Accumulation of high molecular weight kininogen in the brains of Alzheimer's disease patients may affect microglial function by altering phagocytosis and lysosomal cathepsin activity.

Increased activation of the contact system protein high molecular weight kininogen (HK) has been shown in plasma and cerebrospinal fluid of Alzheimer's disease (AD) patients, but its potential role in the brain has not been explored. We assessed HK levels in brain tissue from 20 AD patients and controls and modeled the effects of HK on microglia-like cells in culture. We show increased levels of HK in the hippocampus of AD patients, which colocalized with amyloid beta (Aß) deposits and activated microglia. Treatment of microglia with HK led to cell clustering and elevated levels of phagocytosed Aß. We demonstrate that microglia internalize HK and traffic it to lysosomes, which is accompanied by reduced activity of lysosomal cathepsins L and S. Our results suggest that HK accumulation in the AD hippocampus may alter microglial uptake and degradation of Aß fibrils, possibly contributing to microglial dysfunction in AD.

A Short Isoform of Coagulation Factor XII mRNA Is Expressed by Neurons in the Human Brain.

Coagulation factor XII (FXII) is synthesized in the liver and secreted into the circulation, where it initiates the contact activation system. Although typically thought to be restricted to the circulation, FXII protein has been found in the brain of Alzheimer's disease (AD) and multiple sclerosis patients. Moreover, activation of the contact system has been detected in the cerebrospinal fluid of these patients as well as in the brain of healthy and AD individuals. While FXII protein has been detected in the brain, its source and its potential role in brain physiology and/or pathology have not been elucidated. Using in situ hybridization, we show that a shorter FXII mRNA isoform is expressed by neurons in human brain and in the brain of FXII humanized mice, with the highest expression observed in pyramidal neurons. This shorter FXII transcript contains an open reading frame coding for the portion of FXII that spans its proline-rich and catalytic domains (FXII297-596). We show that a recombinant version of this shorter FXII protein is activated by plasma kallikrein, reciprocally activates prekallikrein, and converts pro-hepatocyte growth factor (HGF) to active HGF in vitro. HGF-Met signaling plays a role in neuronal development and survival, and its dysregulation has been implicated in neurodevelopmental disorders and neurodegeneration. Taken together, our results show that a short isoform of FXII mRNA is expressed in the brain and raise the possibility that brain-derived FXII may be involved in HGF-Met signaling in neurons.

A novel detection method of cleaved plasma high-molecular-weight kininogen reveals its correlation with Alzheimer's pathology and cognitive impairment.

INTRODUCTION: Accumulation of ß-amyloid is a pathological hallmark of Alzheimer's disease (AD). ß-Amyloid activates the plasma contact system leading to kallikrein-mediated cleavage of intact high-molecular-weight kininogen (HKi) to cleaved high-molecular-weight kininogen (HKc). Increased HKi cleavage is observed in plasma of AD patients and mouse models by Western blot. For potential diagnostic purposes, a more quantitative method that can measure HKc levels in plasma with high sensitivity and specificity is needed. METHODS: HKi/c, HKi, and HKc monoclonal antibodies were screened from hybridomas using direct ELISA with a fluorescent substrate. RESULTS: We generated monoclonal antibodies recognizing HKi or HKc specifically and developed sandwich ELISAs that can quantitatively detect HKi and HKc levels in human. These new assays show that decreased HKi and increased HKc levels in AD plasma correlate with dementia and neuritic plaque scores. DISCUSSION: High levels of plasma HKc could be used as an innovative biomarker for AD.

Interactions of ß-amyloid peptide with fibrinogen and coagulation factor XII may contribute to Alzheimer's disease.

PURPOSE OF REVIEW: To review the evidence that the Alzheimer peptide ß-amyloid interacts with the blood coagulation system and influences the pathophysiology of the disease. RECENT FINDINGS: ß-amyloid can interact with fibrinogen and blood coagulation factor XII and trigger ischemia and inflammation. SUMMARY: ß-amyloid interacts with fibrinogen and factor XII. These interactions can lead to increased clotting, abnormal clot formation, persistent fibrin deposition, and generation of proinflammatory molecules. These events can damage neurons and could contribute to the cognitive decline in Alzheimer's disease patients.

Biochemical and structural analysis of the interaction between ß-amyloid and fibrinogen.

