleaved caspase-3) in brain post-stroke. We observed the worst stroke outcomes in the stroke + systemic inflammation group compared to the stroke-alone group. Flow cytometric analysis of different subsets of immune cells in blood, spleen and thymus revealed peripheral immune dysfunction which was restored by both P4 and VDH monotherapy. P4 monotherapy reduced infarction volume, behavioral/functional deficits, peripheral immune dysfunction, neuronal inflammation, and apoptosis induced by post-stroke systemic inflammation. Combination treatment with P4+VDH improved outcomes better than monotherapy. Our findings can be taken to suggest that the current standard of care for stroke and post-stroke infection can be substantially improved by P4 and VDH combination therapy.

Abstract TMP32: Post-Stroke Systemic Inflammation: Immunomodulation by Progesterone and Vitamin D Hormone

Introduction: Post-stroke systemic inflammation due to in-hospital infections increases patient morbidity and mortality. We examined the immunomodulatory effects of progesterone (P4) either alone or in combination with vitamin D hormone (VDH) on post-stroke systemic inflammation-induced peripheral immune dysfunction and functional deficits. Methods: Adult male Sprague-Dawley rats underwent transient middle cerebral artery occlusion/reperfusion (tMCAO) or sham surgery. Delayed systemic inflammation was induced by injections of lipopolysaccharide (LPS; 50 μg/kg; IP) at 24, 28 and 32 h post-occlusion. P4 (8 mg/kg; IP) and VDH (1 μg/kg; IP) were given 5 min prior to reperfusion followed by SC injections on days 1, 2 and 3 post-stroke. Power analysis specified a sample size of 6 rats/group to detect a 30% difference between treatment groups with a power of 80% at alpha 0.05. There were 6 groups (n=6/gp): 1) Sham + vehicle (SHAM); 2) tMCAO + vehicle (VEH); 3) tMCAO + vehicle + LPS (VEH + LPS); 4) tMCAO + LPS + P4 (P4); 5) tMCAO + LPS + VDH (VDH); 6) tMCAO + LPS + P4 + VDH (P4 + VDH). At day 4 post-stroke, rats were tested on spontaneous locomotor activity, grip strength and sensory neglect; peripheral immune dysfunctions were assessed by flow cytometric analysis of subsets of immune cells in blood, spleen and thymus. We measured markers of neuronal inflammation (GFAP, Iba-1, IL-6), apoptosis (cleaved caspase-3, TUNEL), and endoplasmic reticulum (ER) stress (PERK, eIF2α, CHOP) in brain. Data were analyzed using ANOVA with the Tukey-HSD post hoc test. Statistical significance was set at p <0.05. Results: The worst stroke outcomes were seen in the VEH + LPS group. Peripheral immune dysfunctions revealed by flow cytometry were significantly ( p <0.05) restored by both P4 and VDH monotherapy. While P4 and VDH individually improved functional deficits, neuronal inflammation, apoptosis and ER stress induced by post-stroke systemic inflammation, the combination of P4 + VDH improved outcomes significantly ( p <0.05) more than monotherapy. Conclusions: Our findings may have translational relevance because the current standard of care for stroke and post-stroke infections is largely ineffective and our P4+VDH combination therapy may hold promise for stroke therapeutics.

Steroid Hormone Vitamin D: Implications for Cardiovascular Disease.

Understanding of vitamin D physiology is important because about half of the population is being diagnosed with deficiency and treated with supplements. Clinical guidelines were developed based on observational studies showing an association between low serum levels and increased cardiovascular risk. However, new randomized controlled trials have failed to confirm any cardiovascular benefit from supplementation in the general population. A major concern is that excess vitamin D is known to cause calcific vasculopathy and valvulopathy in animal models. For decades, administration of vitamin D has been used in rodents as a reliable experimental model of vascular calcification. Technically, vitamin D is a misnomer. It is not a true vitamin because it can be synthesized endogenously through ultraviolet exposure of the skin. It is a steroid hormone that comes in 3 forms that are sequential metabolites produced by hydroxylases. As a fat-soluble hormone, the vitamin D-hormone metabolites must have special mechanisms for delivery in the aqueous bloodstream. Importantly, endogenously synthesized forms are carried by a binding protein, whereas dietary forms are carried within lipoprotein particles. This may result in distinct biodistributions for sunlight-derived versus supplement-derived vitamin D hormones. Because the cardiovascular effects of vitamin D hormones are not straightforward, both toxic and beneficial effects may result from current recommendations.

