Brain vitamin D3-auto/paracrine system in relation to structural, neurophysiological, and behavioral disturbances associated with glucocorticoid-induced neurotoxicity.

Introduction: Vitamin D3 (VD3) is a potent para/autocrine regulator and neurosteroid that can strongly influence nerve cell function and counteract the negative effects of glucocorticoid (GC) therapy. The aim of the study was to reveal the relationship between VD3 status and behavioral, structural-functional and molecular changes associated with GC-induced neurotoxicity. Methods: Female Wistar rats received synthetic GC prednisolone (5 mg/kg b.w.) with or without VD3 (1000 IU/kg b.w.) for 30 days. Behavioral, histological, physiological, biochemical, molecular biological (RT-PCR, Western blotting) methods, and ELISA were used. Results and discussion: There was no difference in open field test (OFT), while forced swim test (FST) showed an increase in immobility time and a decrease in active behavior in prednisolone-treated rats, indicative of depressive changes. GC increased the perikaryon area, enlarged the size of the nuclei, and caused a slight reduction of cell density in CA1-CA3 hippocampal sections. We established a GC-induced decrease in the long-term potentiation (LTP) in CA1-CA3 hippocampal synapses, the amplitude of high K+-stimulated exocytosis, and the rate of Ca2+-dependent fusion of synaptic vesicles with synaptic plasma membranes. These changes were accompanied by an increase in nitration and poly(ADP)-ribosylation of cerebral proteins, suggesting the development of oxidative-nitrosative stress. Prednisolone upregulated the expression and phosphorylation of NF-κB p65 subunit at Ser311, whereas downregulating IκB. GC loading depleted the circulating pool of 25OHD3 in serum and CSF, elevated VDR mRNA and protein levels but had an inhibitory effect on CYP24A1 and VDBP expression. Vitamin D3 supplementation had an antidepressant-like effect, decreasing the immobility time and stimulating active behavior. VD3 caused a decrease in the size of the perikaryon and nucleus in CA1 hippocampal area. We found a recovery in depolarization-induced fusion of synaptic vesicles and long-term synaptic plasticity after VD3 treatment. VD3 diminished the intensity of oxidative-nitrosative stress, and suppressed the NF-κB activation. Its ameliorative effect on GC-induced neuroanatomical and behavioral abnormalities was accompanied by the 25OHD3 repletion and partial restoration of the VD3-auto/paracrine system. Conclusion: GC-induced neurotoxicity and behavioral disturbances are associated with increased oxidative-nitrosative stress and impairments of VD3 metabolism. Thus, VD3 can be effective in preventing structural and functional abnormalities in the brain and behavior changes caused by long-term GC administration.

The link between vitamin D status and NF-κB-associated renal dysfunction in experimental diabetes mellitus.

BACKGROUND: Type 1 diabetes (T1D) is accompanied by numerous side effects, including renal dysfunction. Mounting evidence suggests that overactivation of nuclear factor ĸB (NF-κB) is one of the key triggers of diabetes-associated chronic kidney disease. Vitamin D3 is considered as a strong modulator of a number of transcription factors, including NF-κB. The purpose of our study was to assess the contribution of NF-κB to type 1 diabetes (T1D)-induced kidney dysfunction and to determine if vitamin D3 supplementation can correct the changes associated with T1D. METHODS: The following animal groups were used: control, diabetic (induced by single i.p. injection of streptozocin at dose 55 mg/kg b.w.), T1D group treated with vitamin D3 (600 IU/kg b.w.), T1D group treated with NF-κB inhibitor - BAY 11-7082. RESULTS: Diabetes led to a decrease in serum 25(OH)D that was accompanied by down-regulation of vitamin D receptor (VDR) expression and up-regulation of hydroxylases CYP24A1 and CYP27B1 synthesis in the kidneys. Diabetes activated the transcription factor NF-κB and increased cleaved (p17) caspase-3 level in renal tissue. Restoration of vitamin D status normalized vitamin D-endocrine system, decreased NF-κB activation and caspase-3 protein level in the kidneys of diabetic animals. BAY 11-7082 partially mimicked the effects of vitamin D3. GENERAL SIGNIFICANCE: Vitamin D3 supplementation counteracts diabetes-induced kidney damage, most likely through VDR-mediated inhibition of NF-κB activation.

Vitamin D Auto-/Paracrine System Is Involved in Modulation of Glucocorticoid-Induced Changes in Angiogenesis/Bone Remodeling Coupling.