The majority of patients with Alzheimer disease (AD) suffer from impaired cerebral circulation. Accumulating evidence suggests that fibrinogen, the main protein component of blood clots, plays an important role in this circulatory dysfunction in AD. Fibrinogen interacts with ß-amyloid (Aß), forming plasmin-resistant abnormal blood clots, and increased fibrin deposition is found in the brains of AD patients and mouse models. In this study, we investigated the biochemical and structural details of the Aß-fibrinogen interaction. We identified the central region of Aß42 as the most critical region for the interaction, which can be inhibited by specific antibodies against the central region of Aß and by naturally occurring p3 peptides, Aß17-40 and Aß17-42. X-ray crystallographic analysis revealed that Aß42 binding to fragment D of fibrinogen induced a structural change in the C-terminal region of the fibrinogen ß-chain (ß384-393). Furthermore, we identified an additional Aß-binding site within the αC region of fibrinogen. Aß binding to this αC region blocked plasmin-mediated fibrin cleavage at this site, resulting in the generation of increased levels of a plasmin-resistant fibrin degradation fragment. Overall, our study elucidates the Aß-fibrinogen interaction and clarifies the mechanism by which Aß-fibrinogen binding delays fibrinolysis by plasmin. These results may facilitate the development of effective therapeutics against the Aß-fibrinogen interaction to treat cerebrovascular abnormalities in AD.

A possible new role for Aß in vascular and inflammatory dysfunction in Alzheimer's disease.

Alzheimer's disease (AD) is often characterized by vascular pathology, a procoagulant state, and chronic inflammation. The mechanisms behind these abnormalities in AD are not clear. Here, we review evidence for the role of the AD-associated peptide Aß in promoting inflammation and thrombosis in AD via its interaction with the circulating proteins factor XII and fibrinogen.

Activation of the factor XII-driven contact system in Alzheimer's disease patient and mouse model plasma.

Alzheimer's disease (AD) is characterized by accumulation of the ß-amyloid peptide (Aß), which likely contributes to disease via multiple mechanisms. Increasing evidence implicates inflammation in AD, the origins of which are not completely understood. We investigated whether circulating Aß could initiate inflammation in AD via the plasma contact activation system. This proteolytic cascade is triggered by the activation of the plasma protein factor XII (FXII) and leads to kallikrein-mediated cleavage of high molecular-weight kininogen (HK) and release of proinflammatory bradykinin. Aß has been shown to promote FXII-dependent cleavage of HK in vitro. In addition, increased cleavage of HK has been found in the cerebrospinal fluid of patients with AD. Here, we show increased activation of FXII, kallikrein activity, and HK cleavage in AD patient plasma. Increased contact system activation is also observed in AD mouse model plasma and in plasma from wild-type mice i.v. injected with Aß42. Our results demonstrate that Aß42-mediated contact system activation can occur in the AD circulation and suggest new pathogenic mechanisms, diagnostic tests, and therapies for AD.

A novel Aß-fibrinogen interaction inhibitor rescues altered thrombosis and cognitive decline in Alzheimer's disease mice.

Many Alzheimer's disease (AD) patients suffer from cerebrovascular abnormalities such as altered cerebral blood flow and cerebral microinfarcts. Recently, fibrinogen has been identified as a strong cerebrovascular risk factor in AD, as it specifically binds to ß-amyloid (Aß), thereby altering fibrin clot structure and delaying clot degradation. To determine if the Aß-fibrinogen interaction could be targeted as a potential new treatment for AD, we designed a high-throughput screen and identified RU-505 as an effective inhibitor of the Aß-fibrinogen interaction. RU-505 restored Aß-induced altered fibrin clot formation and degradation in vitro and inhibited vessel occlusion in AD transgenic mice. Furthermore, long-term treatment of RU-505 significantly reduced vascular amyloid deposition and microgliosis in the cortex and improved cognitive impairment in mouse models of AD. Our studies suggest that inhibitors targeting the Aß-fibrinogen interaction show promise as therapy for treating AD.

Fibrinogen and altered hemostasis in Alzheimer's disease.

