Summary Background Accumulating clinical evidence suggests that hyperhomocysteinemia (HHC) is correlated with Alzheimer’s disease (AD) and vascular dementia. Objective This study was carried out to elucidate the specific role of elevated homocysteine (HC) levels in AD pathophysiology. Methods Immunohistochemistry was used to examine amyloid-beta (Aβ) deposition along blood vessels, also known as cerebral amyloid angiopathy (CAA), fibrin(ogen) deposition, and their correlation to each other in the brains of AD patients with and without HHC. To study AD-HHC comorbidity in detail, an AD mouse model was administered a high methionine diet for several months. Parenchymal Aβ plaques, CAA-positive vessels, and fibrin deposits were then assessed by immunohistochemistry at different stages of AD progression. Memory deficits were evaluated with contextual fear conditioning and the Barnes maze. Additionally, the effect of HC and its metabolite, homocysteine thiolactone (HCTL), on the Aβ-fibrinogen interaction was analyzed by pull-down, ELISA, and fibrin clot formation and fibrinolysis assays in vitro. Results We found increased fibrin(ogen) levels and Aβ deposits in the blood vessels and brain parenchyma of AD patients with HHC. We demonstrate that HC and HCTL enhance the interaction between fibrinogen and Aβ, promote the formation of tighter fibrin clots, and delay clot fibrinolysis. Additionally, we show that diet-induced HHC in an AD mouse model leads to severe CAA and parenchymal Aβ deposition, as well as significant impairments in learning and memory. Conclusions These findings suggest that elevated levels of plasma HC/HCTL contribute to AD pathology via the Aβ-fibrin(ogen) interaction.

The cross‐talk between blood proteins, immune cells, and brain function involves complex mechanisms. Plasma protein C1 inhibitor (C1INH) is an inhibitor of vascular inflammation that is induced by activation of the kallikrein‐kinin system (KKS) and the complement system. Knockout of C1INH was previously correlated with peripheral vascular permeability via the bradykinin pathway, yet there was no evidence of its correlation with blood–brain barrier (BBB) integrity and brain function. In order to understand the effect of plasma C1INH on brain pathology via the vascular system, we knocked down circulating C1INH in wild‐type (WT) mice using an antisense oligonucleotide (ASO), without affecting C1INH expression in peripheral immune cells or the brain, and examined brain pathology. Long‐term elimination of endogenous C1INH in the plasma induced the activation of the KKS and peritoneal macrophages but did not activate the complement system. Bradykinin pathway proteins were elevated in the periphery and the brain, resulting in hypotension. BBB permeability, extravasation of plasma proteins into the brain parenchyma, activation of glial cells, and elevation of pro‐inflammatory response mediators were detected. Furthermore, infiltrating innate immune cells were observed entering the brain through the lateral ventricle walls and the neurovascular unit. Mice showed normal locomotion function, yet cognition was impaired and depressive‐like behavior was evident. In conclusion, our results highlight the important role of regulated plasma C1INH as it acts as a gatekeeper to the brain via the neurovascular system. Thus, manipulation of C1INH in neurovascular disorders might be therapeutically beneficial.

Activation of the innate immune system, mediated by the complement and the contact systems, induces inflammation. The inhibition of these pathways by C1 inhibitor (C1INH) has been shown to decrease pro-inflammatory response and mediate vascular permeability in the periphery. However, the role of peripheral C1INH has not been associated with brain function. Using an antisense oligonucleotide (ASO) to deplete circulating liver-derived C1INH in wildtype (WT) mice, we induced the activation of the kallikrein-kinin system (KKS), which produced bradykinin in the plasma, resulting in hypotension. Interestingly, the complement system was quiescent. Depletion of liver-derived C1INH increased blood-brain barrier (BBB) permeability, increased expression of bradykinin 2 receptor, activated resident glial cells to secrete pro-inflammatory mediators such as Il-1beta, Il-6, TNF-alpha and iNOS, and induced cognitive decline. The results of this study emphasize the important role of circulating C1INH in mediating brain function through the activation of the KKS. Thus, manipulation of C1INH in neuro-vascular disorders might be therapeutically beneficial.