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Vitamin D prevents hypoxia/reoxygenation-induced blood-brain barrier disruption via vitamin D receptor-mediated NF-kB signaling pathways.

Won S, Sayeed I, Peterson BL, Wali B, Kahn JS, Stein DG - PLoS ONE (2015)

Bottom Line: We analyzed the effect of 1,25(OH)2D3 on brain endothelial cell barrier integrity and tight junction proteins after hypoxia/reoxygenation in a mouse brain endothelial cell culture model that closely mimics many of the features of the blood-brain barrier in vitro.These responses were blocked when the interaction of 1,25(OH) )2D3 with the vitamin D receptor (VDR) was inhibited by pyridoxal 5'-phosphate treatment.Our findings show a direct, VDR-mediated, protective effect of 1,25(OH) )2D3 against ischemic injury-induced blood-brain barrier dysfunction in cerebral endothelial cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Emergency Medicine Brain Research Laboratory, Emory University, Atlanta, Georgia, United States of America.

ABSTRACT
Maintaining blood-brain barrier integrity and minimizing neuronal injury are critical components of any therapeutic intervention following ischemic stroke. However, a low level of vitamin D hormone is a risk factor for many vascular diseases including stroke. The neuroprotective effects of 1,25(OH)2D3 (vitamin D) after ischemic stroke have been studied, but it is not known whether it prevents ischemic injury to brain endothelial cells, a key component of the neurovascular unit. We analyzed the effect of 1,25(OH)2D3 on brain endothelial cell barrier integrity and tight junction proteins after hypoxia/reoxygenation in a mouse brain endothelial cell culture model that closely mimics many of the features of the blood-brain barrier in vitro. Following hypoxic injury in bEnd.3 cells, 1,25(OH)2D3 treatment prevented the decrease in barrier function as measured by transendothelial electrical resistance and permeability of FITC-dextran (40 kDa), the decrease in the expression of the tight junction proteins zonula occludin-1, claudin-5, and occludin, the activation of NF-kB, and the increase in matrix metalloproteinase-9 expression. These responses were blocked when the interaction of 1,25(OH) )2D3 with the vitamin D receptor (VDR) was inhibited by pyridoxal 5'-phosphate treatment. Our findings show a direct, VDR-mediated, protective effect of 1,25(OH) )2D3 against ischemic injury-induced blood-brain barrier dysfunction in cerebral endothelial cells.

No MeSH data available.


Related in: MedlinePlus

Effects of 1,25(OH)2D3 on hypoxia/reoxygenation-induced mitochondrial superoxide production and hydrogen peroxide in bEnd.3 cells.(A) Mitochondrial superoxide production in live cells was measured by fluorescence microscopy using MitoSOX Red dye. Representative fluorescence images show localization of MitoSOX Red fluorescence and DAPI fluorescence. Scale bar 50 μm. (B) MitoSOX Red fluorescence per cell was quantified using ImageJ software. Image data from 51–60 cells per treatment condition were averaged (n = 3). (C) Extracellular H2O2 production with the use of Amplex Red (n = 4). Data are expressed as mean ± s.e.m; *p < 0.05, **p < 0.01, and ***p < 0.001.
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pone.0122821.g003: Effects of 1,25(OH)2D3 on hypoxia/reoxygenation-induced mitochondrial superoxide production and hydrogen peroxide in bEnd.3 cells.(A) Mitochondrial superoxide production in live cells was measured by fluorescence microscopy using MitoSOX Red dye. Representative fluorescence images show localization of MitoSOX Red fluorescence and DAPI fluorescence. Scale bar 50 μm. (B) MitoSOX Red fluorescence per cell was quantified using ImageJ software. Image data from 51–60 cells per treatment condition were averaged (n = 3). (C) Extracellular H2O2 production with the use of Amplex Red (n = 4). Data are expressed as mean ± s.e.m; *p < 0.05, **p < 0.01, and ***p < 0.001.

Mentions: To determine whether the protective effect of 1,25(OH)2D3 on BBB damage occurs through inhibition of ROS generation after H/R, we measured mitochondrial superoxide level and intracellular hydrogen peroxide after H/R using MitoSOX Red fluorescence. As shown in Fig. 3, H/R caused a marked increase in MitoSOX Red fluorescence in vehicle-treated cells compared with normoxia. However, this effect was substantially attenuated (58%) in cells pretreated with 100 nmol/L 1,25(OH)2D3. In contrast, when cells were treated with P5P in addition to 1,25(OH)2D3, the protective effect of 1,25(OH)2D3 was substantially decreased.


