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Overlapping Protein Accumulation Profiles of CADASIL and CAA

Is There a Common Mechanism Driving Cerebral Small-Vessel Disease?
  • Kelly Z. Young
    Affiliations
    Department of Neurology, University of Michigan, Ann Arbor, Michigan

    Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan
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  • Gang Xu
    Affiliations
    Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan
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  • Simon G. Keep
    Affiliations
    Department of Neurology, University of Michigan, Ann Arbor, Michigan
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  • Jimo Borjigin
    Affiliations
    Department of Neurology, University of Michigan, Ann Arbor, Michigan

    Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan
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  • Michael M. Wang
    Correspondence
    Address correspondence to Michael M. Wang, M.D., Ph.D. 7725 Medical Science Bldg. II Box 5622, 1137 Catherine St., Ann Arbor, MI 48109-5622.
    Affiliations
    Department of Neurology, University of Michigan, Ann Arbor, Michigan

    Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan

    Department of Neurology Service, VA Ann Arbor Healthcare System, Ann Arbor, Michigan
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Open AccessPublished:December 29, 2020DOI:https://doi.org/10.1016/j.ajpath.2020.11.015
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) and cerebral amyloid angiopathy (CAA) are two distinct vascular angiopathies that share several similarities in clinical presentation and vascular pathology. Given the clinical and pathologic overlap, the molecular overlap between CADASIL and CAA was explored. CADASIL and CAA protein profiles from recently published proteomics-based and immuno-based studies were compared to investigate the potential for shared disease mechanisms. A comparison of affected proteins in each disease highlighted 19 proteins that are regulated in both CADASIL and CAA. Functional analysis of the shared proteins predicts significant interaction between them and suggests that most enriched proteins play roles in extracellular matrix structure and remodeling. Proposed models to explain the observed enrichment of extracellular matrix proteins include both increased protein secretion and decreased protein turnover by sequestration of chaperones and proteases or formation of stable protein complexes. Single-cell RNA sequencing of vascular cells in mice suggested that the vast majority of the genes accounting for the overlapped proteins between CADASIL and CAA are expressed by fibroblasts. Thus, our current understanding of the molecular profiles of CADASIL and CAA appears to support potential for common mechanisms underlying the two disorders.
      Cerebrovascular diseases affect a significant portion of our aging population and are major contributors to cognitive impairment and dementia.
      • Almkvist O.
      • Wahlund L.O.
      • Andersson-Lundman G.
      • Basun H.
      • Bäckman L.
      White-matter hyperintensity and neuropsychological functions in dementia and healthy aging.
      ,
      • Kuller L.H.
      • Longstreth W.T.
      • Arnold A.M.
      • Bernick C.
      • Bryan R.N.
      • Beauchamp N.J.
      Cardiovascular Health Study Collaborative Research Group
      White matter hyperintensity on cranial magnetic resonance imaging: a predictor of stroke.
      Improved understanding of the molecular basis of cerebrovascular diseases is imperative for the development of novel and effective clinical therapeutics. The purpose of this review is to compare two seemingly distinct vascular disease processes with small-vessel disease (SVD) burden to identify potentially shared properties and/or disease pathways. In particular, increasing evidence from two vascular angiopathies, cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) and cerebral amyloid angiopathy (CAA) is discussed, that supports molecular overlap in cerebrovascular disease mechanisms.
      To highlight the extent of overlap between these diseases, first the clinical overviews, genetics, histopathology, protein processing, protein aggregation, and protein accumulation involved in CADASIL and CAA were reviewed.

      Cerebral Autosomal Dominant Arteriopathy with Subcortical Infarcts and Leukoencephalopathy

      Clinical Overview of CADASIL

      Cerebral SVD is a prevalent vascular disorder of the brain and a major contributor to neurologic deterioration in our elderly population. Recent estimates report that sporadic SVD affects over half of the population aged >65 years and significantly elevates the risk of stroke, vascular dementia, and progression of neurologic diseases.
      • Almkvist O.
      • Wahlund L.O.
      • Andersson-Lundman G.
      • Basun H.
      • Bäckman L.
      White-matter hyperintensity and neuropsychological functions in dementia and healthy aging.
      ,
      • Kuller L.H.
      • Longstreth W.T.
      • Arnold A.M.
      • Bernick C.
      • Bryan R.N.
      • Beauchamp N.J.
      Cardiovascular Health Study Collaborative Research Group
      White matter hyperintensity on cranial magnetic resonance imaging: a predictor of stroke.
      In addition to sporadic SVD, monogenic causes of SVD have also been described, which may shed light on pathomechanisms. The most common type of SVD is CADASIL, caused primarily by stereotypical cysteine mutations in the NOTCH3 gene that result in altered cysteine number.
      • Joutel A.
      • Corpechot C.
      • Ducros A.
      • Vahedi K.
      • Chabriat H.
      • Mouton P.
      • Alamowitch S.
      • Domenga V.
      • Cécillion M.
      • Marechal E.
      • Maciazek J.
      • Vayssière C.
      • Cruaud C.
      • Cabanis E.A.
      • Ruchoux M.M.
      • Weissenbach J.
      • Bach J.F.
      • Bousser M.G.
      • Tournier-Lasserve E.
      Notch3 mutations in CADASIL, a hereditary adult-onset condition causing stroke and dementia.
      Clinically, patients with CADASIL present with an accelerated and often more severe clinical course compared with sporadic SVD. Although there can be variability in disease presentations, patients with CADASIL typically present with migraine with aura, subcortical ischemic events, mood disturbances, apathy, and cognitive impairment.
      • Dichgans M.
      • Mayer M.
      • Uttner I.
      • Brüning R.
      • Müller-Höcker J.
      • Rungger G.
      • Ebke M.
      • Klockgether T.
      • Gasser T.
      The phenotypic spectrum of CADASIL: clinical findings in 102 cases.
      In most patients, migraine with aura is the first observed symptom, often occurring roughly 15 years earlier than ischemia.
      • Dichgans M.
      • Mayer M.
      • Uttner I.
      • Brüning R.
      • Müller-Höcker J.
      • Rungger G.
      • Ebke M.
      • Klockgether T.
      • Gasser T.
      The phenotypic spectrum of CADASIL: clinical findings in 102 cases.

      Genetics of CADASIL

      CADASIL was first described in 1993 when several unrelated families presented with a mendelian syndrome that caused recurrent strokes.
      • Choi J.C.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy: a genetic cause of cerebral small vessel disease.
      Initially, these syndromes were reported under different names until the affected gene was mapped to chromosome 19q12.
      • Choi J.C.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy: a genetic cause of cerebral small vessel disease.
      Subsequent linkage analysis of additional families led to the mapping of the CADASIL critical region to the NOTCH3 gene.
      • Joutel A.
      • Corpechot C.
      • Ducros A.
      • Vahedi K.
      • Chabriat H.
      • Mouton P.
      • Alamowitch S.
      • Domenga V.
      • Cécillion M.
      • Marechal E.
      • Maciazek J.
      • Vayssière C.
      • Cruaud C.
      • Cabanis E.A.
      • Ruchoux M.M.
      • Weissenbach J.
      • Bach J.F.
      • Bousser M.G.
      • Tournier-Lasserve E.
      Notch3 mutations in CADASIL, a hereditary adult-onset condition causing stroke and dementia.
      Although the prevalence of genetically proven CADASIL is estimated to be roughly 2 per 100,000 adults, the actual prevalence is thought to be higher when taking into account sporadic mutations.
      • Choi J.C.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy: a genetic cause of cerebral small vessel disease.
      NOTCH3 gene encodes a vascular smooth muscle transmembrane protein involved in smooth muscle cell differentiation and vascular development.
      • Domenga V.
      • Fardoux P.
      • Lacombe P.
      • Monet M.
      • Maciazek J.
      • Krebs L.T.
      • Klonjkowski B.
      • Berrou E.
      • Mericskay M.
      • Li Z.
      • Tournier-Lasserve E.
      • Gridley T.
      • Joutel A.
      Notch3 is required for arterial identity and maturation of vascular smooth muscle cells.
      NOTCH3 contains 34 extracellular epidermal growth factor–like repeats, and each epidermal growth factor–like repeat contains six cysteines that are predicted to form three disulfide bonds important for protein structure.
      • Muiño E.
      • Gallego-Fabrega C.
      • Cullell N.
      • Carrera C.
      • Torres N.
      • Krupinski J.
      • Roquer J.
      • Montaner J.
      • Fernández-Cadenas I.
      Systematic review of cysteine-sparing NOTCH3 missense mutations in patients with clinical suspicion of CADASIL.
      Virtually all CADASIL-causing mutations in NOTCH3 affect cysteines, leading to a change in cysteine number.
      • Joutel A.
      • Vahedi K.
      • Corpechot C.
      • Troesch A.
      • Chabriat H.
      • Vayssière C.
      • Cruaud C.
      • Maciazek J.
      • Weissenbach J.
      • Bousser M.G.
      • Bach J.F.
      • Tournier-Lasserve E.
      Strong clustering and stereotyped nature of Notch3 mutations in CADASIL patients.
      This provides support for the idea that aberrant NOTCH3 undergoes abnormal disulfide bonding that is important for disease pathogenesis. In addition, three-dimensional modeling shows that at least a subset of known NOTCH3 mutations leads to domain misfolding, supporting the notion of misfolded proteins driving CADASIL.
      • Dichgans M.
      • Ludwig H.
      • Müller-Höcker J.
      • Messerschmidt A.
      • Gasser T.
      Small in-frame deletions and missense mutations in CADASIL: 3D models predict misfolding of Notch3 EGF-like repeat domains.
      It is thus postulated that cysteine mutants of NOTCH3 are associated with abnormalities in protein folding and could possess neomorphic properties responsible for CADASIL pathology. Recently, there have also been reports of CADASIL in patients without stereotypical cysteine mutations in NOTCH3.
      • Xiromerisiou G.
      • Marogianni C.
      • Dadouli K.
      • Zompola C.
      • Georgouli D.
      • Provatas A.
      • Theodorou A.
      • Zervas P.
      • Nikolaidou C.
      • Stergiou S.
      • Ntellas P.
      • Sokratous M.
      • Stathis P.
      • Paraskevas G.P.
      • Bonakis A.
      • Voumvourakis K.
      • Hadjichristodoulou C.
      • Hadjigeorgiou G.M.
      • Tsivgoulis G.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy revisited: genotype-phenotype correlations of all published cases.
      Many of the noncysteine involving NOTCH3 mutations are also thought to disrupt NOTCH3 protein structure, and in vitro examination of cysteine-sparing NOTCH3 mutants shows that they also form aggregates similar to those of typical cysteine mutants.
      • Wollenweber F.A.
      • Hanecker P.
      • Bayer-Karpinska A.
      • Malik R.
      • Bäzner H.
      • Moreton F.
      • Muir K.W.
      • Müller S.
      • Giese A.
      • Opherk C.
      • Dichgans M.
      • Haffner C.
      • Duering M.
      Cysteine-sparing CADASIL mutations in NOTCH3 show proaggregatory properties in vitro.
      As of now, no clear genotype-phenotype correlation has been identified, although studies have emerged that suggest a relationship between the position of mutations and disease severity. For example, Rutten et al
      • Rutten J.W.
      • Dauwerse H.G.
      • Gravesteijn G.
      • van Belzen M.J.
      • van der Grond J.
      • Polke J.M.
      • Bernal-Quiros M.
      • Lesnik Oberstein S.A.J.
      Archetypal NOTCH3 mutations frequent in public exome: implications for CADASIL.
      provides evidence to suggest that C-terminal mutants/polymorphisms could contribute to delayed onset small-vessel disease that is less severe compared with the classic CADASIL phenotype. In addition, a recent review of 224 CADASIL case reports suggests that the pathogenicity of CADASIL mutations is related to the location of the mutation.
      • Xiromerisiou G.
      • Marogianni C.
      • Dadouli K.
      • Zompola C.
      • Georgouli D.
      • Provatas A.
      • Theodorou A.
      • Zervas P.
      • Nikolaidou C.
      • Stergiou S.
      • Ntellas P.
      • Sokratous M.
      • Stathis P.
      • Paraskevas G.P.
      • Bonakis A.
      • Voumvourakis K.
      • Hadjichristodoulou C.
      • Hadjigeorgiou G.M.
      • Tsivgoulis G.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy revisited: genotype-phenotype correlations of all published cases.
      Currently, it is thought that both genetic and environmental factors play a role in disease presentation.

      CADASIL Diagnostic Testing

      Presently, the gold standard for diagnosis of CADASIL is genetic testing.
      • Choudhary S.
      • McLeod M.
      • Torchia D.
      • Romanelli P.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL).
      Screening 23 exons of NOTCH3 for mutations has a specificity and sensitivity close to 100%.
      • Choudhary S.
      • McLeod M.
      • Torchia D.
      • Romanelli P.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL).
      Another useful diagnostic tool is skin biopsy. An ultrastructural hallmark of CADASIL is the deposition of granular osmiophilic material (GOM) within vessels.
      • Choudhary S.
      • McLeod M.
      • Torchia D.
      • Romanelli P.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL).
      GOMs are electron-dense extracellular deposits typically found between neighboring vascular smooth muscle cells and most readily visualized by electron microscopy.
      • Choudhary S.
      • McLeod M.
      • Torchia D.
      • Romanelli P.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL).
      In addition to being found in the cerebrovasculature, GOMs can also be detected in extracerebral vasculature, such as that of the skin.
      • Choudhary S.
      • McLeod M.
      • Torchia D.
      • Romanelli P.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL).
      Thus, skin biopsy can be used to diagnose CADASIL with a specificity of 100% and a sensitivity that ranges from 45% to 100%.
      • Di Donato I.
      • Bianchi S.
      • De Stefano N.
      • Dichgans M.
      • Dotti M.T.
      • Duering M.
      • Jouvent E.
      • Korczyn A.D.
      • Lesnik Oberstein S.A.J.
      • Malandrini A.
      • Markus H.S.
      • Pantoni L.
      • Penco S.
      • Rufa A.
      • Sinanović O.
      • Stojanov D.
      • Federico A.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) as a model of small vessel disease: update on clinical, diagnostic, and management aspects.
      Currently, the exact GOM composition remains unclear. However, NOTCH3 ectodomain, N-terminal fragment of NOTCH3 (NTF), metalloproteinase inhibitor 3 (TIMP3), vitronectin (VTN), latent transforming growth factor-β 1 (LTBP1), amyloid P (SAP), annexin 2, and periostin have been identified to be components, suggesting that GOMs consist of abnormal protein aggregates.
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      • Yamamoto Y.
      • Craggs L.J.L.
      • Watanabe A.
      • Booth T.
      • Attems J.
      • Low R.W.C.
      • Oakley A.E.
      • Kalaria R.N.
      Brain microvascular accumulation and distribution of the NOTCH3 ectodomain and granular osmiophilic material in CADASIL.
      • Kast J.
      • Hanecker P.
      • Beaufort N.
      • Giese A.
      • Joutel A.
      • Dichgans M.
      • Opherk C.
      • Haffner C.
      Sequestration of latent TGF-β binding protein 1 into CADASIL-related Notch3-ECD deposits.
      • Nagatoshi A.
      • Ueda M.
      • Ueda A.
      • Tasaki M.
      • Inoue Y.
      • Ma Y.
      • Masuda T.
      • Mizukami M.
      • Matsumoto S.
      • Kosaka T.
      • Kawano T.
      • Ito T.
      • Ando Y.
      Serum amyloid P component: a novel potential player in vessel degeneration in CADASIL.
      • Young K.Z.
      • Lee S.J.
      • Zhang X.
      • Cartee N.M.P.
      • Torres M.
      • Keep S.G.
      • Gabbireddy S.R.
      • Fontana J.L.
      • Qi L.
      • Wang M.M.
      NOTCH3 is non-enzymatically fragmented in inherited cerebral small-vessel disease.
      Finally, presence of subcortical infarcts and leukoencephalopathy is a feature of CADASIL best detected by magnetic resonance imaging (MRI).
      • Chabriat H.
      • Joutel A.
      • Dichgans M.
      • Tournier-Lasserve E.
      • Bousser M.-G.
      Cadasil.
      MRI lesions become apparent at a mean age of 30 years and are found in virtually all patients with CADASIL aged >35 years.
      • Singhal S.
      • Rich P.
      • Markus H.S.
      The spatial distribution of MR imaging abnormalities in cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy and their relationship to age and clinical features.
      Typically, MRI changes precede development of other CADASIL symptoms and worsen with age and disease progression.
      • Chabriat H.
      • Joutel A.
      • Dichgans M.
      • Tournier-Lasserve E.
      • Bousser M.-G.
      Cadasil.
      ,
      • Singhal S.
      • Rich P.
      • Markus H.S.
      The spatial distribution of MR imaging abnormalities in cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy and their relationship to age and clinical features.
      In particular, MRI involvement of the anterior temporal pole has approximately 90% sensitivity and approximately 90% specificity for CADASIL.
      • Di Donato I.
      • Bianchi S.
      • De Stefano N.
      • Dichgans M.
      • Dotti M.T.
      • Duering M.
      • Jouvent E.
      • Korczyn A.D.
      • Lesnik Oberstein S.A.J.
      • Malandrini A.
      • Markus H.S.
      • Pantoni L.
      • Penco S.
      • Rufa A.
      • Sinanović O.
      • Stojanov D.
      • Federico A.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) as a model of small vessel disease: update on clinical, diagnostic, and management aspects.

