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Brain endothelial cells synthesize neurotoxic thrombin in Alzheimer's disease.

ABSTRACT: Alzheimer's disease (AD) is characterized by neuronal death; thus, identifying neurotoxic proteins and their source is central to understanding and treating AD. The multifunctional protease thrombin is neurotoxic and found in AD senile plaques. The objective of this study was to determine whether brain endothelial cells can synthesize thrombin and thus be a source of this neurotoxin in AD brains. Microvessels were isolated from AD patient brains and from age-matched controls. Reverse transcription-PCR demonstrated that thrombin message was highly expressed in microvessels from AD brains but was not detectable in control vessels. Similarly, Western blot analysis of microvessels showed that the thrombin protein was highly expressed in AD- but not control-derived microvessels. In addition, high levels of thrombin were detected in cerebrospinal fluid obtained from AD but not control patients, and sections from AD brains showed reactivity to thrombin antibody in blood vessel walls but not in vessels from controls. Finally, we examined the ability of brain endothelial cells in culture to synthesize thrombin and showed that oxidative stress or cell signaling perturbations led to increased expression of thrombin mRNA in these cells. The results demonstrate, for the first time, that brain endothelial cells can synthesize thrombin, and suggest that novel therapeutics targeting vascular stabilization that prevent or decrease release of thrombin could prove useful in treating this neurodegenerative disease.

SUBMITTER: Yin X

PROVIDER: S-EPMC2843451 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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Brain endothelial cells synthesize neurotoxic thrombin in Alzheimer's disease.

Yin Xiangling X Wright Jill J Wall Trevor T Grammas Paula P

The American journal of pathology 20100211 4

Alzheimer's disease (AD) is characterized by neuronal death; thus, identifying neurotoxic proteins and their source is central to understanding and treating AD. The multifunctional protease thrombin is neurotoxic and found in AD senile plaques. The objective of this study was to determine whether brain endothelial cells can synthesize thrombin and thus be a source of this neurotoxin in AD brains. Microvessels were isolated from AD patient brains and from age-matched controls. Reverse transcripti ...[more]

PMID: 20150433

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Project description:Silence information regulator 3 (SIRT3) is an NAD+ dependent deacetylase enzyme that enhances the function of key mitochondrial proteins. We have earlier demonstrated that deletion of Sirt3 gene leads to downregulation of metabolic enzymes, mitochondrial dysfunction and neuroinflammation in the brain, the major causes of Alzheimer's disease (AD). We also reported recently that Sirt3 gene deletion in Alzheimer's transgenic mice leads to exacerbation of neuroinflammation, amyloid plaque deposition and microglial activation. AD often coexists with other brain lesions caused by comorbidities which can exert their deleterious effects through the neurovascular unit. This unit consists of brain microvascular endothelial cells (BMECs), end feet of astrocytes, and pericytes. BMECs are uniquely different from other vascular endothelial cells because they are glued together by tight-junction proteins. BMECs are in constant contact with circulating factors as they line the luminal side. Therefore, we hypothesized that vascular endothelial injury caused by comorbidities plays a significant role in neuroinflammation. Herein, we investigated the effects of lipotoxicity in BMECs and how Sirt3 deficiency facilitate the deleterious effects of lipotoxicity on them using in vivo and in vitro models. We observed decreases in the levels of SIRT3 and tight junction proteins in the brain samples of western diet-fed APP/PS1 mice. Similar observations were obtained with Alzheimer's post-mortem samples. Exposure of BEND3 cells, mouse brain-derived Endothelial cells3, to a combination of high glucose and palmitic acid resulted in significant (P < 0.01-P < 0.001) decreases in the levels of SIRT3, claudin-5 and ZO-1. Induction of inflammatory mediators, including Cox-2, CXCL1, RANTES, and GADD45β was also observed in these treated cells. Interestingly, the induction was more with Sirt3-silenced BEND3 cells, suggesting that Sirt3 deficiency exacerbates inflammatory response. Palmitic acid was more potent in inducing the inflammatory mediators. Significant cytotoxicity and changes in microglial morphology were observed when cocultures of Sirt3-silenced BEND3 and Sirt3-silenced BV2 cells were exposed to palmitic acid. Transendothelial electrical resistance measurement with these cocultures suggested decreased barrier integrity. The findings of this study suggest that hyperlipidemia in comorbidities can compromise blood brain barrier integrity by inducing inflammatory mediators and decreasing tight junction proteins in the vascular endothelial cells of the AD brain, leading to activation of microglia.

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Brain endothelial cells synthesize neurotoxic thrombin in Alzheimer's disease.

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Brain endothelial cells synthesize neurotoxic thrombin in Alzheimer's disease.

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