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Endothelin-1 Mediates Brain Microvascular Dysfunction Leading to Long-Term Cognitive Impairment in a Model of Experimental Cerebral Malaria.

Freeman BD, Martins YC, Akide-Ndunge OB, Bruno FP, Wang H, Tanowitz HB, Spray DC, Desruisseaux MS - PLoS Pathog. (2016)

Bottom Line: Herein, we demonstrate that an endothelin type A receptor (ETA) antagonist prevented experimental cerebral malaria-induced neurocognitive impairments and improved survival.ETA antagonism prevented blood-brain barrier disruption and cerebral vasoconstriction during experimental cerebral malaria, and reduced brain endothelial activation, diminishing brain microvascular congestion.This vasoactive peptide may thus serve as a potential target for adjunctive therapy in the management of cerebral malaria.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Albert Einstein College of Medicine, Bronx, New York, United States of America.

ABSTRACT
Plasmodium falciparum infection causes a wide spectrum of diseases, including cerebral malaria, a potentially life-threatening encephalopathy. Vasculopathy is thought to contribute to cerebral malaria pathogenesis. The vasoactive compound endothelin-1, a key participant in many inflammatory processes, likely mediates vascular and cognitive dysfunctions in cerebral malaria. We previously demonstrated that C57BL6 mice infected with P. berghei ANKA, our fatal experimental cerebral malaria model, sustained memory loss. Herein, we demonstrate that an endothelin type A receptor (ETA) antagonist prevented experimental cerebral malaria-induced neurocognitive impairments and improved survival. ETA antagonism prevented blood-brain barrier disruption and cerebral vasoconstriction during experimental cerebral malaria, and reduced brain endothelial activation, diminishing brain microvascular congestion. Furthermore, exogenous endothelin-1 administration to P. berghei NK65-infected mice, a model generally regarded as a non-cerebral malaria negative control for P. berghei ANKA infection, led to experimental cerebral malaria-like memory deficits. Our data indicate that endothelin-1 is critical in the development of cerebrovascular and cognitive impairments with experimental cerebral malaria. This vasoactive peptide may thus serve as a potential target for adjunctive therapy in the management of cerebral malaria.

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ET-1 mediates cerebral endothelial activation, neuroinflammation and microvascular congestion during ECM.Vascular congestion was assessed by the degree of intravascular obstruction (A-F). PbA-infected mice showed a significant increase in cerebral microvascular obstruction compared to uninfected controls in the brainstem (A), cerebellum (B), hippocampus (E), and cortex (F). BQ123 treatment significantly reduced the degree of microvascular obstruction of PbA-infected mice in those regions. One-way ANOVA with post-hoc Tukey's comparison. (C,D; arrows) Representative histological image of cerebral blood vessel from PbA-infected and PbA-infected BQ123 treated mice. Endothelial activation was determined by VCAM-1 immunostaining (G,H; cerebellum shown). (K) Representative immunoblot of VCAM-1 protein expression. (L) PbA infection resulted in significantly higher VCAM-1 expression in the brain, and this was partially restored to uninfected levels by BQ123. One-way ANOVA with post-hoc Fisher's LSD comparison. (I,J) Expression of TNF and CCL2 were quantified by real-time PCR. (I) PbA infection resulted significantly higher expression of TNF, and this was prevented by treatment with BQ123. ANOVA with post-hoc Games-Howell comparison. (J) Likewise, PbA induced a significant increase in CCL2 expression which was prevented by BQ123. One-way ANOVA with Tukey's comparison. For graphs A, I, J, a logarithmic transformation was performed to yield variance homogeneity and/ or normal distribution. * = p < 0.05, ** = p < 0.01, *** = p < 0.001 and **** p < 0.0001 n = 5–8/group for vascular congestion, TNF and CCL2 analysis; n = 9–15/group for VCAM analysis.
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ppat.1005477.g006: ET-1 mediates cerebral endothelial activation, neuroinflammation and microvascular congestion during ECM.Vascular congestion was assessed by the degree of intravascular obstruction (A-F). PbA-infected mice showed a significant increase in cerebral microvascular obstruction compared to uninfected controls in the brainstem (A), cerebellum (B), hippocampus (E), and cortex (F). BQ123 treatment significantly reduced the degree of microvascular obstruction of PbA-infected mice in those regions. One-way ANOVA with post-hoc Tukey's comparison. (C,D; arrows) Representative histological image of cerebral blood vessel from PbA-infected and PbA-infected BQ123 treated mice. Endothelial activation was determined by VCAM-1 immunostaining (G,H; cerebellum shown). (K) Representative immunoblot of VCAM-1 protein expression. (L) PbA infection resulted in significantly higher VCAM-1 expression in the brain, and this was partially restored to uninfected levels by BQ123. One-way ANOVA with post-hoc Fisher's LSD comparison. (I,J) Expression of TNF and CCL2 were quantified by real-time PCR. (I) PbA infection resulted significantly higher expression of TNF, and this was prevented by treatment with BQ123. ANOVA with post-hoc Games-Howell comparison. (J) Likewise, PbA induced a significant increase in CCL2 expression which was prevented by BQ123. One-way ANOVA with Tukey's comparison. For graphs A, I, J, a logarithmic transformation was performed to yield variance homogeneity and/ or normal distribution. * = p < 0.05, ** = p < 0.01, *** = p < 0.001 and **** p < 0.0001 n = 5–8/group for vascular congestion, TNF and CCL2 analysis; n = 9–15/group for VCAM analysis.