Osteoporosis is a devastating side effect of chronic glucocorticoid (GC) treatment. Despite the crucial role of vitamin D (VD) in bone homeostasis, the precise molecular mechanisms of its action on GC-induced disturbances of bone remodeling remain undefined. The study was performed to elucidate the relation of VD status to GC-induced changes of the angiogenesis/osteogenesis/bone resorption coupling in bone tissue. Female Wistar rats received prednisolone (5 mg/kg of b.w.) with or without VD3 (1000 IU/kg of b.w., for 30 days). Biomechanical parameters of rat femurs were assessed by the three-point bending test. The levels of calcium, inorganic phosphate, activity of total alkaline phosphatase (ALP), and its isoenzymes were determined spectrophotometrically. Vascular endothelial growth factor-A (VEGF-A) and caspase-3 protein levels were detected by western blotting. Vdr and Cyp27b1 mRNAs were measured by qRT-PCR. Receptor activator of nuclear factor κB (RANK) expression in bone sections was visualized immunohistochemically. Serum 25(OH)D was assayed by ELISA. GC administration led to a decrease in maximal load (by 1.2-fold) and stiffness and toughness (by 1.3-fold), which was accompanied by a 3-fold reduction of 25(OH)D level, an elevation of the ALP bone isoenzyme activity in serum, hypocalcaemia, and hypophosphatemia. Along with prednisolone-induced VD deficiency, an impaired synthesis of Vdr (-30%) and Cyp27b1 (+71%) mRNA was observed, reflecting deregulation of bone tissue VD-auto-/paracrine system. GC caused an increase in caspase-3 content, suppressed the synthesis of the osteoclastic marker RANK, and altered angiogenesis/osteogenesis coupling by significantly reducing the level of VEGF-A.VD3 treatment restored serum 25(OH)D content and the expression of key components of the VD-auto-/paracrine system. VD3 supplementation diminished cell apoptosis and strongly improved angiogenesis/osteogenesis coupling as well as mineral metabolism and biomechanical parameters of femurs in GC-administered rats. Thus, VD3 can have a beneficial effect on the correction of GC-induced pathological changes in bone remodeling.

Vitamin D3 Modulates Impaired Crosstalk Between RANK and Glucocorticoid Receptor Signaling in Bone Marrow Cells After Chronic Prednisolone Administration.

The effectiveness of vitamin D3 (cholecalciferol) in counteracting the side effects of glucocorticoid (GC) therapy has been demonstrated previously. Abnormalities in systemic hormonal and local (cytokine) regulation of bone marrow (BM) cells may underlie GC-induced imbalance between osteosynthesis and bone resorption. The cytokine system receptor activator of nuclear factor kappa-B (RANK)/RANK ligand (RANKL)/osteoprotegerin (OPG) is considered as an integrating link in the NF-κB-mediated interaction of various cells involved in maintaining osteoblastic-osteoclastic balance, which makes it a pharmacological target for regulation and correction of the bone remodeling process. We studied GC-induced impairments of the RANKL/RANK/OPG axis in BM cells depending on vitamin D bioavailability and whether these changes were mediated by glucocorticoid (GR) and/or vitamin D (VDR) receptors. Female Wistar rats administered with prednisolone (5 mg/kg b.w., 30 days) showed a decrease in the GR protein level and the number of GR-positive BM cells. GC caused a marked elevation of RANKL and RANK levels in BM, while OPG decreased. Flow cytometry data indicated GC-elicited increase in the number of circulating RANK-positive osteoclast precursors (OCPs) in BM, peripheral blood, and spleen. In full accordance with the data that the interaction of RANKL-RANK leads to transcriptional activation of NF-κB and subsequent differentiation of osteoclasts, we found an increase in the level of phosphorylated p65 subunit of NF-κB with a simultaneous decrease in the NF-κB inhibitor (IκB) level. These changes were accompanied by vitamin D insufficiency and downregulated expression of CYP27B1 and VDR, which are responsible for synthesis and hormonal signaling of 1,25(OH)2D. Notably, we observed VDR and RANK co-localization in OCPs. Cholecalciferol co-administration (1,000 IU/kg b.w., 30 days) with prednisolone resulted in elevated GR synthesis in BM. Cholecalciferol prevented prednisolone-elicited disturbances of the RANKL/RANK/OPG, which correlated with improved bioavailability and vitamin D signaling through VDR. This caused the lowering of phosphoNF-κB p65 level and inhibiting NF-κB translocation to the nucleus that could reduce the circulating OCPs pool in BM, peripheral blood, and spleen. Our findings suggest that prednisolone-induced abnormalities in GR and RANKL/RANK/OPG signaling pathways are associated with the impairments of vitamin D auto/paracrine system in BM cells and can be ameliorated by cholecalciferol supplementation.

Effects of Cholecalciferol on Key Components of Vitamin D-Endo/Para/Autocrine System in Experimental Type 1 Diabetes.

OBJECTIVES: Recent prospective studies have found the associations between type 1 diabetes (T1D) and vitamin D deficiency. We investigated the role of vitamin D in the regulation of 25OHD-1α-hydroxylase (CYP27B1) and VDR expression in different tissues of T1D rats. DESIGN: T1D was induced in male Wistar rats by streptozotocin (55 mg/k b.w.). After 2 weeks of T1D, the animals were treated orally with or without vitamin D3 (cholecalciferol; 100 IU/rat, 30 days). METHODS: Serum 25-hydroxyvitamin D (25OHD) was detected by ELISA. CYP27A1, CYP2R1, CYP27B1, and VDR were assayed by RT-qPCR and Western blotting or visualized by immunofluorescence staining. RESULTS: We demonstrated that T1D led to a decrease in blood 25OHD, which is probably due to the established downregulation of CYP27A1 and CYP2R1 expression. Vitamin D deficiency was accompanied by elevated synthesis of renal CYP27B1 and VDR. Conversely, CYP27B1 and VDR expression decreased in the liver, bone tissue, and bone marrow. Cholecalciferol administration countered the impairments of the vitamin D-endo/para/autocrine system in the kidneys and extrarenal tissues of diabetic rats. CONCLUSIONS: T1D-induced vitamin D deficiency is associated with impairments of renal and extrarenal CYP27B1 and VDR expression. Cholecalciferol can be effective in the amelioration of diabetes-associated abnormalities in the vitamin D-endo/para/autocrine system.