      Histopathologic Findings in CADASIL

      Pathologic characteristics of CADASIL include GOMs, as mentioned earlier, that surround the vascular smooth muscle cells of arterioles
      • Choudhary S.
      • McLeod M.
      • Torchia D.
      • Romanelli P.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL).
      and the degeneration of vascular smooth muscle cells in arterial walls.
      • Chabriat H.
      • Joutel A.
      • Dichgans M.
      • Tournier-Lasserve E.
      • Bousser M.-G.
      Cadasil.
      In addition, CADASIL brains show significantly thickened arteriolar walls and reduced diameters of small penetrating arteries.
      • Miao Q.
      • Paloneva T.
      • Tuominen S.
      • Pöyhönen M.
      • Tuisku S.
      • Viitanen M.
      • Kalimo H.
      Fibrosis and stenosis of the long penetrating cerebral arteries: the cause of the white matter pathology in cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy.
      It is thought that these thickened arteries and GOMs result from disease-related abnormal protein accumulation that includes molecules such as NOTCH3 ectodomain.
      • Joutel A.
      • Andreux F.
      • Gaulis S.
      • Domenga V.
      • Cécillon M.
      • Battail N.
      • Piga N.
      • Chapon F.
      • Godfrain C.
      • Tournier-Lasserve E.
      The ectodomain of the Notch3 receptor accumulates within the cerebrovasculature of CADASIL patients.

      Protein Processing Involved in CADASIL

      Notch signaling plays a critical role in development and involves processing of NOTCH proteins. NOTCH3 is a transmembrane protein, composed of both an extracellular and a membrane tethered intracellular domain.
      • Wang M.M.
      Notch signaling and Notch signaling modifiers.
      On activation by Notch signaling ligands, a series of proteolytic cleavages occur by ADAM-TACE and γ-secretase, releasing the NOTCH3 intracellular domain to the nucleus to affect gene transcription.
      • Wang M.M.
      Notch signaling and Notch signaling modifiers.
      Although CADASIL is characterized by mutations in NOTCH3, it seems unlikely that a loss of NOTCH3 signaling is the sole driver of disease. Several studies report connections between CADASIL-causing NOTCH3 mutations and NOTCH3 receptor function, which alter canonical Notch signaling.
      • Karlström H.
      • Beatus P.
      • Dannaeus K.
      • Chapman G.
      • Lendahl U.
      • Lundkvist J.
      A CADASIL-mutated Notch 3 receptor exhibits impaired intracellular trafficking and maturation but normal ligand-induced signaling.
      • Arboleda-Velasquez J.F.
      • Manent J.
      • Lee J.H.
      • Tikka S.
      • Ospina C.
      • Vanderburg C.R.
      • Frosch M.P.
      • Rodríguez-Falcón M.
      • Villen J.
      • Gygi S.
      • Lopera F.
      • Kalimo H.
      • Moskowitz M.A.
      • Ayata C.
      • Louvi A.
      • Artavanis-Tsakonas S.
      Hypomorphic Notch 3 alleles link Notch signaling to ischemic cerebral small-vessel disease.
      • Peters N.
      • Opherk C.
      • Zacherle S.
      • Capell A.
      • Gempel P.
      • Dichgans M.
      CADASIL-associated Notch3 mutations have differential effects both on ligand binding and ligand-induced Notch3 receptor signaling through RBP-Jk.
      • Monet-Leprêtre M.
      • Bardot B.
      • Lemaire B.
      • Domenga V.
      • Godin O.
      • Dichgans M.
      • Tournier-Lasserve E.
      • Cohen-Tannoudji M.
      • Chabriat H.
      • Joutel A.
      Distinct phenotypic and functional features of CADASIL mutations in the Notch3 ligand binding domain.
      However, other mouse model studies do not support this idea.
      • Ayata C.
      CADASIL: experimental insights from animal models.
      For example, despite having increased susceptibility to stroke, Notch3 knockout mice do not demonstrate classic features of CADASIL.
      • Domenga V.
      • Fardoux P.
      • Lacombe P.
      • Monet M.
      • Maciazek J.
      • Krebs L.T.
      • Klonjkowski B.
      • Berrou E.
      • Mericskay M.
      • Li Z.
      • Tournier-Lasserve E.
      • Gridley T.
      • Joutel A.
      Notch3 is required for arterial identity and maturation of vascular smooth muscle cells.
      On the other hand, mice that overexpress mutant Notch3 better model the human disorder, although it is also unlikely that CADASIL leads to a gain of Notch signaling function because CADASIL mutations do not enhance downstream signaling.
      • Ayata C.
      CADASIL: experimental insights from animal models.
      The early accumulation of GOMs and NOTCH3 aggregates, found in these mouse models and patients with CADASIL, therefore suggests a neomorphic role of mutant NOTCH3, although it remains unknown whether these aggregates cause disease or are consequences of disease.
      In addition to physiological NOTCH3 processing involved in Notch signaling, post-translational modification of NOTCH3 protein has also been identified as a disease-specific feature of CADASIL. Using conformation-specific antibodies, Zhang et al
      • Zhang X.
      • Lee S.J.
      • Young K.Z.
      • Josephson D.A.
      • Geschwind M.D.
      • Wang M.M.
      Latent NOTCH3 epitopes unmasked in CADASIL and regulated by protein redox state.
      identified a reduced form of NOTCH3 that accumulates specifically in disease-affected CADASIL vessels compared with normal-appearing vessels from age-matched control subjects. It is conceivable that a cysteine-involving mutation results in a change in NOTCH3 tertiary structure or aggregation state. Support for disease-related post-translational alterations of NOTCH3 also comes from Arboleda-Velasquez et al,
      • Arboleda-Velasquez J.F.
      • Rampal R.
      • Fung E.
      • Darland D.C.
      • Liu M.
      • Martinez M.C.
      • Donahue C.P.
      • Navarro-Gonzalez M.F.
      • Libby P.
      • D'Amore P.A.
      • Aikawa M.
      • Haltiwanger R.S.
      • Kosik K.S.
      CADASIL mutations impair Notch3 glycosylation by Fringe.
      who demonstrated impairment of glycosylation and cleavage in mutant CADASIL protein. Both reduced glycosylation and aberrant multimerization of mutant NOTCH3 may play a role in abnormal protein accumulation in CADASIL. In addition, studies identify accumulation of NTF in pathologic vessels compared with normal-appearing vessels, suggesting enhanced cleavage of NOTCH3 protein in disease that is unrelated to Notch signaling.
      • Young K.Z.
      • Lee S.J.
      • Zhang X.
      • Cartee N.M.P.
      • Torres M.
      • Keep S.G.
      • Gabbireddy S.R.
      • Fontana J.L.
      • Qi L.
      • Wang M.M.
      NOTCH3 is non-enzymatically fragmented in inherited cerebral small-vessel disease.
      In vitro studies demonstrate that reduction of NOTCH3, which breaks NOTCH3 disulfide bonding and destabilizes protein structure, enhances nonenzymatic NOTCH3 fragmentation in CADASIL vessels.
      • Young K.Z.
      • Lee S.J.
      • Zhang X.
      • Cartee N.M.P.
      • Torres M.
      • Keep S.G.
      • Gabbireddy S.R.
      • Fontana J.L.
      • Qi L.
      • Wang M.M.
      NOTCH3 is non-enzymatically fragmented in inherited cerebral small-vessel disease.
      Given the disease-specific nature of these NOTCH3 forms, there are likely multiple abnormalities of post-translational processing of NOTCH3 in CADASIL.

      NOTCH3 Assemblies

      Both wild-type NOTCH3 and mutant NOTCH3 are capable of forming dimers, oligomers, and higher-order multimers in vitro.
      • Opherk C.
      • Duering M.
      • Peters N.
      • Karpinska A.
      • Rosner S.
      • Schneider E.
      • Bader B.
      • Giese A.
      • Dichgans M.
      CADASIL mutations enhance spontaneous multimerization of NOTCH3.
      However, mutant NOTCH3 demonstrates increased propensity to form higher-order oligomers compared with wild-type protein.
      • Arboleda-Velasquez J.F.
      • Rampal R.
      • Fung E.
      • Darland D.C.
      • Liu M.
      • Martinez M.C.
      • Donahue C.P.
      • Navarro-Gonzalez M.F.
      • Libby P.
      • D'Amore P.A.
      • Aikawa M.
      • Haltiwanger R.S.
      • Kosik K.S.
      CADASIL mutations impair Notch3 glycosylation by Fringe.
      • Opherk C.
      • Duering M.
      • Peters N.
      • Karpinska A.
      • Rosner S.
      • Schneider E.
      • Bader B.
      • Giese A.
      • Dichgans M.
      CADASIL mutations enhance spontaneous multimerization of NOTCH3.
      • Duering M.
      • Karpinska A.
      • Rosner S.
      • Hopfner F.
      • Zechmeister M.
      • Peters N.
      • Kremmer E.
      • Haffner C.
      • Giese A.
      • Dichgans M.
      • Opherk C.
      Co-aggregate formation of CADASIL-mutant NOTCH3: a single-particle analysis.
      NOTCH3 oligomerization does not require presence of cofactors and is mediated in part by disulfide bonding.
      • Opherk C.
      • Duering M.
      • Peters N.
      • Karpinska A.
      • Rosner S.
      • Schneider E.
      • Bader B.
      • Giese A.
      • Dichgans M.
      CADASIL mutations enhance spontaneous multimerization of NOTCH3.
      ,
      • Duering M.
      • Karpinska A.
      • Rosner S.
      • Hopfner F.
      • Zechmeister M.
      • Peters N.
      • Kremmer E.
      • Haffner C.
      • Giese A.
      • Dichgans M.
      • Opherk C.
      Co-aggregate formation of CADASIL-mutant NOTCH3: a single-particle analysis.
      Furthermore, NOTCH3 binding partners, such as thrombospondin 2 and Fringe, have also been identified to co-aggregate with mutant NOTCH3, suggesting a role of protein aggregation in disease.
      • Arboleda-Velasquez J.F.
      • Rampal R.
      • Fung E.
      • Darland D.C.
      • Liu M.
      • Martinez M.C.
      • Donahue C.P.
      • Navarro-Gonzalez M.F.
      • Libby P.
      • D'Amore P.A.
      • Aikawa M.
      • Haltiwanger R.S.
      • Kosik K.S.
      CADASIL mutations impair Notch3 glycosylation by Fringe.
      ,
      • Duering M.
      • Karpinska A.
      • Rosner S.
      • Hopfner F.
      • Zechmeister M.
      • Peters N.
      • Kremmer E.
      • Haffner C.
      • Giese A.
      • Dichgans M.
      • Opherk C.
      Co-aggregate formation of CADASIL-mutant NOTCH3: a single-particle analysis.
      In addition, NTF, which localizes to pathologically affected vessels in CADASIL, has also been shown to undergo spontaneous thiol-mediated oligomerization in vitro.
      • Young K.Z.
      • Cartee N.M.P.
      • Ivanova M.I.
      • Wang M.M.
      Thiol-mediated and catecholamine-enhanced multimerization of a cerebrovascular disease enriched fragment of NOTCH3.
      NTF can form covalent conjugates with vascular catecholamines that enhance multimerization of the NOTCH3 fragmentation product.
      • Young K.Z.
      • Cartee N.M.P.
      • Ivanova M.I.
      • Wang M.M.
      Thiol-mediated and catecholamine-enhanced multimerization of a cerebrovascular disease enriched fragment of NOTCH3.
      More recent studies have highlighted the presence of NTF multimers in CADASIL tissue, implicating a role for NTF multimers in disease (unpublished data).