Mentions: During CM, there is an increase in brain microvascular endothelial cell activation, critical for parasitized red blood cell (pRBC), platelet and leukocyte adhesion, resulting in sequestration, capillary obstruction, localized hypoxia, tissue injury and subsequent neurocognitive impairment or death [29–31]. Endothelial activation and microvascular obstruction are hallmark features of CM, and are associated with coma in patients with CM [24]. Using a semi-quantitative scoring system we assessed the degree of cellular retention within the cerebral vasculature, measured as percent obstruction of vessel lumen. Mice with ECM displayed a significant degree of vessel congestion, measured by the degree of vessel lumen obstruction (Fig 6C and 6D), that was not evident in uninfected mice in several brain regions, including the brainstem (Fig 6A; Log10 congestion score: 0.37 ± 0.03 vs. 0.27 ± 0.03; P < 0.05), cerebellum (Fig 6B; Score: 2.24 ± 0.15 vs. 1.79 ± 0.08; P < 0.05), hippocampus (Fig 6E; Score: 2.2 ± 0.1 vs. 1.55 ± 0.06; P < 0.0001), and cerebral cortex (Fig 6F; Score: 2.36 ± 0.13 vs. 1.73 ± 0.14; P < 0.01). Treatment with BQ123 prevented cerebral microvascular congestion in the brainstem (Fig 6A; Log10 Score: 0.23 ± 0.01 vs. 0.37 ± 0.03; P < 0.01), cerebellum (Fig 6B; Score: 1.405 ± .0.7 vs. 2.24 ± 0.15; P < 0.001), hippocampus (Fig 6E; Score: 1.24 ± 0.04 vs. 2.2 ± 0.11; P < 0.0001), and cerebral cortex (Fig 6F; Score: 1.46 ± 0.08 vs. 2.36 ± 0.13; P < 0.001) of PbA-infected mice.


Endothelin-1 Mediates Brain Microvascular Dysfunction Leading to Long-Term Cognitive Impairment in a Model of Experimental Cerebral Malaria.

Freeman BD, Martins YC, Akide-Ndunge OB, Bruno FP, Wang H, Tanowitz HB, Spray DC, Desruisseaux MS - PLoS Pathog. (2016)

