Original ContributionDietary vitamin D deficiency in rats from middle to old age leads to elevated tyrosine nitration and proteomics changes in levels of key proteins in brain: Implications for low vitamin D-dependent age-related cognitive decline
Graphical abstract
The figure depicts a potential mechanism for the protein nitration regulatory effects of VitD in brain and the consequences of VitD deficiency. Results obtained in this study supporting this mechanism are marked with wide green arrows, up arrows designating a found increase and down arrows designating a found decrease.
Introduction
The steroid hormone vitamin D (VitD) can be produced by the body or obtained through the diet. VitD is synthesized in the skin from the cholesterol precursor 7-dehydrocholesterol and is converted to cholecalciferol (VitD3) upon exposure to sunlight [1]. VitD3 can also be obtained through several dietary sources and is transported in the blood via vitamin D-binding protein. In the liver, VitD3 is converted to calcidiol, 25-hydroxyvitamin D (25-OH VitD), followed by further conversion to calcitriol, 1α,25-dihydroxyvitamin D (1α,25-(OH)2 VitD), primarily in the kidneys, where it helps to regulate calcium homeostasis [2], [3]. VitD also plays roles in autoimmunity [4], mental health [3], [5], [6], [7], [8], and inhibition of tumor growth through reductions in proliferation and angiogenesis [9], [10], [11], [12].
VitD deficiency has long been associated with osteoporosis, brittle bones, and muscle weakness, but recently low levels of VitD have been linked to increased overall mortality [13], [14]. VitD status is typically assessed using serum concentration of 25-OH VitD because it is longer lived than the biologically active 1α,25-(OH)2 VitD [13], [15], [16].
VitD deficiency is highly prevalent in Europe and North America [1], [17], with the elderly particularly at risk [11], [13], [18], [19], [20]. Current estimates suggest that as many as 40–100% of the elderly populations in these areas are VitD deficient [21]. Poor diet and lower exposure to UV-B from the sun limits VitD synthesis in the skin and an age-related decrease in the VitD synthesis machinery may contribute to the observed lower VitD levels [15].
The elderly represent those at greatest risk of age-related cognitive decline and neurodegenerative disorders [22]. Recent retrospective studies on elderly human subjects provide correlative evidence that those with VitD deficiency have a much higher incidence of cognitive impairment than those with normal VitD levels [23], [24]. Thus, it seems that VitD deficiency may accelerate cognitive decline in aging [25]. A recent meta-analysis also shows that patients with Alzheimer disease (AD) typically have lower serum concentrations of VitD [26]. AD is associated with defects in amyloid-β (Aβ) processing and an upregulation of inflammatory cytokines and nuclear factor κ-light-chain enhancer of activated B cells (NF-κB) [8]. Interestingly, 1α,25(OH)2 VitD helped to reverse soluble Aβ and inflammatory issues [8]. In addition to these actions, VitD is neuroprotective against Ca2+-mediated excitotoxicity, reduces biomarkers of brain aging associated with Ca2+ dyshomeostasis [3], [5], and helps to regulate levels of glutathione, a primary antioxidant in the brain, by modulating γ-glutamyltranspeptidase activity [27]. VitD also prevents onset of autoimmune demyelination in animal models of multiple sclerosis [28], [29].
Here, we manipulated serum VitD status by dietary supplementation with low, moderate/control, or high levels of VitD to identify changes in the VitD-dependent proteome in the brains of rats from middle to old age. Prior studies have shown that cognitively impaired subjects have significant levels of mitochondrial dysfunction and oxidative protein damage. In particular, nitration of protein-resident tyrosine residues is a common marker observed in brain of cognitively impaired subjects [30], [31], [32], [33], [34].Therefore, we tested the hypothesis that manipulating serum VitD levels would alter protein nitration and key protein markers of mitochondrial function. Our results identify several possible targets of VitD action that may mechanistically link circulating VitD levels with risk for age-related cognitive decline.