      Protein Accumulation and Signaling Dysregulation in CADASIL

      CADASIL brains demonstrate abnormal protein accumulation that is often thought to be linked to disease. For example, CADASIL vessels feature accumulation of the NOTCH3 ectodomain without simultaneous accumulation of the intracellular domain, NOTCH3 aggregates, abnormal NOTCH3 conformations enriched in disease, and NOTCH3 fragments, including an NTF.
      • Young K.Z.
      • Lee S.J.
      • Zhang X.
      • Cartee N.M.P.
      • Torres M.
      • Keep S.G.
      • Gabbireddy S.R.
      • Fontana J.L.
      • Qi L.
      • Wang M.M.
      NOTCH3 is non-enzymatically fragmented in inherited cerebral small-vessel disease.
      ,
      • Joutel A.
      • Andreux F.
      • Gaulis S.
      • Domenga V.
      • Cécillon M.
      • Battail N.
      • Piga N.
      • Chapon F.
      • Godfrain C.
      • Tournier-Lasserve E.
      The ectodomain of the Notch3 receptor accumulates within the cerebrovasculature of CADASIL patients.
      ,
      • Young K.Z.
      • Cartee N.M.P.
      • Ivanova M.I.
      • Wang M.M.
      Thiol-mediated and catecholamine-enhanced multimerization of a cerebrovascular disease enriched fragment of NOTCH3.
      In addition, other proteins, such as proteoglycans, collagens (COLs), and other extracellular matrix (ECM) proteins, have been shown to accumulate within the vessels as well, suggesting formation of extracellular protein complexes in disease.
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Dong H.
      • Blaivas M.
      • Wang M.M.
      Bidirectional encroachment of collagen into the tunica media in cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy.
      • Lee S.J.
      • Zhang X.
      • Wang M.M.
      Vascular accumulation of the small leucine-rich proteoglycan decorin in CADASIL.
      • Zhang X.
      • Lee S.J.
      • Young M.F.
      • Wang M.M.
      The small leucine-rich proteoglycan BGN accumulates in CADASIL and binds to NOTCH3.
      The extracellular matrix is a network of highly cross-linked proteins that plays important roles in cell structure and support.
      • Joutel A.
      • Haddad I.
      • Ratelade J.
      • Nelson M.T.
      Perturbations of the cerebrovascular matrisome: a convergent mechanism in small vessel disease of the brain?.
      ECM proteins are also thought to influence fundamental cellular processes, such as differentiation, survival, and proliferation.
      • Joutel A.
      • Haddad I.
      • Ratelade J.
      • Nelson M.T.
      Perturbations of the cerebrovascular matrisome: a convergent mechanism in small vessel disease of the brain?.
      Traditionally, the ECM is thought to consist of collagens, proteoglycans, and glycoproteins.
      • Joutel A.
      • Haddad I.
      • Ratelade J.
      • Nelson M.T.
      Perturbations of the cerebrovascular matrisome: a convergent mechanism in small vessel disease of the brain?.
      However, there are also ECM-associated proteins, such as ECM regulators, enzymes, or proteins, that modify the ECM and ECM-affiliated proteins.
      • Joutel A.
      • Haddad I.
      • Ratelade J.
      • Nelson M.T.
      Perturbations of the cerebrovascular matrisome: a convergent mechanism in small vessel disease of the brain?.
      The observed enrichment of ECM proteins in CADASIL can be explained by increased secretion of proteins or decreased protein turnover in disease. In mouse models of CADASIL, NOTCH3 ectodomain accumulation is one of the earliest observed events in disease pathogenesis, suggesting a role in disease initiation.
      • Joutel A.
      • Monet-Leprêtre M.
      • Gosele C.
      • Baron-Menguy C.
      • Hammes A.
      • Schmidt S.
      • Lemaire-Carrette B.
      • Domenga V.
      • Schedl A.
      • Lacombe P.
      • Hubner N.
      Cerebrovascular dysfunction and microcirculation rarefaction precede white matter lesions in a mouse genetic model of cerebral ischemic small vessel disease.
      One model of enhanced protein recruitment suggests that NOTCH3 mutations lead to abnormal accumulation of mutant NOTCH3 protein, which then plays a role in recruiting and sequestering other proteins. Recent studies have identified the enrichment of ECM proteins in CADASIL, such as TIMP3, VTN, collagens, LTBP1, clusterin, decorin, biglycan, and laminins, among others.
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Dong H.
      • Blaivas M.
      • Wang M.M.
      Bidirectional encroachment of collagen into the tunica media in cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy.
      • Lee S.J.
      • Zhang X.
      • Wang M.M.
      Vascular accumulation of the small leucine-rich proteoglycan decorin in CADASIL.
      • Zhang X.
      • Lee S.J.
      • Young M.F.
      • Wang M.M.
      The small leucine-rich proteoglycan BGN accumulates in CADASIL and binds to NOTCH3.
      ,
      • Manousopoulou A.
      • Gatherer M.
      • Smith C.
      • Nicoll J.A.R.
      • Woelk C.H.
      • Johnson M.
      • Kalaria R.
      • Attems J.
      • Garbis S.D.
      • Carare R.O.
      Systems proteomic analysis reveals that clusterin and tissue inhibitor of metalloproteinases 3 increase in leptomeningeal arteries affected by cerebral amyloid angiopathy.
      In vitro studies suggest potential physical interaction between NOTCH3 and some of the proteins, including TIMP3 and LTBP1.
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Kast J.
      • Hanecker P.
      • Beaufort N.
      • Giese A.
      • Joutel A.
      • Dichgans M.
      • Opherk C.
      • Haffner C.
      Sequestration of latent TGF-β binding protein 1 into CADASIL-related Notch3-ECD deposits.
      Transforming growth factor (TGF)-β signaling activity is linked to regulation of fibrotic events in the vasculature, and is shown to be activated by fibronectin, fibrillin-1, and other members of the LTBP family.
      • Kast J.
      • Hanecker P.
      • Beaufort N.
      • Giese A.
      • Joutel A.
      • Dichgans M.
      • Opherk C.
      • Haffner C.
      Sequestration of latent TGF-β binding protein 1 into CADASIL-related Notch3-ECD deposits.
      This supports a model where dysregulation of TGF-β signaling results from aggregation and/or accumulation of NOTCH3 in the cerebrovasculature of patients with CADASIL and promotes abnormal recruitment of ECM proteins.
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      The resulting aggregation of proteins can result in a change in biological function and contribute to disease. Interestingly, TGF-β activity has been implicated in other vasculopathies, including cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathy (CARASIL) and CAA.
      • Kast J.
      • Hanecker P.
      • Beaufort N.
      • Giese A.
      • Joutel A.
      • Dichgans M.
      • Opherk C.
      • Haffner C.
      Sequestration of latent TGF-β binding protein 1 into CADASIL-related Notch3-ECD deposits.
      Another model of abnormal protein accumulation involves decreased protein turnover and proposes a loss of serine protease HTRA1 activity in CADASIL. HTRA1 is a negative regulator of TGF-β signaling and an extracellular serine protease that is known to be mutated in CARASIL, an autosomal recessive disorder that shares clinical overlap with CADASIL.
      • Nozaki H.
      • Sekine Y.
      • Fukutake T.
      • Nishimoto Y.
      • Shimoe Y.
      • Shirata A.
      • Yanagawa S.
      • Hirayama M.
      • Tamura M.
      • Nishizawa M.
      • Onodera O.
      Characteristic features and progression of abnormalities on MRI for CARASIL.
      In a recent liquid chromatography–tandem mass spectrometry study of CADASIL brain tissue, significantly increased levels of HTRA1 protein and substrates were identified (clusterin, vitronectin, elastin, and LTBP1), suggesting an impairment of HTRA1 activity in CADASIL.
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.

      Cerebral Amyloid Angiopathy

      Clinical Overview of CAA

      CAA is a common age-related disease that can occur both with and without Alzheimer disease (AD).
      • Attems J.
      • Jellinger K.
      • Thal D.R.
      • Van Nostrand W.
      Review: sporadic cerebral amyloid angiopathy.
      Although CAA can be caused by several amyloidogenic proteins, such as cystatin C, transthyretin, and others, this review will focus on amyloid-β (Aβ) CAA. Population-based studies demonstrate that CAA affects roughly 20% to 40% of elderly populations without dementia and 50% to 60% of elderly populations with dementia.
      • Tuttolomondo A.
      • Maugeri R.
      • Orlando E.
      • Giannone G.
      • Ciccia F.
      • Rizzo A.
      • Di Raimondo D.
      • Graziano F.
      • Pecoraro R.
      • Maida C.
      • Simonetta I.
      • Cirrincione A.
      • Portelli F.
      • Corpora F.
      • Iacopino D.G.
      • Pinto A.
      β-Amyloid wall deposit of temporal artery in subjects with spontaneous intracerebral haemorrhage.
      In AD, CAA is predicted to be present in roughly 80% to 90% of patients.
      • Yamada M.
      Cerebral amyloid angiopathy: emerging concepts.
      Classic clinical presentation of CAA involves spontaneous lobar intracerebral hemorrhage, cognitive impairment and/or dementia, and transient focal neurologic episodes.
      • Yamada M.
      Cerebral amyloid angiopathy: emerging concepts.

      Genetics of CAA

      Most cases of CAA are sporadic. However, some CAA has been linked to specific genetic loci that include apolipoprotein E (APOE) e4 and e2 alleles, mutations in presenilin 1 (PS1) and PS2, and mutations in amyloid precursor protein (APP).
      • Yamada M.
      Cerebral amyloid angiopathy: emerging concepts.
      Mutations in APP that cause familial forms of CAA (eg, hereditary cerebral hemorrhage with amyloidosis, Dutch type) tend to cluster around residues 21 to 23 and have been proposed to decrease proteolytic degradation of Aβ or attenuate Aβ protein clearance from the brain into the circulation.
      • Tsubuki S.
      • Takaki Y.
      • Saido T.C.
      Dutch, Flemish, Italian, and Arctic mutations of APP and resistance of Abeta to physiologically relevant proteolytic degradation.
      On the other hand, two isoforms of APOE are thought to promote CAA via different mechanisms. For example, APOE e4 has been linked to increased amyloid deposition, whereas APOE e2 is thought to accelerate the formation of vasculopathies that promote vessel rupture.
      • Tai L.M.
      • Thomas R.
      • Marottoli F.M.
      • Koster K.P.
      • Kanekiyo T.
      • Morris A.W.J.
      • Bu G.
      The role of APOE in cerebrovascular dysfunction.
      Both APOE e2 and e4 alleles have been associated with early recurrence of lobar intracerebral hemorrhage following a prior lobar intracerebral hemorrhage.
      • Yamada M.
      Cerebral amyloid angiopathy: emerging concepts.
      In addition, alterations in TGF-β1 have been linked to CAA, along with polymorphisms in a1-antichymotrypsin, neprilysin, low-density lipoprotein receptor protein 1, CR1, and angiotensin-converting enzyme genes.
      • Yamada M.
      Cerebral amyloid angiopathy: emerging concepts.

      CAA Diagnostic Testing

      Definitive diagnosis of CAA relies on identification of CAA-related vascular damage, multiple lobar hemorrhages, and absence of alternative pathologies on postmortem examination of brain, as assessed by the Boston criteria.
      • Linn J.
      • Halpin A.
      • Demaerel P.
      • Ruhland J.
      • Giese A.D.
      • Dichgans M.
      • van Buchem M.A.
      • Bruckmann H.
      • Greenberg S.M.
      Prevalence of superficial siderosis in patients with cerebral amyloid angiopathy.
      Diagnosis of probable CAA includes use of clinical data and MRI imaging to identify the presence of multiple hemorrhages or microbleeds in regions typical of CAA, and more recently in the modified Boston criteria, the presence of superficial siderosis.
      • Linn J.
      • Halpin A.
      • Demaerel P.
      • Ruhland J.
      • Giese A.D.
      • Dichgans M.
      • van Buchem M.A.
      • Bruckmann H.
      • Greenberg S.M.
      Prevalence of superficial siderosis in patients with cerebral amyloid angiopathy.
      Although many imaging features are shared between CADASIL and CAA (cerebral microbleeds and white matter hyperintensities), cortical superficial siderosis is absent in a large cohort of patients with CADASIL from a clinical study examining imaging features of CADASIL, CAA, and control subjects.
      • Wollenweber F.A.
      • Baykara E.
      • Zedde M.
      • Gesierich B.
      • Achmüller M.
      • Jouvent E.
      • Viswanathan A.
      • Ropele S.
      • Chabriat H.
      • Schmidt R.
      • Opherk C.
      • Dichgans M.
      • Linn J.
      • Duering M.
      Cortical superficial siderosis in different types of cerebral small vessel disease.
      Thus, this study posits that in individuals with imaging features suggestive of small-vessel disease, presence of cortical superficial siderosis is highly suggestive of CAA.
      • Wollenweber F.A.
      • Baykara E.
      • Zedde M.
      • Gesierich B.
      • Achmüller M.
      • Jouvent E.
      • Viswanathan A.
      • Ropele S.
      • Chabriat H.
      • Schmidt R.
      • Opherk C.
      • Dichgans M.
      • Linn J.
      • Duering M.
      Cortical superficial siderosis in different types of cerebral small vessel disease.

      Histopathologic Findings in CAA

      Histopathologically, CAA is characterized by extracellular Aβ protein deposits in the cerebrovasculature.
      • Glenner G.G.
      • Wong C.W.
      Alzheimer's disease: initial report of the purification and characterization of a novel cerebrovascular amyloid protein.
      The most common forms of vascular amyloid deposits include Aβ1-40 and Aβ1-42.
      • Glenner G.G.
      • Wong C.W.
      Alzheimer's disease: initial report of the purification and characterization of a novel cerebrovascular amyloid protein.
      ,
      • Roher A.E.
      • Lowenson J.D.
      • Clarke S.
      • Woods A.S.
      • Cotter R.J.
      • Gowing E.
      • Ball M.J.
      beta-Amyloid-(1-42) is a major component of cerebrovascular amyloid deposits: implications for the pathology of Alzheimer disease.
      Many sources of Aβ have been proposed to explain its deposition within the vessel wall, including transport across the blood-brain barrier (BBB) from the circulation, the vascular smooth muscle cells, and neurons as a result of impaired perivascular drainage.
      • Weller R.O.
      • Massey A.
      • Newman T.A.
      • Hutchings M.
      • Kuo Y.M.
      • Roher A.E.
      Cerebral amyloid angiopathy: amyloid beta accumulates in putative interstitial fluid drainage pathways in Alzheimer's disease.
      • Wisniewski H.M.
      • Fragkowiak J.
      • Zoltowska A.
      • Kim K.S.
      Vascular β-amyloid in Alzheimer's disease angiopathy is produced by proliferating and degenerating smooth muscle cells.
      • Bell R.D.
      • Zlokovic B.V.
      Neurovascular mechanisms and blood-brain barrier disorder in Alzheimer's disease.
      In most cases, there is no direct evidence of overproduction of Aβ within the vessel wall. Instead, the perivascular drainage model suggests that the Aβ accumulation within the vasculature is likely the result of impaired Aβ drainage through perivascular pathways that typically serve as lymphatic-like drainage pathways from the brain.
      • Weller R.O.
      • Subash M.
      • Preston S.D.
      • Mazanti I.
      • Carare R.O.
      Perivascular drainage of amyloid-beta peptides from the brain and its failure in cerebral amyloid angiopathy and Alzheimer's disease.
      The impaired elimination of proteins results in the accumulation of both soluble and insoluble protein aggregates in the extracellular spaces within arterial and capillary walls. Alterations of these extracellular spaces in disease or aging might further contribute to decreases in drainage capacity and increased Aβ deposition.
      • Weller R.O.
      • Subash M.
      • Preston S.D.
      • Mazanti I.
      • Carare R.O.
      Perivascular drainage of amyloid-beta peptides from the brain and its failure in cerebral amyloid angiopathy and Alzheimer's disease.
      CAA vessels also feature marked degeneration of smooth muscle cells within the medial layer of arterioles and hyalinization.
      • Charidimou A.
      • Boulouis G.
      • Gurol M.E.
      • Ayata C.
      • Bacskai B.J.
      • Frosch M.P.
      • Viswanathan A.
      • Greenberg S.M.
      Emerging concepts in sporadic cerebral amyloid angiopathy.
      Severely affected vessels demonstrate disruption of the vascular architecture, leading to microaneurysm formation, fibrinoid necrosis, and Aβ deposition in the surrounding neuropil (alias dysphoric changes).
      • Attems J.
      • Jellinger K.
      • Thal D.R.
      • Van Nostrand W.
      Review: sporadic cerebral amyloid angiopathy.
      The two major types of CAA include type 1, which involves cortical capillaries and/or larger vessels, and type 2, which is limited to larger leptomeningeal and cortical arteries.
      • Attems J.
      • Jellinger K.
      • Thal D.R.
      • Van Nostrand W.
      Review: sporadic cerebral amyloid angiopathy.
      ,
      • Charidimou A.
      • Boulouis G.
      • Gurol M.E.
      • Ayata C.
      • Bacskai B.J.
      • Frosch M.P.
      • Viswanathan A.
      • Greenberg S.M.
      Emerging concepts in sporadic cerebral amyloid angiopathy.
      CAA type 1 leads to capillary occlusion and alterations in vascular flow, contributing to the development of dementia.
      • Yamada M.
      Cerebral amyloid angiopathy: emerging concepts.