Vitamin D prevents hypoxia/reoxygenation-induced blood-brain barrier disruption via vitamin D receptor-mediated NF-kB signaling pathways.

Won S, Sayeed I, Peterson BL, Wali B, Kahn JS, Stein DG - PLoS ONE (2015)

Effects of 1,25(OH)2D3 on hypoxia/reoxygenation-induced mitochondrial superoxide production and hydrogen peroxide in bEnd.3 cells.(A) Mitochondrial superoxide production in live cells was measured by fluorescence microscopy using MitoSOX Red dye. Representative fluorescence images show localization of MitoSOX Red fluorescence and DAPI fluorescence. Scale bar 50 μm. (B) MitoSOX Red fluorescence per cell was quantified using ImageJ software. Image data from 51–60 cells per treatment condition were averaged (n = 3). (C) Extracellular H2O2 production with the use of Amplex Red (n = 4). Data are expressed as mean ± s.e.m; *p < 0.05, **p < 0.01, and ***p < 0.001.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4376709&req=5

pone.0122821.g003: Effects of 1,25(OH)2D3 on hypoxia/reoxygenation-induced mitochondrial superoxide production and hydrogen peroxide in bEnd.3 cells.(A) Mitochondrial superoxide production in live cells was measured by fluorescence microscopy using MitoSOX Red dye. Representative fluorescence images show localization of MitoSOX Red fluorescence and DAPI fluorescence. Scale bar 50 μm. (B) MitoSOX Red fluorescence per cell was quantified using ImageJ software. Image data from 51–60 cells per treatment condition were averaged (n = 3). (C) Extracellular H2O2 production with the use of Amplex Red (n = 4). Data are expressed as mean ± s.e.m; *p < 0.05, **p < 0.01, and ***p < 0.001.
Mentions: To determine whether the protective effect of 1,25(OH)2D3 on BBB damage occurs through inhibition of ROS generation after H/R, we measured mitochondrial superoxide level and intracellular hydrogen peroxide after H/R using MitoSOX Red fluorescence. As shown in Fig. 3, H/R caused a marked increase in MitoSOX Red fluorescence in vehicle-treated cells compared with normoxia. However, this effect was substantially attenuated (58%) in cells pretreated with 100 nmol/L 1,25(OH)2D3. In contrast, when cells were treated with P5P in addition to 1,25(OH)2D3, the protective effect of 1,25(OH)2D3 was substantially decreased.

Bottom Line: We analyzed the effect of 1,25(OH)2D3 on brain endothelial cell barrier integrity and tight junction proteins after hypoxia/reoxygenation in a mouse brain endothelial cell culture model that closely mimics many of the features of the blood-brain barrier in vitro.These responses were blocked when the interaction of 1,25(OH) )2D3 with the vitamin D receptor (VDR) was inhibited by pyridoxal 5'-phosphate treatment.Our findings show a direct, VDR-mediated, protective effect of 1,25(OH) )2D3 against ischemic injury-induced blood-brain barrier dysfunction in cerebral endothelial cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Emergency Medicine Brain Research Laboratory, Emory University, Atlanta, Georgia, United States of America.

ABSTRACT
Maintaining blood-brain barrier integrity and minimizing neuronal injury are critical components of any therapeutic intervention following ischemic stroke. However, a low level of vitamin D hormone is a risk factor for many vascular diseases including stroke. The neuroprotective effects of 1,25(OH)2D3 (vitamin D) after ischemic stroke have been studied, but it is not known whether it prevents ischemic injury to brain endothelial cells, a key component of the neurovascular unit. We analyzed the effect of 1,25(OH)2D3 on brain endothelial cell barrier integrity and tight junction proteins after hypoxia/reoxygenation in a mouse brain endothelial cell culture model that closely mimics many of the features of the blood-brain barrier in vitro. Following hypoxic injury in bEnd.3 cells, 1,25(OH)2D3 treatment prevented the decrease in barrier function as measured by transendothelial electrical resistance and permeability of FITC-dextran (40 kDa), the decrease in the expression of the tight junction proteins zonula occludin-1, claudin-5, and occludin, the activation of NF-kB, and the increase in matrix metalloproteinase-9 expression. These responses were blocked when the interaction of 1,25(OH) )2D3 with the vitamin D receptor (VDR) was inhibited by pyridoxal 5'-phosphate treatment. Our findings show a direct, VDR-mediated, protective effect of 1,25(OH) )2D3 against ischemic injury-induced blood-brain barrier dysfunction in cerebral endothelial cells.

No MeSH data available.


Related in: MedlinePlus