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Ruscogenin Attenuates Cerebral Ischemia-Induced Blood-Brain Barrier Dysfunction by Suppressing TXNIP/NLRP3 Inflammasome Activation and the MAPK Pathway

View Article: PubMed Central - PubMed

ABSTRACT

Ruscogenin, an important steroid sapogenin derived from Ophiopogon japonicus, has been shown to inhibit cerebral ischemic injury. However, its potential molecular action on blood-brain barrier (BBB) dysfunction after stroke remains unclear. This study aimed to investigate the effects of ruscogenin on BBB dysfunction and the underlying mechanisms in middle cerebral artery occlusion/reperfusion (MCAO/R)-injured mice and oxygen–glucose deprivation/reoxygenation (OGD/R)-injured mouse brain microvascular endothelial cells (bEnd.3). The results demonstrated that administration of ruscogenin (10 mg/kg) decreased the brain infarction and edema, improved neurological deficits, increased cerebral brain flow (CBF), ameliorated histopathological damage, reduced evans blue (EB) leakage and upregulated the expression of tight junctions (TJs) in MCAO/R-injured mice. Meanwhile, ruscogenin (0.1–10 µM) treatment increased cell viability and trans-endothelial electrical resistance (TEER) value, decreased sodium fluorescein leakage, and modulated the TJs expression in OGD/R-induced bEnd.3 cells. Moreover, ruscogenin also inhibited the expression of interleukin-1β (IL-1β) and caspase-1, and markedly suppressed the expression of Nucleotide-binding domain (NOD)-like receptor family, pyrin domain containing 3 (NLRP3) and thiredoxin-interactive protein (TXNIP) in vivo and in vitro. Furthermore, ruscogenin decreased reactive oxygen species (ROS) generation and inhibited the mitogen-activated protein kinase (MAPK) pathway in OGD/R-induced bEnd.3 cells. Our findings provide some new insights into its potential application for the prevention and treatment of ischemic stroke.

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Effects of ruscogenin on the expression of tight junction proteins and actin cytoskeleton rearrangement in bEnd.3 cells subjected to OGD/R. (A,B) Representative Western blot bands and quantitative analysis of the expression levels of ZO-1 and occludin in different groups. The band intensities were measured using scanning densitometry. The data were normalized to GAPDH expression (n = 3). The data are expressed as means ± SD. ##p < 0.01 vs. Control, * p < 0.05, ** p < 0.01 vs. Model; (C) representative immunostaining of F-actin in different groups in bEnd.3 cells subjected to OGD/R; the white arrow indicates the stress fiber. Scale bars = 100 µm.
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ijms-17-01418-f006: Effects of ruscogenin on the expression of tight junction proteins and actin cytoskeleton rearrangement in bEnd.3 cells subjected to OGD/R. (A,B) Representative Western blot bands and quantitative analysis of the expression levels of ZO-1 and occludin in different groups. The band intensities were measured using scanning densitometry. The data were normalized to GAPDH expression (n = 3). The data are expressed as means ± SD. ##p < 0.01 vs. Control, * p < 0.05, ** p < 0.01 vs. Model; (C) representative immunostaining of F-actin in different groups in bEnd.3 cells subjected to OGD/R; the white arrow indicates the stress fiber. Scale bars = 100 µm.

Mentions: To further identify the relationship between tight junction’s expression and BBB disruption, we detected the expression of ZO-1 and occludin using Western blot analysis. The Western blot results demonstrated that the expressions of ZO-1 and occludin were decreased in bEnd.3 cells subjected to 6 h of OGD and 18 h reoxygenation, whereas pretreatment with ruscogenin could recover these expressions in response to OGD/R in bEnd.3 cells (Figure 6A,B). Meanwhile, we also found that pretreatment with ruscogenin could ameliorate the OGD/R-induced endothelial cell stress fiber (white arrow) formation (Figure 6C).