      Protein Processing Involved in CAA

      Aβ is generated from cleavage of APP.
      • Yamada M.
      Cerebral amyloid angiopathy: emerging concepts.
      APP is a membrane glycoprotein intimately involved in neuronal development, maintenance of neuronal homeostasis, cellular signaling, and intracellular transport.
      • Chen G.-F.
      • Xu T.-H.
      • Yan Y.
      • Zhou Y.-R.
      • Jiang Y.
      • Melcher K.
      • Xu H.E.
      Amyloid beta: structure, biology and structure-based therapeutic development.
      APP can generate many different cleavage products, some of which are thought to be major contributors to Aβ deposits throughout the brain.
      • Chen G.-F.
      • Xu T.-H.
      • Yan Y.
      • Zhou Y.-R.
      • Jiang Y.
      • Melcher K.
      • Xu H.E.
      Amyloid beta: structure, biology and structure-based therapeutic development.
      Under physiological conditions, APP is cleaved by α-secretase to shed the ectodomain, which does not produce amyloidogenic fragments.
      • Zhang Y.-W.
      • Thompson R.
      • Zhang H.
      • Xu H.
      APP processing in Alzheimer's disease.
      Alternatively, ectodomain shedding can occur by β secretase activity.
      • Zhang Y.-W.
      • Thompson R.
      • Zhang H.
      • Xu H.
      APP processing in Alzheimer's disease.
      Aβ is generated by β-secretase and γ-secretase cleavage to generate two predominant forms: Aβ1-40 and Aβ1-42.
      • Yamada M.
      Cerebral amyloid angiopathy: emerging concepts.
      1-40 is more often found within the vessel wall, whereas Aβ1-42 is more often found in senile plaques and capillaries involved in CAA.
      • Roher A.E.
      • Lowenson J.D.
      • Clarke S.
      • Woods A.S.
      • Cotter R.J.
      • Gowing E.
      • Ball M.J.
      beta-Amyloid-(1-42) is a major component of cerebrovascular amyloid deposits: implications for the pathology of Alzheimer disease.
      The major β-secretase is BACE1, which has a main cleavage site at position 1D of Aβ, but also has an alternative site at 11E, generating Aβ11-40/42.
      • Zhang Y.-W.
      • Thompson R.
      • Zhang H.
      • Xu H.
      APP processing in Alzheimer's disease.
      A second cleavage process occurs via the γ-secretase complex, consisting of presenilin, nicastrin, anterior pharynx-defective-1, and presenilin enhancer-2.
      • Zhang Y.-W.
      • Thompson R.
      • Zhang H.
      • Xu H.
      APP processing in Alzheimer's disease.
      The γ-secretase complex results in the intramembranous proteolysis of the β-secretase cleaved product to generate Aβ1-40/42.
      • Zhang Y.-W.
      • Thompson R.
      • Zhang H.
      • Xu H.
      APP processing in Alzheimer's disease.
      The γ-secretase complex is also essential for physiological processing of NOTCH3.

      Aβ Assemblies

      Aβ monomers can form various types of assemblies, such as oligomers, protofibrils, and amyloid fibrils.
      • Chen G.-F.
      • Xu T.-H.
      • Yan Y.
      • Zhou Y.-R.
      • Jiang Y.
      • Melcher K.
      • Xu H.E.
      Amyloid beta: structure, biology and structure-based therapeutic development.
      In AD, insoluble amyloid fibrils can further assemble into amyloid plaques most commonly found in the neocortex of AD brains, whereas soluble amyloid oligomers can deposit throughout the brain.
      • Chen G.-F.
      • Xu T.-H.
      • Yan Y.
      • Zhou Y.-R.
      • Jiang Y.
      • Melcher K.
      • Xu H.E.
      Amyloid beta: structure, biology and structure-based therapeutic development.
      The dominant form in CAA is fibrillar Aβ, although a recent study found that most mutations in the Aβ sequence that promote CAA and AD do not have obvious stabilizing or destabilizing effects on Aβ fibrils derived from AD brains.
      • Kollmer M.
      • Close W.
      • Funk L.
      • Rasmussen J.
      • Bsoul A.
      • Schierhorn A.
      • Schmidt M.
      • Sigurdson C.J.
      • Jucker M.
      • Fändrich M.
      Cryo-EM structure and polymorphism of Aβ amyloid fibrils purified from Alzheimer's brain tissue.
      Both soluble and insoluble Aβ assemblies are thought to contribute to cerebrovascular dysfunction.
      Accumulation of Aβ has been proposed to exert neuronal toxicity through various means, and it is thought that assemblies of oligomeric Aβ result in activation of microglia and astrocytes, oligomerization and aggregation of tau protein, and progressive neuronal loss.
      • Chen G.-F.
      • Xu T.-H.
      • Yan Y.
      • Zhou Y.-R.
      • Jiang Y.
      • Melcher K.
      • Xu H.E.
      Amyloid beta: structure, biology and structure-based therapeutic development.
      Alternatively, accumulation of fibrillar Aβ in the vasculature has been proposed to impact blood vessel integrity and function.
      • Nicoll J.A.R.
      • Yamada M.
      • Frackowiak J.
      • Mazur-Kolecka B.
      • Weller R.O.
      Cerebral amyloid angiopathy plays a direct role in the pathogenesis of Alzheimer's disease: pro-CAA position statement.
      ,
      • Wisniewski H.M.
      • Wegiel J.
      • Vorbrodt A.W.
      • Mazur-Kolecka B.
      • Frackowiak J.
      Role of perivascular cells and myocytes in vascular amyloidosis.
      Deposition of amyloid in capillaries has been linked to degeneration of the lumen, endothelium, and basal lamina, resulting in ischemia and neurodegeneration.
      • Wisniewski H.M.
      • Wegiel J.
      • Vorbrodt A.W.
      • Mazur-Kolecka B.
      • Frackowiak J.
      Role of perivascular cells and myocytes in vascular amyloidosis.
      Amyloid fibrils found in the perivascular space, indicating dysphoric changes, have also been linked to neurite dystrophy.
      • Wisniewski H.M.
      • Wegiel J.
      • Vorbrodt A.W.
      • Mazur-Kolecka B.
      • Frackowiak J.
      Role of perivascular cells and myocytes in vascular amyloidosis.
      In larger vessels, Aβ deposition is associated with degeneration of vascular smooth muscle cells.
      • Wisniewski H.M.
      • Wegiel J.
      • Vorbrodt A.W.
      • Mazur-Kolecka B.
      • Frackowiak J.
      Role of perivascular cells and myocytes in vascular amyloidosis.
      Furthermore, it is thought that accumulation of Aβ in the vessel wall can impair perivascular draining of Aβ, further promoting accumulation of protein.
      • Nicoll J.A.R.
      • Yamada M.
      • Frackowiak J.
      • Mazur-Kolecka B.
      • Weller R.O.
      Cerebral amyloid angiopathy plays a direct role in the pathogenesis of Alzheimer's disease: pro-CAA position statement.

      Protein Accumulation and Signaling Dysregulation in CAA

      Numerous protein-based studies have highlighted CAA-specific differential regulation of proteins in addition to amyloidogenic proteins, with many of the differentially regulated proteins related to extracellular structure and matrix organization. Several proposed mechanisms exist to explain the increase in ECM proteins in CAA. For example, this could be explained by either increased secretion of extracellular matrix components or decreased turnover.
      One model suggests that tissue injury or cellular processes result in enhanced synthesis of ECM proteins. Disruption of the BBB has been implicated in CAA both with and without AD pathology, with identification of BBB leakage markers, such as fibrinogen, and a decrease in tight junction proteins (occludin, claudin-5, and tight junction protein 1) important for maintenance of the BBB.
      • Magaki S.
      • Tang Z.
      • Tung S.
      • Williams C.K.
      • Lo D.
      • Yong W.H.
      • Khanlou N.
      • Vinters H.V.
      The effects of cerebral amyloid angiopathy on integrity of the blood-brain barrier.
      • Carrano A.
      • Hoozemans J.J.M.
      • van der Vies S.M.
      • Rozemuller A.J.M.
      • van Horssen J.
      • de Vries H.E.
      Amyloid beta induces oxidative stress-mediated blood-brain barrier changes in capillary amyloid angiopathy.
      • Carrano A.
      • Hoozemans J.J.M.
      • van der Vies S.M.
      • van Horssen J.
      • de Vries H.E.
      • Rozemuller A.J.M.
      Neuroinflammation and blood-brain barrier changes in capillary amyloid angiopathy.
      Thus, it is conceivable that enhanced cross-linking of ECM proteins might result as a response to strengthen weakened vessel walls and prevent further BBB leakage.
      • Zipfel G.J.
      • Han H.
      • Ford A.L.
      • Lee J.-M.
      Cerebral amyloid angiopathy: progressive disruption of the neurovascular unit.
      Covalent protein cross-linking results in increased formation and stability of extracellular protein complexes. Tissue transglutaminase is a known modifier of proteins implicated in AD,
      • Appelt D.M.
      • Kopen G.C.
      • Boyne L.J.
      • Balin B.J.
      Localization of transglutaminase in hippocampal neurons: implications for Alzheimer's disease.
      • Johnson G.V.
      • Cox T.M.
      • Lockhart J.P.
      • Zinnerman M.D.
      • Miller M.L.
      • Powers R.E.
      Transglutaminase activity is increased in Alzheimer's disease brain.
      • Wang D.-S.
      • Dickson D.W.
      • Malter J.S.
      Tissue transglutaminase, protein cross-linking and Alzheimer's disease: review and views.
      and enhanced tissue transglutaminase activity has been identified in hereditary cerebral hemorrhage with amyloidosis, Dutch type, and CAA with AD pathology. Tissue transglutaminase cross-links proteins, such as Aβ and APP, and additional ECM proteins, such as fibronectin and laminin, potentially leading to their enhancement in CAA.
      • de Jager M.
      • van der Wildt B.
      • Schul E.
      • Bol J.G.J.M.
      • van Duinen S.G.
      • Drukarch B.
      • Wilhelmus M.M.M.
      Tissue transglutaminase colocalizes with extracellular matrix proteins in cerebral amyloid angiopathy.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      In the ECM, tissue transglutaminase and tissue injury are also capable of activating TGF-β, which stimulates synthesis of ECM proteins and protease inhibitors that prevent enzymatic breakdown of the ECM.
      • Verrecchia F.
      • Mauviel A.
      Transforming growth factor-beta signaling through the Smad pathway: role in extracellular matrix gene expression and regulation.
      ,
      • Verderio E.
      • Gaudry C.
      • Gross S.
      • Smith C.
      • Downes S.
      • Griffin M.
      Regulation of cell surface tissue transglutaminase: effects on matrix storage of latent transforming growth factor-beta binding protein-1.
      In support of this notion, expression of TGF-β1 and TGFβR2 is increased in hereditary cerebral hemorrhage with amyloidosis, Dutch type, a genetic form of CAA that results in both accelerated and more severe CAA pathology and symptoms.
      • Grand Moursel L.
      • Munting L.P.
      • van der Graaf L.M.
      • van Duinen S.G.
      • Goumans M.-J.T.H.
      • Ueberham U.
      • Natté R.
      • van Buchem M.A.
      • van Roon-Mom W.M.C.
      • van der Weerd L.
      TGFβ pathway deregulation and abnormal phospho-SMAD2/3 staining in hereditary cerebral hemorrhage with amyloidosis-Dutch type.
      In addition, immunohistochemical staining of downstream TGF-β pathway signaling molecules, such as phosphorylated SMAD2/3, and TGF-β regulated proteins, such as fibronectin, collagen, and TIMP3, has been identified.
      • de Jager M.
      • van der Wildt B.
      • Schul E.
      • Bol J.G.J.M.
      • van Duinen S.G.
      • Drukarch B.
      • Wilhelmus M.M.M.
      Tissue transglutaminase colocalizes with extracellular matrix proteins in cerebral amyloid angiopathy.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      ,
      • Grand Moursel L.
      • Munting L.P.
      • van der Graaf L.M.
      • van Duinen S.G.
      • Goumans M.-J.T.H.
      • Ueberham U.
      • Natté R.
      • van Buchem M.A.
      • van Roon-Mom W.M.C.
      • van der Weerd L.
      TGFβ pathway deregulation and abnormal phospho-SMAD2/3 staining in hereditary cerebral hemorrhage with amyloidosis-Dutch type.
      ,
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      Similarly, TGF-β activity has been implicated in other ECM affecting diseases, such as AD, CADASIL, and CARASIL.
      • Kast J.
      • Hanecker P.
      • Beaufort N.
      • Giese A.
      • Joutel A.
      • Dichgans M.
      • Opherk C.
      • Haffner C.
      Sequestration of latent TGF-β binding protein 1 into CADASIL-related Notch3-ECD deposits.
      ,
      • Grand Moursel L.
      • Munting L.P.
      • van der Graaf L.M.
      • van Duinen S.G.
      • Goumans M.-J.T.H.
      • Ueberham U.
      • Natté R.
      • van Buchem M.A.
      • van Roon-Mom W.M.C.
      • van der Weerd L.
      TGFβ pathway deregulation and abnormal phospho-SMAD2/3 staining in hereditary cerebral hemorrhage with amyloidosis-Dutch type.
      Another potential explanation for the increase in ECM proteins is the decreased turnover and removal of ECM components. This could be due to aggregation and sequestration of chaperones, contributing to further aggregation and deposition of proteins. In CAA, enrichment of extracellular chaperones, such as clusterin, APOE, and HTRA1, was identified.
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      ,
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      APOE features chaperone function that targets Aβ assemblies, and clusterin and HTRA1 are also thought to be involved in removal of protein aggregates.
      • Castano E.M.
      • Prelli F.
      • Wisniewski T.
      • Golabek A.
      • Kumar R.A.
      • Soto C.
      • Frangione B.
      Fibrillogenesis in Alzheimer's disease of amyloid beta peptides and apolipoprotein E.
      ,
      • Haffner C.
      • Malik R.
      • Dichgans M.
      Genetic factors in cerebral small vessel disease and their impact on stroke and dementia.
      Sequestration of protein chaperones can thus contribute to aggregation of Aβ and other extracellular proteins. In addition, tissue endogenous inhibitors (TIMPs) and matrix metalloproteinases (MMPs) are involved in regulation of the ECM, and dysregulation of these proteases can result in decreased turnover of ECM components and/or damage the integrity of the BBB, contributing to lobar hemorrhage.
      • Zipfel G.J.
      • Han H.
      • Ford A.L.
      • Lee J.-M.
      Cerebral amyloid angiopathy: progressive disruption of the neurovascular unit.
      ,
      • Lepelletier F.-X.
      • Mann D.M.A.
      • Robinson A.C.
      • Pinteaux E.
      • Boutin H.
      Early changes in extracellular matrix in Alzheimer's disease.
      ,
      • Jäkel L.
      • Kuiperij H.B.
      • Gerding L.P.
      • Custers E.E.M.
      • van den Berg E.
      • Jolink W.M.T.
      • Schreuder F.H.B.M.
      • Küsters B.
      • Klijn C.J.M.
      • Verbeek M.M.
      Disturbed balance in the expression of MMP9 and TIMP3 in cerebral amyloid angiopathy-related intracerebral haemorrhage.
      MMP2 and MMP9 are enhanced in CAA with AD pathology, and matrix metalloproteinase inhibitor TIMP3 is enhanced in CAA both with and without underlying AD pathology.
      • Manousopoulou A.
      • Gatherer M.
      • Smith C.
      • Nicoll J.A.R.
      • Woelk C.H.
      • Johnson M.
      • Kalaria R.
      • Attems J.
      • Garbis S.D.
      • Carare R.O.
      Systems proteomic analysis reveals that clusterin and tissue inhibitor of metalloproteinases 3 increase in leptomeningeal arteries affected by cerebral amyloid angiopathy.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      ,
      • Jäkel L.
      • Kuiperij H.B.
      • Gerding L.P.
      • Custers E.E.M.
      • van den Berg E.
      • Jolink W.M.T.
      • Schreuder F.H.B.M.
      • Küsters B.
      • Klijn C.J.M.
      • Verbeek M.M.
      Disturbed balance in the expression of MMP9 and TIMP3 in cerebral amyloid angiopathy-related intracerebral haemorrhage.
      ,
      • Hernandez-Guillamon M.
      • Martinez-Saez E.
      • Delgado P.
      • Domingues-Montanari S.
      • Boada C.
      • Penalba A.
      • Boada M.
      • Pagola J.
      • Maisterra O.
      • Rodriguez-Luna D.
      • Molina C.A.
      • Rovira A.
      • Alvarez-Sabin J.
      • Ortega-Aznar A.
      • Montaner J.
      MMP-2/MMP-9 plasma level and brain expression in cerebral amyloid angiopathy-associated hemorrhagic stroke.
      MMP2 and MMP9 have also been shown to proteolyze various Aβ peptides in AD.
      • Hernandez-Guillamon M.
      • Mawhirt S.
      • Fossati S.
      • Blais S.
      • Pares M.
      • Penalba A.
      • Boada M.
      • Couraud P.-O.
      • Neubert T.A.
      • Montaner J.
      • Ghiso J.
      • Rostagno A.
      Matrix metalloproteinase 2 (MMP-2) degrades soluble vasculotropic amyloid-beta E22Q and L34V mutants, delaying their toxicity for human brain microvascular endothelial cells.
      ,
      • Yan P.
      • Hu X.
      • Song H.
      • Yin K.
      • Bateman R.J.
      • Cirrito J.R.
      • Xiao Q.
      • Hsu F.F.
      • Turk J.W.
      • Xu J.
      • Hsu C.Y.
      • Holtzman D.M.
      • Lee J.-M.
      Matrix metalloproteinase-9 degrades amyloid-beta fibrils in vitro and compact plaques in situ.
      Thus, dysregulation of proteases can contribute to accumulation of ECM proteins identified in CAA.