Section snippets
Chemicals
Criterion precast polyacrylamide gels, Tris–glycine–sodium dodecyl sulfate (SDS) (TGS) and Mes electrophoresis running buffers, ReadyStrip IPG strips, mineral oil, Precision Plus Protein All Blue standards, SYPRO Ruby protein stain, nitrocellulose membranes, dithiothreitol (DTT), iodoacetamide (IA), Biolytes, and urea were purchased from Bio-Rad (Hercules, CA, USA). Chemicals, proteases, protease inhibitors, and antibodies used in this study were purchased from Sigma–Aldrich (St. Louis, MO,
Vitamin D deficiency leads to increased nitrosative protein damage in brain
Tyrosine nitration is a common indicator/biomarker of the aging brain and of age-related neurodegenerative disorders [30], [34], both of which typically are accompanied by different extents of cognitive deficit. Here, we tested for indicators of oxidative and nitrosative stress in brain tissue samples from rats in which we manipulated serum VitD levels from middle age to old age. Significantly increased global 3-NT (Fig. 2) in the brains of rats on a low-VitD diet compared to rats on control or
Discussion
We have previously shown that tyrosine nitration occurs early in neurodegenerative processes, i.e., in mild cognitive impairment, arguably the earliest form of AD [31]. Nitration of tyrosine occurs from the reaction of NO with O2− through the reactive intermediate ONOO− in the presence of CO2 [32], [58], leading to tyrosine nitration by the NO2 radical. Nitrosative stress measures on these cortical samples showed approximately a 25% elevation in 3-NT globally in brain protein in the low-VitD
Conclusions
This study is the first to demonstrate that a chronic low-VitD diet and consequential low levels of VitD in the bloodstream result in significant increases in tyrosine nitration in brain proteins, alterations in glucose metabolism, and mitochondrial changes in brain of elderly rats, an animal model of brain in older human subjects (Fig. 8). A shift from the TCA cycle to glycolysis may be indicative of metabolic dysfunction. Further, ATP generated from glycolysis is important for maintaining a
Acknowledgment
This work was supported by the following grants from the National Institute on Aging: AG05119 (D.A.B.), AG010836 (P.W. Landfield, D.A.B., N.M.P.), AG034605 (P.W. Landfield, N.M.P.), and T32 AG0000242 (C.S.L., G. Gerhardt).
References (122)
Sunlight and vitamin D for bone health and prevention of autoimmune diseases, cancers, and cardiovascular disease
Am. J. Clin. Nutr.
(2004)- et al.
Chronic 1alpha,25-(OH)2 vitamin D3 treatment reduces Ca2+-mediated hippocampal biomarkers of aging
Cell Calcium
(2006) The vitamin D deficiency pandemic and consequences for nonskeletal health: mechanisms of action
Mol. Aspects Med.
(2008)- et al.
Vitamin D deficiency: a worldwide problem with health consequences
Am. J. Clin. Nutr.
(2008) - et al.
Low serum 25-hydroxyvitamin D concentrations are associated with greater all-cause mortality in older community-dwelling women
Nutr. Res.
(2009) - et al.
Vitamin D requirements: current and future
Am. J. Clin. Nutr.
(2004) - et al.
The urgent need to recommend an intake of vitamin D that is effective
Am. J. Clin. Nutr.
(2007) - et al.
Proteomics analysis provides insight into caloric restriction mediated oxidation and expression of brain proteins associated with age-related impaired cellular processes: mitochondrial dysfunction, glutamate dysregulation and impaired protein synthesis
Neurobiol. Aging
(2006) - et al.
Is vitamin D important for preserving cognition? A positive correlation of serum 25-hydroxyvitamin D concentration with cognitive function
Arch. Biochem. Biophys
(2007) - et al.
New clues about vitamin D functions in the nervous system
Trends Endocrinol. Metab.
(2002)
Sultana, R. Elevated levels of 3-nitrotyrosine in brain from subjects with amnestic mild cognitive impairment: implications for the role of nitration in the progression of Alzheimer's disease
Brain Res.
Differential expression and redox proteomics analyses of an Alzheimer disease transgenic mouse model: effects of the amyloid-beta peptide of amyloid precursor protein
Neuroscience
Slot-blot analysis of 3-nitrotyrosine-modified brain proteins
Methods Enzymol.
2-Mercaptoethane sulfonate prevents doxorubicin-induced plasma protein oxidation and TNF-α release: implications for the reactive oxygen species-mediated mechanisms of chemobrain
Free Radic. Biol. Med.
Evidence that light modulates protein nitration in rat retina
Mol. Cell. Proteomics
Expression of cytokine genes and increased nuclear factor-kappa B activity in the brains of scrapie-infected mice
Brain Res. Mol. Brain Res.
Tumor necrosis factor alpha-mediated nitric oxide production enhances manganese superoxide dismutase nitration and mitochondrial dysfunction in primary neurons: an insight into the role of glial cells
Neuroscience
Beta-amyloid stimulation of inducible nitric-oxide synthase in astrocytes is interleukin-1beta- and tumor necrosis factor-alpha (TNFalpha)-dependent, and involves a TNFalpha receptor-associated factor- and NFkappaB-inducing kinase-dependent signaling mechanism
J. Biol. Chem.
Tumour necrosis factor-alpha (TNF-alpha) increases nuclear factor kappaB (NFkappaB) activity in and interleukin-8 (IL-8) release from bovine mammary epithelial cells
Vet. Immunol. Immunopathol.
NF-kappaB, inflammation, and metabolic disease
Cell Metab.
Two pathways to NF-kappaB
Mol. Cell
Monocytes from type 2 diabetic patients have a pro-inflammatory profile: 1,25-dihydroxyvitamin D(3) works as anti-inflammatory
Diabetes Res. Clin. Pract.
Calcitriol inhibits TNF-alpha-induced inflammatory cytokines in human trophoblasts
J. Reprod. Immunol.
Redox proteomic identification of 4-hydroxy-2-nonenal-modified brain proteins in amnestic mild cognitive impairment: insight into the role of lipid peroxidation in the progression and pathogenesis of Alzheimer's disease
Neurobiol. Dis.