Progesterone and vitamin D: Improvement after traumatic brain injury in middle-aged rats

Progesterone (PROG) and vitamin D hormone (VDH) have both shown promise in treating traumatic brain injury (TBI). Both modulate apoptosis, inflammation, oxidative stress, and excitotoxicity. We investigated whether 21days of VDH deficiency would alter cognitive behavior after TBI and whether combined PROG and VDH would improve behavioral and morphological outcomes more than either hormone alone in VDH-deficient middle-aged rats given bilateral contusions of the medial frontal cortex. PROG (16mg/kg) and VDH (5μg/kg) were injected intraperitoneally 1h post-injury. Eight additional doses of PROG were injected subcutaneously over 7days post-injury. VDH deficiency itself did not significantly reduce baseline behavioral functions or aggravate impaired cognitive outcomes. Combination therapy showed moderate improvement in preserving spatial and reference memory but was not significantly better than PROG monotherapy. However, combination therapy significantly reduced neuronal loss and the proliferation of reactive astrocytes, and showed better efficacy compared to VDH or PROG alone in preventing MAP-2 degradation. VDH+PROG combination therapy may attenuate some of the potential long-term, subtle, pathophysiological consequences of brain injury in older subjects.

Combination treatment with progesterone and vitamin D hormone is more effective than monotherapy in ischemic stroke: The role of BDNF/TrkB/Erk1/2 signaling in neuroprotection

We investigated whether combinatorial post-injury treatment with progesterone (P4) and vitamin D hormone (VDH) would reduce ischemic injury more effectively than P4 alone in an oxygen glucose deprivation (OGD) model in primary cortical neurons and in a transient middle cerebral artery occlusion (tMCAO) model in rats. In the OGD model, P4 and VDH each showed neuroprotection individually, but combination of the “best” doses did not show substantial efficacy; instead, the lower dose of VDH in combination with P4 was the most effective. In the tMCAO model, P4 and VDH were given alone or in combination at different times post-occlusion for 7 days. In vivo data confirmed the in vitro findings and showed better infarct reduction at day 7 and functional outcomes (at 3, 5 and 7 days post-occlusion) after combinatorial treatment than when either agent was given alone. VDH, but not P4, upregulated heme oxygenase-1, suggesting a pathway for the neuroprotective effects of VDH differing from that of P4. The combination of P4 and VDH activated brain-derived neurotrophic factor and its specific receptor, tyrosine kinase receptor-B. Under specific conditions VDH potentiates P4's neuroprotective efficacy and should be considered as a potential partner of P4 in a low-cost, safe and effective combinatorial treatment for stroke.

Progesterone and low-dose vitamin D hormone treatment enhances sparing of memory following traumatic brain injury

Progesterone (PROG) has been shown to protect the brain from traumatic injury and is now in Phase III clinical trials. Our work shows that PROG's beneficial effects can be reduced in vitamin D hormone (VDH)-deficient subjects. VDH can modulate neuronal apoptosis, trophic factors, inflammation, oxidative stress, excitotoxicity, and myelin and axon repair. We investigated whether VDH combined with PROG could improve behavioral outcomes more than PROG alone in VDH-sufficient rats given bilateral contusions of the medial frontal cortex. PROG and different doses of VDH (1μg/kg, VDH1; 2.5μg/kg, VDH2; 5μg/kg, VDH3) were injected intraperitoneally 1h post-injury. Eight additional doses of PROG were given subcutaneously over 8days with tapering over the last 2days. Neurobehavioral tests, necrotic cavity, neuronal death and activation of astrocytes were evaluated 21days post-injury. We found that PROG and PROG+VDH preserve spatial memory processing. VDH1+PROG improved performance in acquisition more effectively than PROG alone, indicating that the low VDH dose is optimal for combination therapy. There were no significant differences in necrotic cavity size among the groups. The density of positive staining for reactive astrocytes (glial fibrillary acidic protein (GFAP)) increased and the cell bodies and processes of GFAP-positive cells were enlarged in the PROG+VDH1 group. Our data indicate that the combination of PROG and VDH is more effective than PROG alone in preserving spatial and reference memory, and that PROG plus low-dose VDH can activate GFAP reactions up to 21days after injury. This effect may be one of the mechanisms underlying PROG's neuroprotective effects in combination with VDH.

