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In Vitro Assays to Assess Blood-brain Barrier Mesh-like Vessel Formation and Disruption
Published on: June 20, 2017
Homocysteine potentiates amyloid β -induced death receptor 4- and 5-mediated cerebral endothelial cell apoptosis,
Ashley Carey1, Rebecca Parodi-Rullan1, Rafael Vazquez-Torres1
1Department of Neural Sciences, Alzheimer's Center at Temple, Temple University Lewis Katz School of Medicine, Philadelphia, Pennsylvania, USA.
Insights
Cardiovascular risk factor hyperhomocysteinemia (Hhcy) worsens Alzheimer's-related brain cell damage and blood-brain barrier issues by amplifying amyloid-beta effects on cell death and blood flow. This reveals shared molecular pathways for combined therapeutic targets.
Area of Science:
- Neuroscience
- Vascular Biology
- Molecular Biology
Background:
- Cerebrovascular dysfunction is a key factor in Alzheimer's Disease (AD) pathology.
- Cardiovascular risk factors like hyperhomocysteinemia (Hhcy) may worsen AD.
- Amyloid-beta (Aβ) species, particularly AβQ22, induce brain endothelial cell (cEC) apoptosis and barrier dysfunction.
Purpose of the Study:
- To investigate if Hhcy exacerbates Aβ-induced cEC apoptosis, barrier dysfunction, and angiogenesis defects.
- To elucidate the molecular mechanisms underlying the interaction between Hhcy and Aβ in cECs.
- To identify potential therapeutic targets for comorbid AD and Hhcy conditions.
Main Methods:
- Utilized human cECs to study the effects of Hhcy and AβQ22.
- Assessed apoptosis via DR4/DR5 pathway activation, caspase activity, and DNA fragmentation.
- Evaluated blood-brain barrier (BBB) integrity by examining cEC junction proteins.
- Measured angiogenic capacity by analyzing VEGF-A/VEGFR2 signaling and VEGFR2 trafficking.
Main Results:
- Hhcy potentiates AβQ22-induced cEC apoptosis through the extrinsic death receptor pathway.
- Hhcy intensifies Aβ-mediated deregulation of cEC junction proteins, increasing BBB permeability.
- Hhcy and AβQ22 additively impair cEC angiogenic capabilities by disrupting VEGF-A/VEGFR2 signaling.
Conclusions:
- Hhcy significantly exacerbates Aβ-induced cEC apoptosis, barrier dysfunction, and angiogenic impairment.
- This study reveals molecular mechanisms by which amyloidosis and Hhcy jointly damage brain endothelial cells.
- Findings highlight potential molecular targets for treating vascular pathology in comorbid AD/CAA and Hhcy.
Abstract:
Cerebrovascular dysfunction has been implicated as a major contributor to Alzheimer's Disease (AD) pathology, with cerebral endothelial cell (cEC) stress promoting ischemia, cerebral-blood flow impairments and blood-brain barrier (BBB) permeability. Recent evidence suggests that cardiovascular (CV)/cerebrovascular risk factors, including hyperhomocysteinemia (Hhcy), exacerbate AD pathology and risk. Yet, the underlying molecular mechanisms for this interaction remain unclear. Our lab has demonstrated that amyloid beta 40 (Aβ40) species, and particularly Aβ40-E22Q (AβQ22; vasculotropic Dutch mutant), promote death receptor 4 and 5 (DR4/DR5)-mediated apoptosis in human cECs, barrier permeability, and angiogenic impairment. Previous studies show that Hhcy also induces EC dysfunction, but it remains unknown whether Aβ and homocysteine function through common molecular mechanisms. We tested the hypotheses that Hhcy exacerbates Aβ-induced cEC DR4/5-mediated apoptosis, barrier dysfunction, and angiogenesis defects. This study was the first to demonstrate that Hhcy specifically potentiates AβQ22-mediated activation of the DR4/5-mediated extrinsic apoptotic pathway in cECs, including DR4/5 expression, caspase 8/9/3 activation, cytochrome-c release and DNA fragmentation. Additionally, we revealed that Hhcy intensifies the deregulation of the same cEC junction proteins mediated by Aβ, precipitating BBB permeability. Furthermore, Hhcy and AβQ22, impairing VEGF-A/VEGFR2 signaling and VEGFR2 endosomal trafficking, additively decrease cEC angiogenic capabilities. Overall, these results show that the presence of the CV risk factor Hhcy exacerbates Aβ-induced cEC apoptosis, barrier dysfunction, and angiogenic impairment. This study reveals specific mechanisms through which amyloidosis and Hhcy jointly operate to produce brain EC dysfunction and death, highlighting new potential molecular targets against vascular pathology in comorbid AD/CAA and Hhcy conditions.
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