Identification of nitrated proteins in Alzheimer's disease brain using a redox proteomics approach
Neurobiol. Dis.
Park7, a novel locus for autosomal recessive early-onset parkinsonism, on chromosome 1p36
Am. J. Hum. Genet.
Parkinson disease-associated DJ-1 is required for the expression of the glial cell line-derived neurotrophic factor receptor RET in human neuroblastoma cells
J. Biol. Chem.
Proline cis-trans isomerization controls autoinhibition of a signaling protein
Mol. Cell
Inhibition of cell cycle progression by the novel cyclophilin ligand sanglifehrin A is mediated through the NFkappa B-dependent activation of p53
J. Biol. Chem
Cyclophilin A binds to peroxiredoxins and activates its peroxidase activity
J. Biol. Chem.
Removal of hydrogen peroxide by thiol-specific antioxidant enzyme (TSA) is involved with its antioxidant properties: TSA possesses thiol peroxidase activity
J. Biol. Chem.
Mammalian peroxiredoxin isoforms can reduce hydrogen peroxide generated in response to growth factors and tumor necrosis factor-alpha
J. Biol. Chem.
Protein tyrosine nitration—an update
Arch. Biochem. Biophys.
1alpha,25-Dihydroxyvitamin D3 inducible transcription factor and its role in the vitamin D action
Endocr. Regul.
Mounting evidence for vitamin D as an environmental factor affecting autoimmune disease prevalence
Exp. Biol. Med. (Maywood)
Vitamin D hormone confers neuroprotection in parallel with downregulation of L-type calcium channel expression in hippocampal neurons
J. Neurosci.
Correlation between serum 25-hydroxyvitamin D concentrations and regional cerebral blood flow in degenerative dementia
Nucl. Med. Commun.
Vitamin D, a neuro-immunomodulator: implications for neurodegenerative and autoimmune diseases
Psychoneuroendocrinology
1alpha,25-Dihydroxyvitamin D3 and resolvin D1 retune the balance between amyloid-beta phagocytosis and inflammation in Alzheimer's disease patients
J. Alzheimers Dis.
Vitamin D signalling pathways in cancer: potential for anticancer therapeutics
Nat. Rev. Cancer
Evidence-based D-bate on health benefits of vitamin D revisited
Dermatoendocrinology
Vitamin D deficiency and mortality
Curr. Opin. Clin. Nutr. Metab. Care
Vitamin D deficiency, atherosclerosis and cancer
Bratisl. Lek. Listy
The vitamin D deficiency pandemic: a forgotten hormone important for health
Public Health Rev
Resurrection of vitamin D deficiency and rickets
J. Clin. Invest.
Vitamin D and cognitive impairment in the elderly U.S. population
J. Gerontol. A Biol. Sci. Med. Sci.
Vitamin D deficiency
N. Engl. J. Med.
Serum 25-hydroxyvitamin D concentration and cognitive impairment
J. Geriatr. Psychiatry Neurol.
Vitamin D and risk of cognitive decline in elderly persons
Arch. Intern. Med.
Low serum vitamin D concentrations in Alzheimer's disease: a systematic review and meta-analysis
J. Alzheimers Dis.
Cited by (65)
Supplementation with vitamins D3 and a mitigates Parkinsonism in a haloperidol mice model
2024, Journal of Chemical NeuroanatomyAdult vitamin D deficiency and adverse brain outcomes
2023, Feldman and Pike's Vitamin D: Volume One: Biochemistry, Physiology and DiagnosticsUbiquitin carboxyl-terminal hydrolase L-1 in brain: Focus on its oxidative/nitrosative modification and role in brains of subjects with Alzheimer disease and mild cognitive impairment
2021, Free Radical Biology and MedicineCitation Excerpt :Then, in a rapid radical-radical recombination reaction, NO2 formed as from above binds to the free electron in the 3-position of Tyr to form 3-NT (Fig. 2), which is often detected in samples immunochemically by reaction with specific antibodies to protein-resident 3-NT [14]. Specificity of the antibody used is shown by the absence of binding to samples that had been pretreated with the powerful reducing reagent, Na2S2O4 that converts 3-NT to 3-amino-Tyr that is no longer recognized by antibodies against 3-NT [15]. When present on proteins, steric hindrance of 3-NT of the OH group on the aromatic ring of Tyr interferes with cell-signaling processes involving receptor tyrosine kinases, i.e.,3-NT is highly detrimental to intracellular signaling [16].
The treatment of Goji berry (Lycium barbarum) improves the neuroplasticity of the prefrontal cortex and hippocampus in aged rats
2020, Journal of Nutritional BiochemistryThe triangle of death of neurons: Oxidative damage, mitochondrial dysfunction, and loss of choline-containing biomolecules in brains of mice treated with doxorubicin. Advanced insights into mechanisms of chemotherapy induced cognitive impairment (“chemobrain”) involving TNF-α
2019, Free Radical Biology and MedicineVitamin D improves the antidiabetic effectiveness of aerobic training via modulation of Akt, PEPCK, and G6Pase expression
2023, Diabetology and Metabolic Syndrome