The effects of cerebral amyloid angiopathy on integrity of the blood-brain barrier.

Cerebral amyloid angiopathy (CAA), in which amyloid accumulates predominantly in the walls of arterioles and capillaries, is seen in most patients with Alzheimer disease (AD) and may contribute to compromise of blood-brain barrier (BBB) function seen in AD. We investigated the effects of CAA on BBB integrity by examining the expression of the endothelial marker CD31, basement membrane protein collagen IV (COL4), tight junction protein claudin-5, and fibrinogen, a marker of BBB leakage, by immunohistochemistry in the occipital cortex of autopsy brains with AD and capillary CAA (CAA type 1; n = 8), AD with noncapillary CAA (CAA type 2; n = 10), and AD without CAA (n = 7) compared with elderly controls (n = 10). Given the difference in pathogenesis of capillary and noncapillary CAA, we hypothesize that features of BBB breakdown are observed only in capillary CAA. We found decreased expression of CD31 in AD subjects with CAA types 1 and 2 compared with AD without CAA and an increase in COL4 in AD without CAA compared with controls. Furthermore, there was increased immunoreactivity for fibrinogen in AD with CAA type 1 compared with controls. These findings suggest that capillary CAA is associated with morphologic and possibly physiologic alterations of the neurovascular unit and increased BBB permeability in AD.

Molecular Disorganization of Axons Adjacent to Human Cortical Microinfarcts.

Cortical microinfarcts (CMIs) are microscopically identified wedge-shaped ischemic lesions that occur at or near the cortical surface and result from occlusion of penetrating arterioles. These microscopic lesions can be observed with high-resolution magnetic resonance imaging in aging brains and in patients with cerebrovascular disease. Recent studies have suggested that strategically located microinfarcts strongly correlate with cognitive deficits, which can contribute to Alzheimer's disease as well as other forms of dementia. We have recently shown that the molecular organization of axons into functional microdomains is altered in areas adjacent to white matter lacunar and microinfarcts, creating a peri-infarct penumbral injury in surviving axons. Whether similar changes in nodal, adjacent paranodal, and proximal axon initial segment molecular organization occur in the cortex adjacent to human CMIs is not known. Paraffin-embedded sections of autopsy brain tissue from five patients with CMIs were immunofluorescently labeled for nodal and paranodal markers including beta-IV spectrin, ankyrin-G, and contactin-associated protein. High magnification images from the peri-infarct cortical tissue were generated using confocal microscopy. In surviving cortical tissue adjacent to microinfarcts, we observed a dramatic loss of axon initial segments, suggesting that neuronal firing capacity in adjacent cortical tissue is likely compromised. The number of identifiable nodal/paranodal complexes in surviving cortical tissue is reduced adjacent to microinfarcts, while the average paranodal length is increased indicating a breakdown of axoglial contact. This axonal microdomain disorganization occurs in the relative absence of changes in the structural integrity of myelinated axons as measured by myelin basic protein and neurofilament staining. These findings indicate that the molecular organization of surviving axons adjacent to human CMIs is abnormal, reflecting lost axoglial contact and the functional elements necessary for neural transmission. This study provides support for the concept of a microinfarct penumbral injury that may account for the cumulative cognitive effect of these tiny strokes.

Associations between hippocampal morphometry and neuropathologic markers of Alzheimer's disease using 7 T MRI.

Hippocampal atrophy, amyloid plaques, and neurofibrillary tangles are established pathologic markers of Alzheimer's disease. We analyzed the temporal lobes of 9 Alzheimer's dementia (AD) and 7 cognitively normal (NC) subjects. Brains were scanned post-mortem at 7 Tesla. We extracted hippocampal volumes and radial distances using automated segmentation techniques. Hippocampal slices were stained for amyloid beta (Aß), tau, and cresyl violet to evaluate neuronal counts. The hippocampal subfields, CA1, CA2, CA3, CA4, and subiculum were manually traced so that the neuronal counts, Aß, and tau burden could be obtained for each region. We used linear regression to detect associations between hippocampal atrophy in 3D, clinical diagnosis and total as well as subfield pathology burden measures. As expected, we found significant correlations between hippocampal radial distance and mean neuronal count, as well as diagnosis. There were subfield specific associations between hippocampal radial distance and tau in CA2, and cresyl violet neuronal counts in CA1 and subiculum. These results provide further validation for the European Alzheimer's Disease Consortium Alzheimer's Disease Neuroimaging Initiative Center Harmonized Hippocampal Segmentation Protocol (HarP).

Huntington's disease accelerates epigenetic aging of human brain and disrupts DNA methylation levels.