Alzheimer's disease (AD) is characterized by amyloid-ß (Aß) plaques, tau tangles, brain atrophy, and vascular pathology. Vascular defects include cerebrovascular dysfunction, decreased cerebral blood flow, and blood brain barrier (BBB) disruption, among others. Here, we review the evidence that links Aß with the vascular pathology present in AD, with a specific focus on the hemostatic system and the clotting protein fibrinogen. Fibrinogen is normally found circulating in blood, but in AD it deposits with Aß in the brain parenchyma and cerebral blood vessels. We found that Aß and fibrin(ogen) interact, and their binding leads to increased fibrinogen aggregation, Aß fibrillization, and the formation of degradation-resistant fibrin clots. Decreasing fibrinogen levels not only lessens cerebral amyloid angiopathy and BBB permeability, but it also reduces microglial activation and improves cognitive performance in AD mouse models. Moreover, a prothrombotic state in AD is evidenced by increased clot formation, decreased fibrinolysis, and elevated levels of coagulation factors and activated platelets. Abnormal deposition and persistence of fibrin(ogen) in AD may result from Aß-fibrin(ogen) binding and altered hemostasis and could thus contribute to Aß deposition, decreased cerebral blood flow, exacerbated neuroinflammation, and eventual neurodegeneration. Blocking the interaction between fibrin(ogen) and Aß may be a promising therapeutic target for AD.

Aß delays fibrin clot lysis by altering fibrin structure and attenuating plasminogen binding to fibrin.

Alzheimer disease is characterized by the presence of increased levels of the ß-amyloid peptide (Aß) in the brain parenchyma and cerebral blood vessels. This accumulated Aß can bind to fibrin(ogen) and render fibrin clots more resistant to degradation. Here, we demonstrate that Aß(42) specifically binds to fibrin and induces a tighter fibrin network characterized by thinner fibers and increased resistance to lysis. However, Aß(42)-induced structural changes cannot be the sole mechanism of delayed lysis because Aß overlaid on normal preformed clots also binds to fibrin and delays lysis without altering clot structure. In this regard, we show that Aß interferes with the binding of plasminogen to fibrin, which could impair plasmin generation and fibrin degradation. Indeed, plasmin generation by tissue plasminogen activator (tPA), but not streptokinase, is slowed in fibrin clots containing Aß(42), and clot lysis by plasmin, but not trypsin, is delayed. Notably, plasmin and tPA activities, as well as tPA-dependent generation of plasmin in solution, are not decreased in the presence of Aß(42). Our results indicate the existence of 2 mechanisms of Aß(42) involvement in delayed fibrinolysis: (1) through the induction of a tighter fibrin network composed of thinner fibers, and (2) through inhibition of plasmin(ogen)-fibrin binding.

Alzheimer's disease peptide beta-amyloid interacts with fibrinogen and induces its oligomerization.

Increasing evidence supports a vascular contribution to Alzheimer's disease (AD), but a direct connection between AD and the circulatory system has not been established. Previous work has shown that blood clots formed in the presence of the ß-amyloid peptide (Aß), which has been implicated in AD, have an abnormal structure and are resistant to degradation in vitro and in vivo. In the present study, we show that Aß specifically interacts with fibrinogen with a K(d) of 26.3 ± 6.7 nM, that the binding site is located near the C terminus of the fibrinogen ß-chain, and that the binding causes fibrinogen to oligomerize. These results suggest that the interaction between Aß and fibrinogen modifies fibrinogen's structure, which may then lead to abnormal fibrin clot formation. Overall, our study indicates that the interaction between Aß and fibrinogen may be an important contributor to the vascular abnormalities found in AD.

Fibrinogen and beta-amyloid association alters thrombosis and fibrinolysis: a possible contributing factor to Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative disorder in which vascular pathology plays an important role. Since the beta-amyloid peptide (Abeta) is a critical factor in this disease, we examined its relationship to fibrin clot formation in AD. In vitro and in vivo experiments showed that fibrin clots formed in the presence of Abeta are structurally abnormal and resistant to degradation. Fibrin(ogen) was observed in blood vessels positive for amyloid in mouse and human AD samples, and intravital brain imaging of clot formation and dissolution revealed abnormal thrombosis and fibrinolysis in AD mice. Moreover, depletion of fibrinogen lessened cerebral amyloid angiopathy pathology and reduced cognitive impairment in AD mice. These experiments suggest that one important contribution of Abeta to AD is via its effects on fibrin clots, implicating fibrin(ogen) as a potential critical factor in this disease.