Ruscogenin Attenuates Cerebral Ischemia-Induced Blood-Brain Barrier Dysfunction by Suppressing TXNIP/NLRP3 Inflammasome Activation and the MAPK Pathway
Effects of ruscogenin on the expression of tight junction proteins and actin cytoskeleton rearrangement in bEnd.3 cells subjected to OGD/R. (A,B) Representative Western blot bands and quantitative analysis of the expression levels of ZO-1 and occludin in different groups. The band intensities were measured using scanning densitometry. The data were normalized to GAPDH expression (n = 3). The data are expressed as means ± SD. ##p < 0.01 vs. Control, * p < 0.05, ** p < 0.01 vs. Model; (C) representative immunostaining of F-actin in different groups in bEnd.3 cells subjected to OGD/R; the white arrow indicates the stress fiber. Scale bars = 100 µm.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC5037697&req=5

ijms-17-01418-f006: Effects of ruscogenin on the expression of tight junction proteins and actin cytoskeleton rearrangement in bEnd.3 cells subjected to OGD/R. (A,B) Representative Western blot bands and quantitative analysis of the expression levels of ZO-1 and occludin in different groups. The band intensities were measured using scanning densitometry. The data were normalized to GAPDH expression (n = 3). The data are expressed as means ± SD. ##p < 0.01 vs. Control, * p < 0.05, ** p < 0.01 vs. Model; (C) representative immunostaining of F-actin in different groups in bEnd.3 cells subjected to OGD/R; the white arrow indicates the stress fiber. Scale bars = 100 µm.
Mentions: To further identify the relationship between tight junction’s expression and BBB disruption, we detected the expression of ZO-1 and occludin using Western blot analysis. The Western blot results demonstrated that the expressions of ZO-1 and occludin were decreased in bEnd.3 cells subjected to 6 h of OGD and 18 h reoxygenation, whereas pretreatment with ruscogenin could recover these expressions in response to OGD/R in bEnd.3 cells (Figure 6A,B). Meanwhile, we also found that pretreatment with ruscogenin could ameliorate the OGD/R-induced endothelial cell stress fiber (white arrow) formation (Figure 6C).

View Article: PubMed Central - PubMed

ABSTRACT

Ruscogenin, an important steroid sapogenin derived from Ophiopogon japonicus, has been shown to inhibit cerebral ischemic injury. However, its potential molecular action on blood-brain barrier (BBB) dysfunction after stroke remains unclear. This study aimed to investigate the effects of ruscogenin on BBB dysfunction and the underlying mechanisms in middle cerebral artery occlusion/reperfusion (MCAO/R)-injured mice and oxygen&ndash;glucose deprivation/reoxygenation (OGD/R)-injured mouse brain microvascular endothelial cells (bEnd.3). The results demonstrated that administration of ruscogenin (10 mg/kg) decreased the brain infarction and edema, improved neurological deficits, increased cerebral brain flow (CBF), ameliorated histopathological damage, reduced evans blue (EB) leakage and upregulated the expression of tight junctions (TJs) in MCAO/R-injured mice. Meanwhile, ruscogenin (0.1&ndash;10 &micro;M) treatment increased cell viability and trans-endothelial electrical resistance (TEER) value, decreased sodium fluorescein leakage, and modulated the TJs expression in OGD/R-induced bEnd.3 cells. Moreover, ruscogenin also inhibited the expression of interleukin-1&beta; (IL-1&beta;) and caspase-1, and markedly suppressed the expression of Nucleotide-binding domain (NOD)-like receptor family, pyrin domain containing 3 (NLRP3) and thiredoxin-interactive protein (TXNIP) in vivo and in vitro. Furthermore, ruscogenin decreased reactive oxygen species (ROS) generation and inhibited the mitogen-activated protein kinase (MAPK) pathway in OGD/R-induced bEnd.3 cells. Our findings provide some new insights into its potential application for the prevention and treatment of ischemic stroke.

No MeSH data available.


Related in: MedlinePlus