Age of Huntington's disease (HD) motoric onset is strongly related to the number of CAG trinucleotide repeats in the huntingtin gene, suggesting that biological tissue age plays an important role in disease etiology. Recently, a DNA methylation based biomarker of tissue age has been advanced as an epigenetic aging clock. We sought to inquire if HD is associated with an accelerated epigenetic age. DNA methylation data was generated for 475 brain samples from various brain regions of 26 HD cases and 39 controls. Overall, brain regions from HD cases exhibit a significant epigenetic age acceleration effect (p=0.0012). A multivariate model analysis suggests that HD status increases biological age by 3.2 years. Accelerated epigenetic age can be observed in specific brain regions (frontal lobe, parietal lobe, and cingulate gyrus). After excluding controls, we observe a negative correlation (r=-0.41, p=5.5×10-8) between HD gene CAG repeat length and the epigenetic age of HD brain samples. Using correlation network analysis, we identify 11 co-methylation modules with a significant association with HD status across 3 broad cortical regions. In conclusion, HD is associated with an accelerated epigenetic age of specific brain regions and more broadly with substantial changes in brain methylation levels.

Neuropathology of Autosomal Dominant Alzheimer Disease in the National Alzheimer Coordinating Center Database.

Alzheimer disease (AD) represents a genetically heterogeneous entity. To elucidate neuropathologic features of autosomal dominant AD ([ADAD] due to PSEN1, APP, or PSEN2 mutations), we compared hallmark AD pathologic findings in 60 cases of ADAD and 120 cases of sporadic AD matched for sex, race, ethnicity, and disease duration. Greater degrees of neuritic plaque and neurofibrillary tangle formation and cerebral amyloid angiopathy (CAA) were found in ADAD (p values < 0.01). Moderate to severe CAA was more prevalent in ADAD (63.3% vs. 39.2%, p = 0.003), and persons with PSEN1 mutations beyond codon 200 had higher average Braak scores and severity and prevalence of CAA than those with mutations before codon 200. Lewy body pathology was less extensive in ADAD but was present in 27.1% of cases. We also describe a novel pathogenic PSEN1 mutation (P267A). The finding of more severe neurofibrillary pathology and CAA in ADAD, particularly in carriers of PSEN1 mutations beyond codon 200, warrants consideration when designing trials to treat or prevent ADAD. The finding of Lewy body pathology in a substantial minority of ADAD cases supports the assertion that development of Lewy bodies may be in part driven by abnormal ß-amyloid protein precursor processing.

GDF10 is a signal for axonal sprouting and functional recovery after stroke.

Stroke produces a limited process of neural repair. Axonal sprouting in cortex adjacent to the infarct is part of this recovery process, but the signal that initiates axonal sprouting is not known. Growth and differentiation factor 10 (GDF10) is induced in peri-infarct neurons in mice, non-human primates and humans. GDF10 promotes axonal outgrowth in vitro in mouse, rat and human neurons through TGFßRI and TGFßRII signaling. Using pharmacogenetic gain- and loss-of-function studies, we found that GDF10 produced axonal sprouting and enhanced functional recovery after stroke; knocking down GDF10 blocked axonal sprouting and reduced recovery. RNA sequencing from peri-infarct cortical neurons revealed that GDF10 downregulated PTEN, upregulated PI3 kinase signaling and induced specific axonal guidance molecules. Using unsupervised genome-wide association analysis of the GDF10 transcriptome, we found that it was not related to neurodevelopment, but may partially overlap with other CNS injury patterns. Thus, GDF10 is a stroke-induced signal for axonal sprouting and functional recovery.

The cerebellum ages slowly according to the epigenetic clock.

Studies that elucidate why some human tissues age faster than others may shed light on how we age, and ultimately suggest what interventions may be possible. Here we utilize a recent biomarker of aging (referred to as epigenetic clock) to assess the epigenetic ages of up to 30 anatomic sites from supercentenarians (subjects who reached an age of 110 or older) and younger subjects. Using three novel and three published human DNA methylation data sets, we demonstrate that the cerebellum ages more slowly than other parts of the human body. We used both transcriptional data and genetic data to elucidate molecular mechanisms which may explain this finding. The two largest superfamilies of helicases (SF1 and SF2) are significantly over-represented (p=9.2x10-9) among gene transcripts that are over-expressed in the cerebellum compared to other brain regions from the same subject. Furthermore, SNPs that are associated with epigenetic age acceleration in the cerebellum tend to be located near genes from helicase superfamilies SF1 and SF2 (enrichment p=5.8x10-3). Our genetic and transcriptional studies of epigenetic age acceleration support the hypothesis that the slow aging rate of the cerebellum is due to processes that involve RNA helicases.

Accelerated epigenetic aging in Down syndrome.

Down Syndrome (DS) entails an increased risk of many chronic diseases that are typically associated with older age. The clinical manifestations of accelerated aging suggest that trisomy 21 increases the biological age of tissues, but molecular evidence for this hypothesis has been sparse. Here, we utilize a quantitative molecular marker of aging (known as the epigenetic clock) to demonstrate that trisomy 21 significantly increases the age of blood and brain tissue (on average by 6.6 years, P = 7.0 × 10(-14)).