      CADASIL and CAA

      CADASIL and CAA Exhibit Distinct Histopathologies

      CADASIL and CAA can be clearly distinguished by histologic evaluation. Key histologic findings of CADASIL include accumulation of NOTCH3 protein ectodomain within the vasculature and the pathognomonic presence of ultrastructural extracellular GOMs.
      • Domenga V.
      • Fardoux P.
      • Lacombe P.
      • Monet M.
      • Maciazek J.
      • Krebs L.T.
      • Klonjkowski B.
      • Berrou E.
      • Mericskay M.
      • Li Z.
      • Tournier-Lasserve E.
      • Gridley T.
      • Joutel A.
      Notch3 is required for arterial identity and maturation of vascular smooth muscle cells.
      ,
      • Joutel A.
      • Andreux F.
      • Gaulis S.
      • Domenga V.
      • Cécillon M.
      • Battail N.
      • Piga N.
      • Chapon F.
      • Godfrain C.
      • Tournier-Lasserve E.
      The ectodomain of the Notch3 receptor accumulates within the cerebrovasculature of CADASIL patients.
      In addition, CADASIL vessels demonstrate dramatic intimal hyperplasia and accumulation of intimal proteins (Figure 1A).
      • Gatti J.R.
      • Zhang X.
      • Korcari E.
      • Lee S.J.
      • Greenstone N.
      • Dean J.G.
      • Maripudi S.
      • Wang M.M.
      Redistribution of mature smooth muscle markers in brain arteries in cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy.
      ,
      • Dong H.
      • Ding H.
      • Young K.
      • Blaivas M.
      • Christensen P.J.
      • Wang M.M.
      Advanced intimal hyperplasia without luminal narrowing of leptomeningeal arteries in CADASIL.
      In comparison, CAA is characterized by cerebrovascular accumulation of amyloidogenic protein, such as Aβ (Figure 1A).
      • Glenner G.G.
      • Wong C.W.
      Alzheimer's disease: initial report of the purification and characterization of a novel cerebrovascular amyloid protein.
      ,
      • Roher A.E.
      • Lowenson J.D.
      • Clarke S.
      • Woods A.S.
      • Cotter R.J.
      • Gowing E.
      • Ball M.J.
      beta-Amyloid-(1-42) is a major component of cerebrovascular amyloid deposits: implications for the pathology of Alzheimer disease.
      Routine histopathologic distinction is straightforward, because CADASIL vessels have not been shown to contain amyloid.
      • Xiromerisiou G.
      • Marogianni C.
      • Dadouli K.
      • Zompola C.
      • Georgouli D.
      • Provatas A.
      • Theodorou A.
      • Zervas P.
      • Nikolaidou C.
      • Stergiou S.
      • Ntellas P.
      • Sokratous M.
      • Stathis P.
      • Paraskevas G.P.
      • Bonakis A.
      • Voumvourakis K.
      • Hadjichristodoulou C.
      • Hadjigeorgiou G.M.
      • Tsivgoulis G.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy revisited: genotype-phenotype correlations of all published cases.
      Figure thumbnail gr1
      Figure 1Differences and similarities between cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) and cerebral amyloid angiopathy (CAA) histopathology and protein profiles. A: The histopathologies of CADASIL and CAA vessels are compared. Both CADASIL and CAA feature abnormal protein accumulation. In CADASIL, the major protein involved is NOTCH3 ectodomain (brown deposits), whereas in CAA, the major protein involved is amyloid-β (Aβ; green deposits). CADASIL vessels also demonstrate dramatic intimal hyperplasia with accumulation of intimal proteins. Both CADASIL and CAA vessels involve significant smooth muscle cell degeneration in the medial layer and hyalinization of the vessel walls. B: Examination of published literature indicated 378 proteins differentially regulated in CADASIL and 58 proteins differentially regulated in vascular CAA. Of these, approximately 33% of the proteins differentially regulated in vascular CAA overlap with those differentially regulated in CADASIL. C: STRING version 11 analysis shows a high degree of interconnectedness among the shared proteins.
      • Szklarczyk D.
      • Gable A.L.
      • Lyon D.
      • Junge A.
      • Wyder S.
      • Huerta-Cepas J.
      • Simonovic M.
      • Doncheva N.T.
      • Morris J.H.
      • Bork P.
      • Jensen L.J.
      • Mering C.V.
      STRING v11: protein-protein association networks with increased coverage, supporting functional discovery in genome-wide experimental datasets.
      Only norrin (NDP) does not have any known or predicted interactions with the other proteins. APCS, serum amyloid protein; APOE, apolipoprotein E; CLU, clusterin; COL1A2, collagen α-2(I) chain; COL6A2, collagen α-2(VI) chain; COL6A3, collagen α-3(VI) chain; C4A, complement C4-A; CST3, cystatin C; FN1, fibronectin; GFAP, glial fibrillary acidic protein; HSPG2, basement membrane–specific heparan sulfate proteoglycan core protein; HTRA1, serine protease HTRA1; LAMC1, laminin subunit γ 1; NFASC, neurofascin; PTGDS, prostaglandin H2 D isomerase; TIMP3, metalloproteinase inhibitor 3; TPM1, tropomyosin α 1 chain; VTN, vitronectin.

      Shared Features between CADASIL and CAA

      There is significant clinical overlap in CADASIL and CAA disease presentation, with both disorders resulting in increased risk of dementia, stroke, and intracerebral hemorrhage.
      • Di Donato I.
      • Bianchi S.
      • De Stefano N.
      • Dichgans M.
      • Dotti M.T.
      • Duering M.
      • Jouvent E.
      • Korczyn A.D.
      • Lesnik Oberstein S.A.J.
      • Malandrini A.
      • Markus H.S.
      • Pantoni L.
      • Penco S.
      • Rufa A.
      • Sinanović O.
      • Stojanov D.
      • Federico A.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) as a model of small vessel disease: update on clinical, diagnostic, and management aspects.
      ,
      • Attems J.
      • Jellinger K.
      • Thal D.R.
      • Van Nostrand W.
      Review: sporadic cerebral amyloid angiopathy.
      At the histopathologic level, CADASIL and CAA both harbor abnormal accumulation of proteins within the vessel wall, although in CADASIL the major protein is NOTCH3 ectodomain and in CAA the major protein is Aβ (Figure 1A).
      • Joutel A.
      • Andreux F.
      • Gaulis S.
      • Domenga V.
      • Cécillon M.
      • Battail N.
      • Piga N.
      • Chapon F.
      • Godfrain C.
      • Tournier-Lasserve E.
      The ectodomain of the Notch3 receptor accumulates within the cerebrovasculature of CADASIL patients.
      ,
      • Glenner G.G.
      • Wong C.W.
      Alzheimer's disease: initial report of the purification and characterization of a novel cerebrovascular amyloid protein.
      Furthermore, both diseases feature degeneration of the vascular smooth muscle cells with accompanying hyalinization of vessels (Figure 1A).
      • Di Donato I.
      • Bianchi S.
      • De Stefano N.
      • Dichgans M.
      • Dotti M.T.
      • Duering M.
      • Jouvent E.
      • Korczyn A.D.
      • Lesnik Oberstein S.A.J.
      • Malandrini A.
      • Markus H.S.
      • Pantoni L.
      • Penco S.
      • Rufa A.
      • Sinanović O.
      • Stojanov D.
      • Federico A.
      Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) as a model of small vessel disease: update on clinical, diagnostic, and management aspects.
      ,
      • Attems J.
      • Jellinger K.
      • Thal D.R.
      • Van Nostrand W.
      Review: sporadic cerebral amyloid angiopathy.
      Finally, many characteristics of CADASIL at the molecular level are highly reminiscent of known amyloid pathology. For example, as discussed above, abnormal protein cleavage, processing, and oligomerization have been identified in both CADASIL and CAA. Both disorders are also thought to recruit abnormal accumulation of additional proteins other than NOTCH3 and Aβ. Given the numerous common qualities, shared molecular mechanisms may be involved in CADASIL and CAA. Thus, published work was reviewed to compare and contrast the protein profiles of the two cerebrovascular diseases.