ET-1 mediates cerebral endothelial activation, neuroinflammation and microvascular congestion during ECM.Vascular congestion was assessed by the degree of intravascular obstruction (A-F). PbA-infected mice showed a significant increase in cerebral microvascular obstruction compared to uninfected controls in the brainstem (A), cerebellum (B), hippocampus (E), and cortex (F). BQ123 treatment significantly reduced the degree of microvascular obstruction of PbA-infected mice in those regions. One-way ANOVA with post-hoc Tukey's comparison. (C,D; arrows) Representative histological image of cerebral blood vessel from PbA-infected and PbA-infected BQ123 treated mice. Endothelial activation was determined by VCAM-1 immunostaining (G,H; cerebellum shown). (K) Representative immunoblot of VCAM-1 protein expression. (L) PbA infection resulted in significantly higher VCAM-1 expression in the brain, and this was partially restored to uninfected levels by BQ123. One-way ANOVA with post-hoc Fisher's LSD comparison. (I,J) Expression of TNF and CCL2 were quantified by real-time PCR. (I) PbA infection resulted significantly higher expression of TNF, and this was prevented by treatment with BQ123. ANOVA with post-hoc Games-Howell comparison. (J) Likewise, PbA induced a significant increase in CCL2 expression which was prevented by BQ123. One-way ANOVA with Tukey's comparison. For graphs A, I, J, a logarithmic transformation was performed to yield variance homogeneity and/ or normal distribution. * = p < 0.05, ** = p < 0.01, *** = p < 0.001 and **** p < 0.0001 n = 5–8/group for vascular congestion, TNF and CCL2 analysis; n = 9–15/group for VCAM analysis.
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ppat.1005477.g006: ET-1 mediates cerebral endothelial activation, neuroinflammation and microvascular congestion during ECM.Vascular congestion was assessed by the degree of intravascular obstruction (A-F). PbA-infected mice showed a significant increase in cerebral microvascular obstruction compared to uninfected controls in the brainstem (A), cerebellum (B), hippocampus (E), and cortex (F). BQ123 treatment significantly reduced the degree of microvascular obstruction of PbA-infected mice in those regions. One-way ANOVA with post-hoc Tukey's comparison. (C,D; arrows) Representative histological image of cerebral blood vessel from PbA-infected and PbA-infected BQ123 treated mice. Endothelial activation was determined by VCAM-1 immunostaining (G,H; cerebellum shown). (K) Representative immunoblot of VCAM-1 protein expression. (L) PbA infection resulted in significantly higher VCAM-1 expression in the brain, and this was partially restored to uninfected levels by BQ123. One-way ANOVA with post-hoc Fisher's LSD comparison. (I,J) Expression of TNF and CCL2 were quantified by real-time PCR. (I) PbA infection resulted significantly higher expression of TNF, and this was prevented by treatment with BQ123. ANOVA with post-hoc Games-Howell comparison. (J) Likewise, PbA induced a significant increase in CCL2 expression which was prevented by BQ123. One-way ANOVA with Tukey's comparison. For graphs A, I, J, a logarithmic transformation was performed to yield variance homogeneity and/ or normal distribution. * = p < 0.05, ** = p < 0.01, *** = p < 0.001 and **** p < 0.0001 n = 5–8/group for vascular congestion, TNF and CCL2 analysis; n = 9–15/group for VCAM analysis.
Mentions: During CM, there is an increase in brain microvascular endothelial cell activation, critical for parasitized red blood cell (pRBC), platelet and leukocyte adhesion, resulting in sequestration, capillary obstruction, localized hypoxia, tissue injury and subsequent neurocognitive impairment or death [29–31]. Endothelial activation and microvascular obstruction are hallmark features of CM, and are associated with coma in patients with CM [24]. Using a semi-quantitative scoring system we assessed the degree of cellular retention within the cerebral vasculature, measured as percent obstruction of vessel lumen. Mice with ECM displayed a significant degree of vessel congestion, measured by the degree of vessel lumen obstruction (Fig 6C and 6D), that was not evident in uninfected mice in several brain regions, including the brainstem (Fig 6A; Log10 congestion score: 0.37 ± 0.03 vs. 0.27 ± 0.03; P < 0.05), cerebellum (Fig 6B; Score: 2.24 ± 0.15 vs. 1.79 ± 0.08; P < 0.05), hippocampus (Fig 6E; Score: 2.2 ± 0.1 vs. 1.55 ± 0.06; P < 0.0001), and cerebral cortex (Fig 6F; Score: 2.36 ± 0.13 vs. 1.73 ± 0.14; P < 0.01). Treatment with BQ123 prevented cerebral microvascular congestion in the brainstem (Fig 6A; Log10 Score: 0.23 ± 0.01 vs. 0.37 ± 0.03; P < 0.01), cerebellum (Fig 6B; Score: 1.405 ± .0.7 vs. 2.24 ± 0.15; P < 0.001), hippocampus (Fig 6E; Score: 1.24 ± 0.04 vs. 2.2 ± 0.11; P < 0.0001), and cerebral cortex (Fig 6F; Score: 1.46 ± 0.08 vs. 2.36 ± 0.13; P < 0.001) of PbA-infected mice.

Bottom Line: Herein, we demonstrate that an endothelin type A receptor (ETA) antagonist prevented experimental cerebral malaria-induced neurocognitive impairments and improved survival.ETA antagonism prevented blood-brain barrier disruption and cerebral vasoconstriction during experimental cerebral malaria, and reduced brain endothelial activation, diminishing brain microvascular congestion.This vasoactive peptide may thus serve as a potential target for adjunctive therapy in the management of cerebral malaria.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Albert Einstein College of Medicine, Bronx, New York, United States of America.

ABSTRACT
Plasmodium falciparum infection causes a wide spectrum of diseases, including cerebral malaria, a potentially life-threatening encephalopathy. Vasculopathy is thought to contribute to cerebral malaria pathogenesis. The vasoactive compound endothelin-1, a key participant in many inflammatory processes, likely mediates vascular and cognitive dysfunctions in cerebral malaria. We previously demonstrated that C57BL6 mice infected with P. berghei ANKA, our fatal experimental cerebral malaria model, sustained memory loss. Herein, we demonstrate that an endothelin type A receptor (ETA) antagonist prevented experimental cerebral malaria-induced neurocognitive impairments and improved survival. ETA antagonism prevented blood-brain barrier disruption and cerebral vasoconstriction during experimental cerebral malaria, and reduced brain endothelial activation, diminishing brain microvascular congestion. Furthermore, exogenous endothelin-1 administration to P. berghei NK65-infected mice, a model generally regarded as a non-cerebral malaria negative control for P. berghei ANKA infection, led to experimental cerebral malaria-like memory deficits. Our data indicate that endothelin-1 is critical in the development of cerebrovascular and cognitive impairments with experimental cerebral malaria. This vasoactive peptide may thus serve as a potential target for adjunctive therapy in the management of cerebral malaria.

Show MeSH
Related in: MedlinePlus