Lovastatin exhibits synergy with Vitamin D hormone to confer protection from experimental autoimmune encephalomylitis (148.5)

Abstract Recent studies provide sufficient evidence that disease modifying agents (DMAs) i.e., vitamin D hormone (VDH) and statin (lovastatin) were immunomodulatory to attenuate experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS). Interestingly, irrespective of their different mechanism of action these DMAs produce similar protective activities to attenuate EAE. Therefore, we asked whether statin regulate the vitamin D precursor (VDP; vitamin D3) homeostasis in treated EAE model. Interestingly, we observed a significant increase in the serum level of both VDP and VDH in lovastatin treated EAE animals as compared to those treated with vehicle. It was associated with diminished level of CYP24A1 (VDH inactivating enzyme) and elevated level of CYP27B1 (VDH synthesizing enzyme) involved in VDH metabolism in the blood cells and spinal cord of EAE treated with lovastatin than vehicle. Next, we tested the potential of lovastatin and VDH in combination to confer protection from EAE. Interestingly, suboptimal doses of these DMAs significantly attenuated acute clinical exacerbations, cellular infiltration, inflammatory response, axonal loss and demyelination in the spinal cord of EAE animals compared with their individual treatment analyzed by using quantitative and qualitative analyses. Altogether, these findings provide initial evidence that lovastatin maintains VDH homeostasis and its synergy with VDH enhances neuroprotection in EAE/MS brain.

Vitamin D metabolism in thyrotoxicosis. Therapeutic aspects derived from an old observation

To the Editor: In an article of 2007 about vitamin D deficiency, it was reported that low 25OH-vitD (25hydroxyvitaminD) plasma levels are found in thyrotoxicosis, with thyrotoxicosis being the last in the list of acquired causes of vitamin D deficiency (1). We first demonstrated low 25OH-vitD plasma levels in thyrotoxicosis (2), as well as the fact that the transformation of vitamin D into 25OH-vitD is 2.5 to eight times faster in thyrotoxic subjects in comparison to healthy controls (3). In an unpublished report, the acceleration of hydroxylation of vitamin D into 25OH-vitD in thyrotoxic subjects (HC) was depicted against that of healthy controls (HC) (Figure 1). 25hydroxyvitaminD plasma levels estimated the same period in three thyrotoxic subjects (TS) and three healthy controls (HC), all males matched for age and body weight after i.v. injection of 1 mg of vitamin D When our observations were made, the lower normal limits of 25OH-vitD plasma levels were ill defined as various cut points were used for a long time to define vitamin D inadequacy fluctuating from 9 ng/ml to 30 ng/ml (4). Therefore, it was inconvenient to consider hypovitaminosis D with 25OH-vitD plasma levels between 9 ng/ml and 30 ng/ml. These nebulous data about the lower normal limits of 25OH-vitD plasma levels resulted in the lack of concern regarding the acceleration of vitamin D metabolism in thyrotoxicosis, although the osteomalacic component has been mentioned long ago in thyrotoxic bone disease (5). In recent years, a level of 30 ng/ml or greater of 25OH-vitD plasma levels was approved definitely as the lower normal level in humans, while the toxic levels were defined as greater than 150 ng/ml (1). The new data validated the ascertained acceleration of vitamin metabolism in thyrotoxicosis as thyrotoxic subjects are at risk for drop of 25OH-vitD plasma levels below the level of 30 ng/ml. In addition, hyperthyroidism is an endocrinopathy, which may run for a long time without the patient seeking medical intervention. Hence, the vitamin D depot in the body may decline in the levels of inadequacy or overt deficiency with the all known consequences. Especially prone to this risk are the thyrotoxic subjects of old age or institutionalised as well as those confined at home. A similar condition may be observed in patients taking suppressive doses of thyroxine or incorrectly increased doses of replacement therapy. The aim of this issue is to establish in all patients who are under the influence of increased thyroid hormones, a testing of their vitamin D status parallel to the testing of their thyroid status, as well as the follow-up of their 25OH-vitD plasma levels during the medical supervision of the patient. The intervention with vitamin D when insufficiency or deficiency of this vitamin ensues will avert the deleterious effect of this inadequacy especially to the bones. This substance ‘vitamin D-hormone’, on the basis of the above mentioned, must be considered as a serious agent in the restoration of health after the elapse of toxicosis in thyrotoxic patients, given the fact that many are unaware of this kind of inadequacy in thyrotoxic subjects.