      Overlap of CADASIL and CAA Protein Profiles

      A recent review of CADASIL and CAA proteomics-based studies by Haffner
      • Haffner C.
      Proteostasis in cerebral small vessel disease.
      identified six proteins that demonstrated increased abundance in both diseases compared with controls: SAP, TIMP3, VTN, APOE, clusterin, and HTRA1.
      • Haffner C.
      Proteostasis in cerebral small vessel disease.
      To expand on this, both proteomics-based studies and immuno-based studies of human CADASIL and CAA tissue were examined. In addition, CAA studies that did not include patients with underlying AD pathology were specifically targeted to compare CADASIL and CAA as primarily vascular disorders. The criteria for inclusion of studies in this review can be found in Table 1.
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Kast J.
      • Hanecker P.
      • Beaufort N.
      • Giese A.
      • Joutel A.
      • Dichgans M.
      • Opherk C.
      • Haffner C.
      Sequestration of latent TGF-β binding protein 1 into CADASIL-related Notch3-ECD deposits.
      • Nagatoshi A.
      • Ueda M.
      • Ueda A.
      • Tasaki M.
      • Inoue Y.
      • Ma Y.
      • Masuda T.
      • Mizukami M.
      • Matsumoto S.
      • Kosaka T.
      • Kawano T.
      • Ito T.
      • Ando Y.
      Serum amyloid P component: a novel potential player in vessel degeneration in CADASIL.
      • Young K.Z.
      • Lee S.J.
      • Zhang X.
      • Cartee N.M.P.
      • Torres M.
      • Keep S.G.
      • Gabbireddy S.R.
      • Fontana J.L.
      • Qi L.
      • Wang M.M.
      NOTCH3 is non-enzymatically fragmented in inherited cerebral small-vessel disease.
      ,
      • Joutel A.
      • Andreux F.
      • Gaulis S.
      • Domenga V.
      • Cécillon M.
      • Battail N.
      • Piga N.
      • Chapon F.
      • Godfrain C.
      • Tournier-Lasserve E.
      The ectodomain of the Notch3 receptor accumulates within the cerebrovasculature of CADASIL patients.
      ,
      • Arboleda-Velasquez J.F.
      • Manent J.
      • Lee J.H.
      • Tikka S.
      • Ospina C.
      • Vanderburg C.R.
      • Frosch M.P.
      • Rodríguez-Falcón M.
      • Villen J.
      • Gygi S.
      • Lopera F.
      • Kalimo H.
      • Moskowitz M.A.
      • Ayata C.
      • Louvi A.
      • Artavanis-Tsakonas S.
      Hypomorphic Notch 3 alleles link Notch signaling to ischemic cerebral small-vessel disease.
      ,
      • Zhang X.
      • Lee S.J.
      • Young K.Z.
      • Josephson D.A.
      • Geschwind M.D.
      • Wang M.M.
      Latent NOTCH3 epitopes unmasked in CADASIL and regulated by protein redox state.
      ,
      • Lee S.J.
      • Zhang X.
      • Wang M.M.
      Vascular accumulation of the small leucine-rich proteoglycan decorin in CADASIL.
      ,
      • Zhang X.
      • Lee S.J.
      • Young M.F.
      • Wang M.M.
      The small leucine-rich proteoglycan BGN accumulates in CADASIL and binds to NOTCH3.
      ,
      • Manousopoulou A.
      • Gatherer M.
      • Smith C.
      • Nicoll J.A.R.
      • Woelk C.H.
      • Johnson M.
      • Kalaria R.
      • Attems J.
      • Garbis S.D.
      • Carare R.O.
      Systems proteomic analysis reveals that clusterin and tissue inhibitor of metalloproteinases 3 increase in leptomeningeal arteries affected by cerebral amyloid angiopathy.
      ,
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Bell R.D.
      • Zlokovic B.V.
      Neurovascular mechanisms and blood-brain barrier disorder in Alzheimer's disease.
      ,
      • Magaki S.
      • Tang Z.
      • Tung S.
      • Williams C.K.
      • Lo D.
      • Yong W.H.
      • Khanlou N.
      • Vinters H.V.
      The effects of cerebral amyloid angiopathy on integrity of the blood-brain barrier.
      • Carrano A.
      • Hoozemans J.J.M.
      • van der Vies S.M.
      • Rozemuller A.J.M.
      • van Horssen J.
      • de Vries H.E.
      Amyloid beta induces oxidative stress-mediated blood-brain barrier changes in capillary amyloid angiopathy.
      • Carrano A.
      • Hoozemans J.J.M.
      • van der Vies S.M.
      • van Horssen J.
      • de Vries H.E.
      • Rozemuller A.J.M.
      Neuroinflammation and blood-brain barrier changes in capillary amyloid angiopathy.
      ,
      • de Jager M.
      • van der Wildt B.
      • Schul E.
      • Bol J.G.J.M.
      • van Duinen S.G.
      • Drukarch B.
      • Wilhelmus M.M.M.
      Tissue transglutaminase colocalizes with extracellular matrix proteins in cerebral amyloid angiopathy.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      ,
      • Grand Moursel L.
      • Munting L.P.
      • van der Graaf L.M.
      • van Duinen S.G.
      • Goumans M.-J.T.H.
      • Ueberham U.
      • Natté R.
      • van Buchem M.A.
      • van Roon-Mom W.M.C.
      • van der Weerd L.
      TGFβ pathway deregulation and abnormal phospho-SMAD2/3 staining in hereditary cerebral hemorrhage with amyloidosis-Dutch type.
      ,
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      ,
      • Lepelletier F.-X.
      • Mann D.M.A.
      • Robinson A.C.
      • Pinteaux E.
      • Boutin H.
      Early changes in extracellular matrix in Alzheimer's disease.
      • Jäkel L.
      • Kuiperij H.B.
      • Gerding L.P.
      • Custers E.E.M.
      • van den Berg E.
      • Jolink W.M.T.
      • Schreuder F.H.B.M.
      • Küsters B.
      • Klijn C.J.M.
      • Verbeek M.M.
      Disturbed balance in the expression of MMP9 and TIMP3 in cerebral amyloid angiopathy-related intracerebral haemorrhage.
      • Hernandez-Guillamon M.
      • Martinez-Saez E.
      • Delgado P.
      • Domingues-Montanari S.
      • Boada C.
      • Penalba A.
      • Boada M.
      • Pagola J.
      • Maisterra O.
      • Rodriguez-Luna D.
      • Molina C.A.
      • Rovira A.
      • Alvarez-Sabin J.
      • Ortega-Aznar A.
      • Montaner J.
      MMP-2/MMP-9 plasma level and brain expression in cerebral amyloid angiopathy-associated hemorrhagic stroke.
      ,
      • Dong H.
      • Ding H.
      • Young K.
      • Blaivas M.
      • Christensen P.J.
      • Wang M.M.
      Advanced intimal hyperplasia without luminal narrowing of leptomeningeal arteries in CADASIL.
      ,
      • Chen C.-H.
      • Cheng Y.-W.
      • Chen Y.-F.
      • Tang S.-C.
      • Jeng J.-S.
      Plasma neurofilament light chain and glial fibrillary acidic protein predict stroke in CADASIL.
      • Zhang X.
      • Meng H.
      • Blaivas M.
      • Rushing E.J.
      • Moore B.E.
      • Schwartz J.
      • Lopes M.B.S.
      • Worrall B.B.
      • Wang M.M.
      von Willebrand factor permeates small vessels in CADASIL and inhibits smooth muscle gene expression.
      • van Horssen J.
      • Otte-Höller I.
      • David G.
      • Maat-Schieman M.L.
      • van den Heuvel L.P.
      • Wesseling P.
      • De Waal R.M.
      • Verbeek M.M.
      Heparan sulfate proteoglycan expression in cerebrovascular amyloid beta deposits in Alzheimer's disease and hereditary cerebral hemorrhage with amyloidosis (Dutch) brains.
      • McCarron M.O.
      • Nicoll J.A.
      • Stewart J.
      • Ironside J.W.
      • Mann D.M.
      • Love S.
      • Graham D.I.
      • Dewar D.
      The apolipoprotein E epsilon2 allele and the pathological features in cerebral amyloid angiopathy-related hemorrhage.
      • Tagliavini F.
      • Ghiso J.
      • Timmers W.F.
      • Giaccone G.
      • Bugiani O.
      • Frangione B.
      Coexistence of Alzheimer's amyloid precursor protein and amyloid protein in cerebral vessel walls.
      • Inoue Y.
      • Ueda M.
      • Tasaki M.
      • Takeshima A.
      • Nagatoshi A.
      • Masuda T.
      • Misumi Y.
      • Kosaka T.
      • Nomura T.
      • Mizukami M.
      • Matsumoto S.
      • Yamashita T.
      • Takahashi H.
      • Kakita A.
      • Ando Y.
      Sushi repeat-containing protein 1: a novel disease-associated molecule in cerebral amyloid angiopathy.
      • Merlini M.
      • Wanner D.
      • Nitsch R.M.
      Tau pathology-dependent remodelling of cerebral arteries precedes Alzheimer's disease-related microvascular cerebral amyloid angiopathy.
      • Keable A.
      • Fenna K.
      • Yuen H.M.
      • Johnston D.A.
      • Smyth N.R.
      • Smith C.
      • Al-Shahi Salman R.
      • Samarasekera N.
      • Nicoll J.A.R.
      • Attems J.
      • Kalaria R.N.
      • Weller R.O.
      • Carare R.O.
      Deposition of amyloid β in the walls of human leptomeningeal arteries in relation to perivascular drainage pathways in cerebral amyloid angiopathy.
      • Xiong H.
      • Callaghan D.
      • Jones A.
      • Bai J.
      • Rasquinha I.
      • Smith C.
      • Pei K.
      • Walker D.
      • Lue L.-F.
      • Stanimirovic D.
      • Zhang W.
      ABCG2 is upregulated in Alzheimer's brain with cerebral amyloid angiopathy and may act as a gatekeeper at the blood-brain barrier for Abeta(1-40) peptides.
      • Stopa E.G.
      • Butala P.
      • Salloway S.
      • Johanson C.E.
      • Gonzalez L.
      • Tavares R.
      • Hovanesian V.
      • Hulette C.M.
      • Vitek M.P.
      • Cohen R.A.
      Cerebral cortical arteriolar angiopathy, vascular beta-amyloid, smooth muscle actin, Braak stage, and APOE genotype.
      • Miners J.S.
      • Ashby E.
      • Van Helmond Z.
      • Chalmers K.A.
      • Palmer L.E.
      • Love S.
      • Kehoe P.G.
      Angiotensin-converting enzyme (ACE) levels and activity in Alzheimer's disease, and relationship of perivascular ACE-1 to cerebral amyloid angiopathy.
      • Tian J.
      • Shi J.
      • Smallman R.
      • Iwatsubo T.
      • Mann D.M.A.
      Relationships in Alzheimer's disease between the extent of Abeta deposition in cerebral blood vessel walls, as cerebral amyloid angiopathy, and the amount of cerebrovascular smooth muscle cells and collagen.
      • Ervin J.F.
      • Pannell C.
      • Szymanski M.
      • Welsh-Bohmer K.
      • Schmechel D.E.
      • Hulette C.M.
      Vascular smooth muscle actin is reduced in Alzheimer disease brain: a quantitative analysis.
      • Zhang W.W.
      • Lempessi H.
      • Olsson Y.
      Amyloid angiopathy of the human brain: immunohistochemical studies using markers for components of extracellular matrix, smooth muscle actin and endothelial cells.
      • Verbeek M.M.
      • Otte-Höller I.
      • Ruiter D.J.
      • De Waal R.M.
      Distribution of Aβ-associated proteins in cerebrovascular amyloid of Alzheimer's disease.
      • Kalaria R.N.
      Cerebral vessels in ageing and Alzheimer's disease.
      • Powers J.M.
      • Schlaepfer W.W.
      • Willingham M.C.
      • Hall B.J.
      An immunoperoxidase study of senile cerebral amyloidosis with pathogenetic considerations.
      • Ishii T.
      • Haga S.
      Identification of components of immunoglobulins in senile plaques by means of fluorescent antibody technique.
      Limitations of including immuno-based studies include the lack of specificity for some protein subtypes. For example, numerous immunohistochemical studies have observed enrichment of various collagens in both CADASIL and CAA. However, unlike with proteomics-based approaches, immunohistochemical studies often cannot discern the specific collagen subtype (eg, COL1A1 versus COL1A2). In these cases, only the more specific collagen subtypes from liquid chromatography–tandem mass spectrometry studies were included, when available. When unavailable, the general protein (eg, COL4) was included. In later gene expression analyses, all subtypes of the protein were included. Occasionally, studies indicated opposite directions of change for some proteins (eg, glial fibrillary acidic protein and basement membrane–specific heparan sulfate proteoglycan core protein). These cases were noted, and the proteins were graded in the direction supported by most studies (≥50%).
      Table 1Inclusion Criteria for CADASIL, Vascular CAA, and CAA with Underlying AD Pathology Studies
      CADASIL studiesMethodTissueBrain regionSample sizeInclusion criteria
      Chen et al
      • Chen C.-H.
      • Cheng Y.-W.
      • Chen Y.-F.
      • Tang S.-C.
      • Jeng J.-S.
      Plasma neurofilament light chain and glial fibrillary acidic protein predict stroke in CADASIL.
      Plasma biomarkerPlasmaNA63 CADASIL, 17 controlP < 0.05
      Young et al
      • Young K.Z.
      • Lee S.J.
      • Zhang X.
      • Cartee N.M.P.
      • Torres M.
      • Keep S.G.
      • Gabbireddy S.R.
      • Fontana J.L.
      • Qi L.
      • Wang M.M.
      NOTCH3 is non-enzymatically fragmented in inherited cerebral small-vessel disease.
      IHCLeptomeningeal arteries, small penetrating arteries of white matterFrontal lobe19 CADASIL, 10 controlQualitative
      Zellner et al
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      LC-MS/MSIsolated cerebral vesselsFrontal lobe6 CADASIL, 6 controlP < 0.05
      Nagatoshi et al
      • Nagatoshi A.
      • Ueda M.
      • Ueda A.
      • Tasaki M.
      • Inoue Y.
      • Ma Y.
      • Masuda T.
      • Mizukami M.
      • Matsumoto S.
      • Kosaka T.
      • Kawano T.
      • Ito T.
      • Ando Y.
      Serum amyloid P component: a novel potential player in vessel degeneration in CADASIL.
      LC-MS/MSIsolated leptomeningeal arteries/arterioles or superficial temporal arteryNA3 CADASIL (2 autopsy, 1 biopsy), 6 control (5 autopsy, 1 biopsy)Top 25 changed proteins (up/down) in nondetected/detected and detected/detected conditions
      Zhang et al
      • Zhang X.
      • Lee S.J.
      • Young M.F.
      • Wang M.M.
      The small leucine-rich proteoglycan BGN accumulates in CADASIL and binds to NOTCH3.
      WB, IHCLeptomeningeal arteries, small penetrating arteries of white matterFrontal lobe8 CADASIL, 6 controlP < 0.05
      Zhang et al
      • Zhang X.
      • Lee S.J.
      • Young K.Z.
      • Josephson D.A.
      • Geschwind M.D.
      • Wang M.M.
      Latent NOTCH3 epitopes unmasked in CADASIL and regulated by protein redox state.
      IHCLeptomeningeal arteries, small penetrating arteries of white matterFrontal lobe8 CADASIL, 6 controlQualitative
      Lee et al
      • Lee S.J.
      • Zhang X.
      • Wang M.M.
      Vascular accumulation of the small leucine-rich proteoglycan decorin in CADASIL.
      WB, IHCLeptomeningeal arteries, small penetrating arteries of white matter, and capillariesAnterior temporal lobe6 CADASIL, 6 controlP < 0.05
      Kast et al
      • Kast J.
      • Hanecker P.
      • Beaufort N.
      • Giese A.
      • Joutel A.
      • Dichgans M.
      • Opherk C.
      • Haffner C.
      Sequestration of latent TGF-β binding protein 1 into CADASIL-related Notch3-ECD deposits.
      WB, IHCArteriolesFrontal subcortex5 CADASIL, 4 controlQualitative
      Monet-Leprêtre et al
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      Nano–LC-MS/MSIsolated microvesselsFrontal or occipital lobe7 CADASIL, 9 control>6 Peptide changes from control
      Dong et al
      • Dong H.
      • Blaivas M.
      • Wang M.M.
      Bidirectional encroachment of collagen into the tunica media in cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy.
      IHCLeptomeningeal arteries, small penetrating arteries of white matter, and capillariesAnterior temporal lobe6 CADASIL, 6 controlP < 0.05
      Zhang et al
      • Zhang X.
      • Meng H.
      • Blaivas M.
      • Rushing E.J.
      • Moore B.E.
      • Schwartz J.
      • Lopes M.B.S.
      • Worrall B.B.
      • Wang M.M.
      von Willebrand factor permeates small vessels in CADASIL and inhibits smooth muscle gene expression.
      IHCLeptomeningeal arteries, small penetrating arteries of white matter, and capillariesAnterior temporal lobe6 CADASIL, 25 controlQualitative
      Arboleda-Velasquez et al
      • Arboleda-Velasquez J.F.
      • Manent J.
      • Lee J.H.
      • Tikka S.
      • Ospina C.
      • Vanderburg C.R.
      • Frosch M.P.
      • Rodríguez-Falcón M.
      • Villen J.
      • Gygi S.
      • Lopera F.
      • Kalimo H.
      • Moskowitz M.A.
      • Ayata C.
      • Louvi A.
      • Artavanis-Tsakonas S.
      Hypomorphic Notch 3 alleles link Notch signaling to ischemic cerebral small-vessel disease.
      LC-MS/MSArteriolesSubcortical white matter2 CADASIL (R1031C mutation), 2 controlAbsolute value of the difference between the average peptide counts (ie, PSM) in the experimental sample minus the average PSM in the control sample exceeded three times the variation and a value of 4
      Joutel et al
      • Joutel A.
      • Andreux F.
      • Gaulis S.
      • Domenga V.
      • Cécillon M.
      • Battail N.
      • Piga N.
      • Chapon F.
      • Godfrain C.
      • Tournier-Lasserve E.
      The ectodomain of the Notch3 receptor accumulates within the cerebrovasculature of CADASIL patients.
      WB, IHCCerebral arteries, veins, and capillariesFrontal lobe9 CADASIL, 13 controlQualitative
      CAA studiesMethodTissueBrain regionSample sizeInclusion criteria
      Endo et al
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      LC-MS/MSLaser capture microdissected tissue; leptomeningeal and cortical vesselsCortical tissue6 CAA, 5 control≥50% Detection in CAA patients
      Hondius et al