Potential of alfacalcidol for reducing increased risk of falls and fractures

There are no general accepted strategies for combined drug treatments in osteoporosis, while in other important chronic diseases combinations of different medications are used as a rule to improve therapeutic results and reduce the risk of adverse events. It is suggested that the success of combined treatments is related to the different modes of action of the respective single therapies. On the other hand it was shown that a strong antiresorptive bisphosphonate is able to blunt at least in part the effects of anabolic parathyroid hormone peptides Calcitriol, the active vitamin D-hormone and its prodrug alfacalcidol lead to pleiotropic effects on bone remodelling (antiresorptive, anabolic and enhancing mineralization) and in addition to effects on other important target tissues (e.g. gut, parathyroid glands, muscle). With active D-analogs significant improvements in the therapeutic outcome of osteoporosis can be achieved by the resulting improvements of bone quality, calcium absorption and risk reduction of falling. The same beneficial effects cannot be achieved with plain vitamin D due to feedback controlled, limited renal activation or insufficient conversion in the elderly with impairment of renal function. Accordingly alfacalcidol, approved as a treatment for different forms of osteoporosis, is besides adoption as a mono-therapy an interesting candidate for combined therapies. There are interesting preclinical trials and clinical pilote studies in the literature proving that a parallel therapy with selectively anti-osteoclastic bisphophonates and pleiotropically acting D-analogs is able to optimize therapeutic results in osteoporosis. In the AAC-Trial (Alfacalcidol-Alendronate-Combined) we studied 90 patients with established osteoporosis (57 women, 33 men) over two years after alternate allocation to three treatment arms (alfacalcidol plus calcium, alendronate plus plain vitamin D and Ca, and alendronate plus alfacalcidol and Ca). During the 2-year-study we observed the significantly highest lumbar spine and hip BMD increases in the combined treatment group (p < 0.001). The number of patients with new vertebral and non-vertebral fractures after 2 years was 9 with alfacalcidol alone, 10 with alfacalcidol and plain vitamin D and 2 in the group receiving alendronate plus alfacalidol (p < 0.02). Furthermore there was a lower rate of falls and an earlier reduction in back pain in the patients treated with the active combination. This trial confirms the demonstrated highly significant advantages of this combined treatment regimen used in the pilote studies. Especially in patients with severe osteoporosis this interesting combination of two substances with complete different mechanisms of action should be taken into consideration.

Differential Effects of D-Hormone Analogs and Native Vitamin D on the Risk of Falls: A Comparative Meta-Analysis

The aim of this meta-analysis was to compare the antifall efficacy of native vitamin D to that of its hydroxylated analogs alfacalcidol and calcitriol. Randomized clinical trials comparing oral native vitamin D and its analogs alfacalcidol and calcitriol to a placebo were included. Sources included the Cochrane Controlled Trials Register, EMBASE, MEDLINE, a hand search of abstracts, as well as reference lists. The time range was January 1995 to May 2007. Data were abstracted and scored by two investigators. The core analysis was based on double-blind trials, while open trials were included as a robustness analysis. Relative risks (RRs) for falls while allocated to D-hormone analogs or vitamin D were calculated. Publication bias and robustness were formally tested. Fourteen trials including 21,268 subjects were included. Using double-blind data only, vitamin D-hormone analogs provided a statistically significant lower level of risk for falling compared to native vitamin D: RR = 0.79 (95% confidence interval 0.64-0.96) vs. 0.94 (0.87-1.01) (intergroup difference P = 0.049). The dropout rates observed in the two sets of trials were comparable: 0.33% per month. Publication bias investigation did not report any significant trend for selective publication favoring active treatment arms. Upon current evidence, D-hormone analogs seem to prevent falls to a greater extent than their native compound. Long-term, prospective, head-to-head, confirmatory trials are required to address the exact role of vitamin D and D-hormone analogs in the prevention of falls and fractures.

Influence of hormones on osteogenic differentiation processes of mesenchymal stem cells

Bone development, regeneration and maintenance are governed by osteogenic differentiation processes from mesenchymal stem cells through to mature bone cells, which are directed by local growth and differentiation factors and modulated strongly by hormones. Mesenchymal stem cells develop from both mesoderm and neural crest and can give rise to development, regeneration and maintenance of mesenchymal tissues, such as bone, cartilage, muscle, tendons and discs. There are only limited data regarding the effects of hormones on early events, such as regulation of stemness and maintenance of the mesenchymal stem cell pool. Hormones, such as estrogens, vitamin D-hormone and parathyroid hormone, besides others, are important modulators of osteogenic differentiation processes and bone formation, starting off with fate decision and the development of osteogenic offspring from mesenchymal stem cells, which end up in osteoblasts and osteocytes. Hormones are involved in fetal bone development and regeneration and, in childhood, adolescence and adulthood, they control adaptive needs for growth and reproduction, nutrition, physical power and crisis adaptation. As in other tissues, aging in mesenchymal stem cells and their osteogenic offspring is accompanied by the accumulation of genomic and proteomic damage caused by oxidative burden and insufficient repair. Failsafe programs, such as apoptosis and cellular senescence avoid tumorigenesis. Hormones can influence the pace of such events, thus supporting the quality of tissue regeneration in aging organisms in vivo; for example, by delaying osteoporosis development. The potential for hormones in systemic therapeutic strategies is well appreciated and some concepts are approved for clinical use already. Their potential for cell-based therapeutic strategies for tissue regeneration is probably underestimated and could enhance the quality of tissue-engineering constructs for transplantation and the concept of in situ-guided tissue regeneration.