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      LC-MS/MSLaser capture microdissected tissueCortical layers II to VI7 AD, 7 CAA, 6 controlP < 0.05
      Grand Moursel et al
      • Grand Moursel L.
      • Munting L.P.
      • van der Graaf L.M.
      • van Duinen S.G.
      • Goumans M.-J.T.H.
      • Ueberham U.
      • Natté R.
      • van Buchem M.A.
      • van Roon-Mom W.M.C.
      • van der Weerd L.
      TGFβ pathway deregulation and abnormal phospho-SMAD2/3 staining in hereditary cerebral hemorrhage with amyloidosis-Dutch type.
      IHC, qPCRParenchymal angiopathic arteriolesFrontal and occipital cortex11 HCHWA-D, 11 sCAA, 11 controlP < 0.05
      de Jager et al
      • de Jager M.
      • van der Wildt B.
      • Schul E.
      • Bol J.G.J.M.
      • van Duinen S.G.
      • Drukarch B.
      • Wilhelmus M.M.M.
      Tissue transglutaminase colocalizes with extracellular matrix proteins in cerebral amyloid angiopathy.
      IHCLeptomeningeal and parenchymal vesselsNeocortex5 AD, 2 CAA, 5 HCHWA-D, 7 controlQualitative
      Carrano et al
      • Carrano A.
      • Hoozemans J.J.M.
      • van der Vies S.M.
      • van Horssen J.
      • de Vries H.E.
      • Rozemuller A.J.M.
      Neuroinflammation and blood-brain barrier changes in capillary amyloid angiopathy.
      IHCNormal and Ab-laden capillariesOccipital cortex23 CAAP < 0.05
      van Horssen et al
      • van Horssen J.
      • Otte-Höller I.
      • David G.
      • Maat-Schieman M.L.
      • van den Heuvel L.P.
      • Wesseling P.
      • De Waal R.M.
      • Verbeek M.M.
      Heparan sulfate proteoglycan expression in cerebrovascular amyloid beta deposits in Alzheimer's disease and hereditary cerebral hemorrhage with amyloidosis (Dutch) brains.
      IHCLeptomeningeal and cortical vesselsFrontal cortex7 AD, 4 HCHWA-D, 3 controlQualitative
      McCarron et al
      • McCarron M.O.
      • Nicoll J.A.
      • Stewart J.
      • Ironside J.W.
      • Mann D.M.
      • Love S.
      • Graham D.I.
      • Dewar D.
      The apolipoprotein E epsilon2 allele and the pathological features in cerebral amyloid angiopathy-related hemorrhage.
      IHCLeptomeningeal and cortical vesselsFrontal, parietal, occipital, temporal or GFM26 AD, 37 CAA, 20 controlP < 0.05
      Tagliavini et al
      • Tagliavini F.
      • Ghiso J.
      • Timmers W.F.
      • Giaccone G.
      • Bugiani O.
      • Frangione B.
      Coexistence of Alzheimer's amyloid precursor protein and amyloid protein in cerebral vessel walls.
      WB, IHCIsolated leptomeningeal and cortical microvesselsCortical tissue3 AD, 2 HCHWA-D, 2 controlQualitative
      CAA studies with underlying AD pathologyMethodTissueBrain regionSample sizeInclusion criteria
      Jäkel et al
      • Jäkel L.
      • Kuiperij H.B.
      • Gerding L.P.
      • Custers E.E.M.
      • van den Berg E.
      • Jolink W.M.T.
      • Schreuder F.H.B.M.
      • Küsters B.
      • Klijn C.J.M.
      • Verbeek M.M.
      Disturbed balance in the expression of MMP9 and TIMP3 in cerebral amyloid angiopathy-related intracerebral haemorrhage.
      IHCLeptomeningeal and cortical vesselsOccipital cortex18 CAA (nonhemorrhagic), 11 CAA (hemorrhagic), 11 controlP < 0.05
      Magaki et al
      • Magaki S.
      • Tang Z.
      • Tung S.
      • Williams C.K.
      • Lo D.
      • Yong W.H.
      • Khanlou N.
      • Vinters H.V.
      The effects of cerebral amyloid angiopathy on integrity of the blood-brain barrier.
      IHCGray matter and white matterFrontal, temporal, parietal, and occipital cortices, hippocampus, entorhinal cortex and amygdala, basal ganglia, brainstem, and cerebellum7 AD, 8 type 1 CAA, 10 type 2 CAA, 10 controlP < 0.05
      Inoue et al
      • Inoue Y.
      • Ueda M.
      • Tasaki M.
      • Takeshima A.
      • Nagatoshi A.
      • Masuda T.
      • Misumi Y.
      • Kosaka T.
      • Nomura T.
      • Mizukami M.
      • Matsumoto S.
      • Yamashita T.
      • Takahashi H.
      • Kakita A.
      • Ando Y.
      Sushi repeat-containing protein 1: a novel disease-associated molecule in cerebral amyloid angiopathy.
      LC-MS/MSCerebral neocortical tissueNeocortical tissue8 Severe CAA, 12 mild CAA, and 10 controlP < 0.05
      Lepelletier et al
      • Lepelletier F.-X.
      • Mann D.M.A.
      • Robinson A.C.
      • Pinteaux E.
      • Boutin H.
      Early changes in extracellular matrix in Alzheimer's disease.
      IHCCortical gray matterSuperior frontal gyrus and inferior temporal gyrus17 AD (10 subclinical, 8 clinical), 1 CAA, 12 controlP < 0.05
      Merlini et al
      • Merlini M.
      • Wanner D.
      • Nitsch R.M.
      Tau pathology-dependent remodelling of cerebral arteries precedes Alzheimer's disease-related microvascular cerebral amyloid angiopathy.
      IHCLeptomeningeal arterioles, small arteries, medium arteriesGFM and hippocampus17 AD, 28 controlP < 0.05
      Keable et al
      • Keable A.
      • Fenna K.
      • Yuen H.M.
      • Johnston D.A.
      • Smyth N.R.
      • Smith C.
      • Al-Shahi Salman R.
      • Samarasekera N.
      • Nicoll J.A.R.
      • Attems J.
      • Kalaria R.N.
      • Weller R.O.
      • Carare R.O.
      Deposition of amyloid β in the walls of human leptomeningeal arteries in relation to perivascular drainage pathways in cerebral amyloid angiopathy.
      IFLeptomeningeal and intraparenchymal vesselsOccipital cortex20 Severe CAA, 14 young controls, 20 aged controlsP < 0.05
      Manousopoulou et al
      • Manousopoulou A.
      • Gatherer M.
      • Smith C.
      • Nicoll J.A.R.
      • Woelk C.H.
      • Johnson M.
      • Kalaria R.
      • Attems J.
      • Garbis S.D.
      • Carare R.O.
      Systems proteomic analysis reveals that clusterin and tissue inhibitor of metalloproteinases 3 increase in leptomeningeal arteries affected by cerebral amyloid angiopathy.
      LC-MSIsolated leptomeningeal arteriesOccipital cortex4 CAA, 2 old control (females only)>Average ± 2.4 log2 ratio
      Hernandez-Guillamon et al
      • Hernandez-Guillamon M.
      • Martinez-Saez E.
      • Delgado P.
      • Domingues-Montanari S.
      • Boada C.
      • Penalba A.
      • Boada M.
      • Pagola J.
      • Maisterra O.
      • Rodriguez-Luna D.
      • Molina C.A.
      • Rovira A.
      • Alvarez-Sabin J.
      • Ortega-Aznar A.
      • Montaner J.
      MMP-2/MMP-9 plasma level and brain expression in cerebral amyloid angiopathy-associated hemorrhagic stroke.
      WB, IHC, ELISAHemorrhagic stroke areaPerihematoma and contralateral area4 CAA, 3 controlP < 0.05
      Carrano et al
      • Carrano A.
      • Hoozemans J.J.M.
      • van der Vies S.M.
      • Rozemuller A.J.M.
      • van Horssen J.
      • de Vries H.E.
      Amyloid beta induces oxidative stress-mediated blood-brain barrier changes in capillary amyloid angiopathy.
      IHCNormal and Ab-laden capillariesOccipital cortex6 CAA, 2 controlP < 0.05
      Xiong et al
      • Xiong H.
      • Callaghan D.
      • Jones A.
      • Bai J.
      • Rasquinha I.
      • Smith C.
      • Pei K.
      • Walker D.
      • Lue L.-F.
      • Stanimirovic D.
      • Zhang W.
      ABCG2 is upregulated in Alzheimer's brain with cerebral amyloid angiopathy and may act as a gatekeeper at the blood-brain barrier for Abeta(1-40) peptides.
      WB, IHC, qPCRCerebral vasculatureOccipital cortex13 AD, 13 CAA/AD, 12 normalP < 0.05
      Bell and Zlokovic
      • Bell R.D.
      • Zlokovic B.V.
      Neurovascular mechanisms and blood-brain barrier disorder in Alzheimer's disease.
      WBLeptomeningeal vesselsBrodmann areas 9/103 AD, 3 controlP < 0.05
      Stopa et al
      • Stopa E.G.
      • Butala P.
      • Salloway S.
      • Johanson C.E.
      • Gonzalez L.
      • Tavares R.
      • Hovanesian V.
      • Hulette C.M.
      • Vitek M.P.
      • Cohen R.A.
      Cerebral cortical arteriolar angiopathy, vascular beta-amyloid, smooth muscle actin, Braak stage, and APOE genotype.
      IHCArteriolesFrontal cortex76 AD, 19 controlP < 0.05
      Miners et al
      • Miners J.S.
      • Ashby E.
      • Van Helmond Z.
      • Chalmers K.A.
      • Palmer L.E.
      • Love S.
      • Kehoe P.G.
      Angiotensin-converting enzyme (ACE) levels and activity in Alzheimer's disease, and relationship of perivascular ACE-1 to cerebral amyloid angiopathy.
      IHCCerebral vasculatureFrontal cortex5 AD, 5 AD with CAA, 5 controlP < 0.05
      Tian et al
      • Tian J.
      • Shi J.
      • Smallman R.
      • Iwatsubo T.
      • Mann D.M.A.
      Relationships in Alzheimer's disease between the extent of Abeta deposition in cerebral blood vessel walls, as cerebral amyloid angiopathy, and the amount of cerebrovascular smooth muscle cells and collagen.
      IHCLeptomeningeal arteries and intraparenchymal blood vesselsFrontal cortex70 Late-stage AD patientsP < 0.05
      Ervin et al
      • Ervin J.F.
      • Pannell C.
      • Szymanski M.
      • Welsh-Bohmer K.
      • Schmechel D.E.
      • Hulette C.M.
      Vascular smooth muscle actin is reduced in Alzheimer disease brain: a quantitative analysis.
      IHCArteriesFrontal cortex10 AD (APOE4), 10 AD (APOE3), 10 controlP < 0.05
      Zhang et al
      • Zhang W.W.
      • Lempessi H.
      • Olsson Y.
      Amyloid angiopathy of the human brain: immunohistochemical studies using markers for components of extracellular matrix, smooth muscle actin and endothelial cells.
      IHCLeptomeningeal and cortical vesselsCerebral cortex10 CAA (5 AD, 1 Down syndrome, 4 unknown CAA), 8 controlQualitative
      Verbeek et al
      • Verbeek M.M.
      • Otte-Höller I.
      • Ruiter D.J.
      • De Waal R.M.
      Distribution of Aβ-associated proteins in cerebrovascular amyloid of Alzheimer's disease.
      IHCLeptomeningeal and cortical vesselsFrontal, parietal, temporal, and occipital cortex11 AD, 1 FTD, 11 controlQualitative
      Kalaria
      • Kalaria R.N.
      Cerebral vessels in ageing and Alzheimer's disease.
      WB, IHCCerebral microvessels, large meningeal vessels, and choroidal samplesFrontal, occipital, temporal lobes and cerebellar cortex22 AD, 8 controlP < 0.05
      Powers et al
      • Powers J.M.
      • Schlaepfer W.W.
      • Willingham M.C.
      • Hall B.J.
      An immunoperoxidase study of senile cerebral amyloidosis with pathogenetic considerations.
      IHCArteries and plaqueCerebral cortex2 Presenile AD, 5 senile AD, 5 controlQualitative
      Ishii and Haga
      • Ishii T.
      • Haga S.
      Identification of components of immunoglobulins in senile plaques by means of fluorescent antibody technique.
      IHCVessels and plaquesFrontal cortex2 AD, 2 controlQualitative
      A total of 13 CADASIL studies, 8 vascular CAA studies, and 20 CAA with underlying AD pathology studies were examined to identify differentially regulated proteins in disease. Detection methods included LC-MS/MS, IHC, and WB. Sample types include isolated vessels and/or brain tissue. When applicable, the brain region is specified. Sample sizes are noted for each study. When classifying studies examining vascular CAA, patients with known AD or cognitive changes indicative of AD were excluded. From each of these studies, a protein was denoted as differentially regulated if it was significantly changed in disease compared with control (P < 0.05), qualitatively changed, as in the case of IHC, or otherwise specified if no statistical analysis was available.
      Ab, antibody; AD, Alzheimer disease; APOE, apolipoprotein E; CAA, cerebral amyloid angiopathy; CADASIL, cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy; ELISA, enzyme-linked immunosorbent assay; FTD, frontotemporal dementia; GFM, gyrus frontalis medialis; HCHWA-D, hereditary cerebral hemorrhage with amyloidosis, Dutch type; IF, immunofluorescence; IHC, immunohistochemistry; LC-MS, liquid chromatography–mass spectrometry; LC-MS/MS, liquid chromatography–tandem mass spectrometry; NA, not applicable; PSM, peptide spectrum matches; qPCR, quantitative PCR; WB, Western blot analysis.
      Analysis of existing literature identified significant overlap of proteins differentially regulated in CADASIL and CAA compared with controls. The vast majority of these proteins come from liquid chromatography–tandem mass spectrometry studies. The advantages of proteomics-based studies include the ability to assess global protein content from disease tissue.
      • Haffner C.
      • Malik R.
      • Dichgans M.
      Genetic factors in cerebral small vessel disease and their impact on stroke and dementia.
      In CAA without underlying AD pathology, 58 proteins from eight studies were found to be enhanced or decreased. A comparison between proteins differentially regulated in CADASIL and CAA studies highlighted 19 shared proteins changed in the same direction, which made up roughly 33% of the enhanced or decreased CAA proteins (Figure 1B and Table 2
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Kast J.
      • Hanecker P.
      • Beaufort N.
      • Giese A.
      • Joutel A.
      • Dichgans M.
      • Opherk C.
      • Haffner C.
      Sequestration of latent TGF-β binding protein 1 into CADASIL-related Notch3-ECD deposits.
      ,
      • Nagatoshi A.
      • Ueda M.
      • Ueda A.
      • Tasaki M.
      • Inoue Y.
      • Ma Y.
      • Masuda T.
      • Mizukami M.
      • Matsumoto S.
      • Kosaka T.
      • Kawano T.
      • Ito T.
      • Ando Y.
      Serum amyloid P component: a novel potential player in vessel degeneration in CADASIL.
      ,
      • Arboleda-Velasquez J.F.
      • Manent J.
      • Lee J.H.
      • Tikka S.
      • Ospina C.
      • Vanderburg C.R.
      • Frosch M.P.
      • Rodríguez-Falcón M.
      • Villen J.
      • Gygi S.
      • Lopera F.
      • Kalimo H.
      • Moskowitz M.A.
      • Ayata C.
      • Louvi A.
      • Artavanis-Tsakonas S.
      Hypomorphic Notch 3 alleles link Notch signaling to ischemic cerebral small-vessel disease.
      ,
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • de Jager M.
      • van der Wildt B.
      • Schul E.
      • Bol J.G.J.M.
      • van Duinen S.G.
      • Drukarch B.
      • Wilhelmus M.M.M.
      Tissue transglutaminase colocalizes with extracellular matrix proteins in cerebral amyloid angiopathy.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      ,
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      ,
      • Chen C.-H.
      • Cheng Y.-W.
      • Chen Y.-F.
      • Tang S.-C.
      • Jeng J.-S.
      Plasma neurofilament light chain and glial fibrillary acidic protein predict stroke in CADASIL.
      ,
      • McCarron M.O.
      • Nicoll J.A.
      • Stewart J.
      • Ironside J.W.
      • Mann D.M.
      • Love S.
      • Graham D.I.
      • Dewar D.
      The apolipoprotein E epsilon2 allele and the pathological features in cerebral amyloid angiopathy-related hemorrhage.
      ,
      • Cifuentes D.
      • Poittevin M.
      • Dere E.
      • Broquères-You D.
      • Bonnin P.
      • Benessiano J.
      • Pocard M.
      • Mariani J.
      • Kubis N.
      • Merkulova-Rainon T.
      • Lévy B.I.
      Hypertension accelerates the progression of Alzheimer-like pathology in a mouse model of the disease.
      ). Functional STRING version 11 (https://string-db.org/) analysis demonstrated that 18 of the 19 shared proteins are known or predicted to interact with each other, with an average per protein 5.6 predicted and known interactions with other shared proteins listed in Table 2 (Figure 1C).
      • Szklarczyk D.
      • Gable A.L.
      • Lyon D.
      • Junge A.
      • Wyder S.
      • Huerta-Cepas J.
      • Simonovic M.
      • Doncheva N.T.
      • Morris J.H.
      • Bork P.
      • Jensen L.J.
      • Mering C.V.
      STRING v11: protein-protein association networks with increased coverage, supporting functional discovery in genome-wide experimental datasets.
      Only norrin did not demonstrate any known or predicted interactions with the other overlapped proteins.
      • Szklarczyk D.
      • Gable A.L.
      • Lyon D.
      • Junge A.
      • Wyder S.
      • Huerta-Cepas J.
      • Simonovic M.
      • Doncheva N.T.
      • Morris J.H.
      • Bork P.
      • Jensen L.J.
      • Mering C.V.
      STRING v11: protein-protein association networks with increased coverage, supporting functional discovery in genome-wide experimental datasets.
      Interestingly, the vast majority of the shared enriched proteins are components of the ECM.
      • Szklarczyk D.
      • Gable A.L.
      • Lyon D.
      • Junge A.
      • Wyder S.
      • Huerta-Cepas J.
      • Simonovic M.
      • Doncheva N.T.
      • Morris J.H.
      • Bork P.
      • Jensen L.J.
      • Mering C.V.
      STRING v11: protein-protein association networks with increased coverage, supporting functional discovery in genome-wide experimental datasets.
      The high degree of similarity and interconnectedness of shared proteins suggests mechanistic overlap between the two vascular angiopathies at the molecular level.
      Table 2Proteins That Are Differentially Regulated in Both CADASIL and CAA
      Regulated proteins in CADASIL and CAACoding geneSubcellular localizationSTRING

      interactions, N
      • Szklarczyk D.
      • Gable A.L.
      • Lyon D.
      • Junge A.
      • Wyder S.
      • Huerta-Cepas J.
      • Simonovic M.
      • Doncheva N.T.
      • Morris J.H.
      • Bork P.
      • Jensen L.J.
      • Mering C.V.
      STRING v11: protein-protein association networks with increased coverage, supporting functional discovery in genome-wide experimental datasets.
      Serum amyloid protein (SAP)
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Nagatoshi A.
      • Ueda M.
      • Ueda A.
      • Tasaki M.
      • Inoue Y.
      • Ma Y.
      • Masuda T.
      • Mizukami M.
      • Matsumoto S.
      • Kosaka T.
      • Kawano T.
      • Ito T.
      • Ando Y.
      Serum amyloid P component: a novel potential player in vessel degeneration in CADASIL.
      ,
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      APCSExtracellular7
      Apolipoprotein E
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      ,
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      ,
      • McCarron M.O.
      • Nicoll J.A.
      • Stewart J.
      • Ironside J.W.
      • Mann D.M.
      • Love S.
      • Graham D.I.
      • Dewar D.
      The apolipoprotein E epsilon2 allele and the pathological features in cerebral amyloid angiopathy-related hemorrhage.
      APOEExtracellular10
      Complement C4-A
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      C4AExtracellular5
      Clusterin
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Arboleda-Velasquez J.F.
      • Manent J.
      • Lee J.H.
      • Tikka S.
      • Ospina C.
      • Vanderburg C.R.
      • Frosch M.P.
      • Rodríguez-Falcón M.
      • Villen J.
      • Gygi S.
      • Lopera F.
      • Kalimo H.
      • Moskowitz M.A.
      • Ayata C.
      • Louvi A.
      • Artavanis-Tsakonas S.
      Hypomorphic Notch 3 alleles link Notch signaling to ischemic cerebral small-vessel disease.
      ,
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      ,
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      CLUExtracellular, cytoplasm, mitochondria, endoplasmic reticulum, nucleus7
      Collagen α-2(I) chain
      • Arboleda-Velasquez J.F.
      • Manent J.
      • Lee J.H.
      • Tikka S.
      • Ospina C.
      • Vanderburg C.R.
      • Frosch M.P.
      • Rodríguez-Falcón M.
      • Villen J.
      • Gygi S.
      • Lopera F.
      • Kalimo H.
      • Moskowitz M.A.
      • Ayata C.
      • Louvi A.
      • Artavanis-Tsakonas S.
      Hypomorphic Notch 3 alleles link Notch signaling to ischemic cerebral small-vessel disease.
      ,
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      COL1A2Extracellular5
      Collagen α-2(VI) chain
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Nagatoshi A.
      • Ueda M.
      • Ueda A.
      • Tasaki M.
      • Inoue Y.
      • Ma Y.
      • Masuda T.
      • Mizukami M.
      • Matsumoto S.
      • Kosaka T.
      • Kawano T.
      • Ito T.
      • Ando Y.
      Serum amyloid P component: a novel potential player in vessel degeneration in CADASIL.
      ,
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      COL6A2Extracellular5
      Collagen α-3(VI) chain
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      COL6A3Extracellular5
      Cystatin C
      • Nagatoshi A.
      • Ueda M.
      • Ueda A.
      • Tasaki M.
      • Inoue Y.
      • Ma Y.
      • Masuda T.
      • Mizukami M.
      • Matsumoto S.
      • Kosaka T.
      • Kawano T.
      • Ito T.
      • Ando Y.
      Serum amyloid P component: a novel potential player in vessel degeneration in CADASIL.
      ,
      • McCarron M.O.
      • Nicoll J.A.
      • Stewart J.
      • Ironside J.W.
      • Mann D.M.
      • Love S.
      • Graham D.I.
      • Dewar D.
      The apolipoprotein E epsilon2 allele and the pathological features in cerebral amyloid angiopathy-related hemorrhage.
      CST3Extracellular8
      Fibronectin
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Kast J.
      • Hanecker P.
      • Beaufort N.
      • Giese A.
      • Joutel A.
      • Dichgans M.
      • Opherk C.
      • Haffner C.
      Sequestration of latent TGF-β binding protein 1 into CADASIL-related Notch3-ECD deposits.
      ,
      • de Jager M.
      • van der Wildt B.
      • Schul E.
      • Bol J.G.J.M.
      • van Duinen S.G.
      • Drukarch B.
      • Wilhelmus M.M.M.
      Tissue transglutaminase colocalizes with extracellular matrix proteins in cerebral amyloid angiopathy.
      FN1Extracellular14
      Glial fibrillary acidic protein
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      ,
      • Chen C.-H.
      • Cheng Y.-W.
      • Chen Y.-F.
      • Tang S.-C.
      • Jeng J.-S.
      Plasma neurofilament light chain and glial fibrillary acidic protein predict stroke in CADASIL.
      GFAPCytoplasm4
      Basement membrane–specific heparan sulfate proteoglycan core protein
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Arboleda-Velasquez J.F.
      • Manent J.
      • Lee J.H.
      • Tikka S.
      • Ospina C.
      • Vanderburg C.R.
      • Frosch M.P.
      • Rodríguez-Falcón M.
      • Villen J.
      • Gygi S.
      • Lopera F.
      • Kalimo H.
      • Moskowitz M.A.
      • Ayata C.
      • Louvi A.
      • Artavanis-Tsakonas S.
      Hypomorphic Notch 3 alleles link Notch signaling to ischemic cerebral small-vessel disease.
      ,
      • van Horssen J.
      • Otte-Höller I.
      • David G.
      • Maat-Schieman M.L.
      • van den Heuvel L.P.
      • Wesseling P.
      • De Waal R.M.
      • Verbeek M.M.
      Heparan sulfate proteoglycan expression in cerebrovascular amyloid beta deposits in Alzheimer's disease and hereditary cerebral hemorrhage with amyloidosis (Dutch) brains.
      ,
      • Cifuentes D.
      • Poittevin M.
      • Dere E.
      • Broquères-You D.
      • Bonnin P.
      • Benessiano J.
      • Pocard M.
      • Mariani J.
      • Kubis N.
      • Merkulova-Rainon T.
      • Lévy B.I.
      Hypertension accelerates the progression of Alzheimer-like pathology in a mouse model of the disease.
      HSPG2Extracellular9
      Serine protease HTRA1
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      HTRA1Extracellular, cytosol, plasma membrane2
      Laminin subunit γ 1
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Arboleda-Velasquez J.F.
      • Manent J.
      • Lee J.H.
      • Tikka S.
      • Ospina C.
      • Vanderburg C.R.
      • Frosch M.P.
      • Rodríguez-Falcón M.
      • Villen J.
      • Gygi S.
      • Lopera F.
      • Kalimo H.
      • Moskowitz M.A.
      • Ayata C.
      • Louvi A.
      • Artavanis-Tsakonas S.
      Hypomorphic Notch 3 alleles link Notch signaling to ischemic cerebral small-vessel disease.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      LAMC1Extracellular7
      Norrin
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      NDPExtracellular0
      Neurofascin
      • Arboleda-Velasquez J.F.
      • Manent J.
      • Lee J.H.
      • Tikka S.
      • Ospina C.
      • Vanderburg C.R.
      • Frosch M.P.
      • Rodríguez-Falcón M.
      • Villen J.
      • Gygi S.
      • Lopera F.
      • Kalimo H.
      • Moskowitz M.A.
      • Ayata C.
      • Louvi A.
      • Artavanis-Tsakonas S.
      Hypomorphic Notch 3 alleles link Notch signaling to ischemic cerebral small-vessel disease.
      ,
      • Hondius D.C.
      • Eigenhuis K.N.
      • Morrema T.H.J.
      • van der Schors R.C.
      • van Nierop P.
      • Bugiani M.
      • Li K.W.
      • Hoozemans J.J.M.
      • Smit A.B.
      • Rozemuller A.J.M.
      Proteomics analysis identifies new markers associated with capillary cerebral amyloid angiopathy in Alzheimer's disease.
      NFASCPlasma membrane1
      Prostaglandin H2 D isomerase
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      PTGDSExtracellular, endoplasmic reticulum, nucleus, Golgi apparatus3
      Metalloproteinase inhibitor 3
      • Monet-Leprêtre M.
      • Haddad I.
      • Baron-Menguy C.
      • Fouillot-Panchal M.
      • Riani M.
      • Domenga-Denier V.
      • Dussaule C.
      • Cognat E.
      • Vinh J.
      • Joutel A.
      Abnormal recruitment of extracellular matrix proteins by excess Notch3 ECD: a new pathomechanism in CADASIL.
      ,
      • Zellner A.
      • Scharrer E.
      • Arzberger T.
      • Oka C.
      • Domenga-Denier V.
      • Joutel A.
      • Lichtenthaler S.F.
      • Müller S.A.
      • Dichgans M.
      • Haffner C.
      CADASIL brain vessels show a HTRA1 loss-of-function profile.
      ,
      • Endo Y.
      • Hasegawa K.
      • Nomura R.
      • Arishima H.
      • Kikuta K.-I.
      • Yamashita T.
      • Inoue Y.
      • Ueda M.
      • Ando Y.
      • Wilson M.R.
      • Hamano T.
      • Nakamoto Y.
      • Naiki H.
      Apolipoprotein E and clusterin inhibit the early phase of amyloid-β aggregation in an in vitro model of cerebral amyloid angiopathy.
      TIMP3Extracellular7
      Tropomyosin α 1 chain
      • Nagatoshi A.
      • Ueda M.
      • Ueda A.
      • Tasaki M.
      • Inoue Y.
      • Ma Y.
      • Masuda T.
      • Mizukami M.
      • Matsumoto S.
      • Kosaka T.
      • Kawano T.
      • Ito T.
      • Ando Y.
      Serum amyloid P component: a novel potential player in vessel degeneration in CADASIL.