A dialogue on analogues Newer vitamin-D drugs for use in bone disease, psoriasis, and cancer

Many new analogues of the vitamin-D hormone, 1 α,25-dihydroxy-vitamin D 3 [1 α,25-(OH) 2D 3; calcitriol], have emerged that can mimic its various actions in classic calcium transport systems and/or in the regulation of cell proliferation and cell differentiation. Though some of these analogues have accentuated activity in cell differentiation assays in vitro, they lack appreciable “calcemic” activity in vivo, leading to the name “noncalcemic analogues.” Several of these analogues are promising candidates for use in treatment of psoriasis and in tumor suppression, one of them, calcipotriol, being already widely approved for the former indication. New generations of calcemic analogues with altered pharmacokinetics are appearing for use in secondary hyperparathyroidism and osteoporosis. We believe that the selective properties of both types of analogues stem from altered receptor binding, blood protein binding, and rate of catabolism.

Calbindin D 28K Regulation in Precociously Matured Chick Egg Shell Gland in Vitro

Egg shell calcification in the hen uterus (egg shell gland, ESG) depends primarily on intestinal absorption of dietary Ca 2+ as well as ESG Ca 2+ transport into the shell. Intestinal Ca 2+ absorption is linked to vitamin D-induced calbindin D 28K (D28K) concentration. The ESG also contains D28K, and Ca 2+ transport into the shell appears to be linked to D28K gene expression, but until this report, there was no direct proof that ESG D28K was or was not vitamin D-dependent. To address this issue, highly developed ESG from estradiol (E 2)injected, severely vitamin D-depleted chicks were cultured in serum-free medium with excellent viability. Addition of the vitamin D-hormone, 1,25(OH) 2 vitamin D 3 (1,25), to the culture medium increased ESG D28K levels as much as 70%. E 2 alone had no effect, but E 2 plus 1,25 further increased ESG D28K levels up to 160%. By contrast, progesterone (P 4) prevented the 1,25-stimulated increase in D28K, while having no effect on basal D28K level. Of considerable interest, thapsigargin (THAPS), which increases intracellular Ca 2+ concentration ([Ca 2+]i) in many cell types, stimulated D28K synthesis in a concentration-dependent manner in the complete absence of 1,25 and independent of the [Ca 2+] of the medium. These results are the first direct evidence that ESG D28K is under direct control of 1,25 and that both gonadal steroid hormones, E 2 and P 4, may be coregulators. Further, the effects of THAPS suggest that [Ca 2+]i itself may also regulate D28K. This new in vitro model clearly represents a unique opportunity to study the regulation of the ESG calcium transport mechanism under stringently defined conditions.

Reduction of vitamin D hormone receptor mRNA levels in Alzheimer as compared to Huntington hippocampus: correlation with calbindin-28k mRNA levels

Receptors for vitamin D hormone (VDR) and the calcium binding protein, calbindin-28k, have been localized in many tissues, including brain. In brain, VDR and calbindin-28k were reported to colocalize in hippocampal CA1 cells. We have shown that mRNA pool size for calbindin-28k was reduced, on average, by 35% in Alzheimer hippocampal CA1 cells, as compared to Huntington control (manuscript in preparation). In the present study, in situ hybridization with tritiated antisense RNA probes was used to examine VDR expression in paired Alzheimer and Huntington brain tissue. Message levels for VDR were reduced, on average, by 34% and 31%, respectively, in Alzheimer hippocampal CA1 and CA2 pyramidal cells, as compared to Huntington control. However, VDR message levels were not significantly different from control in Alzheimer temporal cortex or cerebellum. There was no correlation between VDR message levels and brain weight, autopsy interval, patient age or the extent of neurofibrillary degeneration. Instead, VDR mRNA pool size in hippocampal CA1 cells correlated significantly with calbindin-28k message levels ( r = 0.52, P < 0.001). Decreased message levels for VDR and calbindin-28k in these cells were due to an increased percentage of cells expressing lower message levels for these proteins. These results show that in Alzheimer hippocampal CA1 cells, VDR mRNA pool size is downregulated and that this downregulation may play a role in the reduction of calbindin-28k expression.