Relationship between hippocampal atrophy and neuropathology markers: a 7T MRI validation study of the EADC-ADNI Harmonized Hippocampal Segmentation Protocol.

OBJECTIVE: The pathologic validation of European Alzheimer's Disease Consortium Alzheimer's Disease Neuroimaging Initiative Center Harmonized Hippocampal Segmentation Protocol (HarP). METHODS: Temporal lobes of nine Alzheimer's disease (AD) and seven cognitively normal subjects were scanned post-mortem at 7 Tesla. Hippocampal volumes were obtained with HarP. Six-micrometer-thick hippocampal slices were stained for amyloid beta (Aß), tau, and cresyl violet. Hippocampal subfields were manually traced. Neuronal counts, Aß, and tau burden for each hippocampal subfield were obtained. RESULTS: We found significant correlations between hippocampal volume and Braak and Braak staging (ρ = -0.75, P = .001), tau (ρ = -0.53, P = .034), Aß burden (ρ = -0.61, P = .012), and neuronal count (ρ = 0.77, P < .001). Exploratory subfield-wise significant associations were found for Aß in Cornu Ammonis (CA)1 (ρ = -0.58, P = .019) and subiculum (ρ = -0.75, P = .001), tau in CA2 (ρ = -0.59, P = .016), and CA3 (ρ = -0.5, P = .047), and neuronal count in CA1 (ρ = 0.55, P = .028), CA3 (ρ = 0.65, P = .006), and CA4 (ρ = 0.76, P = .001). CONCLUSIONS: The observed associations provide pathological confirmation of hippocampal morphometry as a valid biomarker for AD and pathologic validation of HarP.

Molecular disorganization of axons adjacent to human lacunar infarcts.

Cerebral microvascular disease predominantly affects brain white matter and deep grey matter, resulting in ischaemic damage that ranges from lacunar infarcts to white matter hyperintensities seen on magnetic resonance imaging. These lesions are common and result in both clinical stroke syndromes and accumulate over time, resulting in cognitive deficits and dementia. Magnetic resonance imaging studies suggest that these lesions progress over time, accumulate adjacent to prior lesions and have a penumbral region susceptible to further injury. The pathological correlates of this adjacent injury in surviving myelinated axons have not been previously defined. In this study, we sought to determine the molecular organization of axons in tissue adjacent to lacunar infarcts and in the regions surrounding microinfarcts, by determining critical elements in axonal function: the morphology and length of node of Ranvier segments and adjacent paranodal segments. We examined post-mortem brain tissue from six patients with lacunar infarcts and tissue from two patients with autosomal dominant retinal vasculopathy and cerebral leukoencephalopathy (previously known as hereditary endotheliopathy with retinopathy, nephropathy and stroke) who accumulate progressive white matter ischaemic lesions in the form of lacunar and microinfarcts. In axons adjacent to lacunar infarcts yet extending up to 150% of the infarct diameter away, both nodal and paranodal length increase by ∼20% and 80%, respectively, reflecting a loss of normal cell-cell adhesion and signalling between axons and oligodendrocytes. Using premorbid magnetic resonance images, brain regions from patients with retinal vasculopathy and cerebral leukoencephalopathy that harboured periventricular white matter hyperintensities were selected and the molecular organization of axons was determined within these regions. As in regions adjacent to lacunar infarcts, nodal and paranodal length in white matter of these patients is increased. Myelin basic protein and neurofilament immunolabelling demonstrates that axons in these adjacent regions have preserved axonal cytoskeleton organization and are generally myelinated. This indicates that the loss of normal axonal microdomain architecture results from disrupted axoglial signalling in white matter adjacent to lacunar and microinfarcts. The loss of the normal molecular organization of nodes and paranodes is associated with axonal degeneration and may lead to impaired conduction velocity across surviving axons after stroke. These findings demonstrate that the degree of white matter injury associated with cerebral microvascular disease extends well beyond what can be identified using imaging techniques and that an improved understanding of the neurobiology in these regions can drive new therapeutic strategies for this disease entity.

Comorbidity in dementia: update of an ongoing autopsy study.

OBJECTIVES: To examine systemic and central nervous system (CNS) comorbidities of individuals with dementia evaluated during general autopsy. DESIGN: Retrospective cohort study. SETTING: A large tertiary academic medical center in Los Angeles, California. PARTICIPANTS: Individuals with clinically and neuropathologically diagnosed dementia who received complete autopsies (n = 86) and individuals with dementia who received partial (brain only) autopsies (n = 132). MEASUREMENTS: Information on cause of death and systemic and CNS comorbidities was obtained from autopsy reports and clinical information as available from the medical records. Findings were tabulated with respect to type of dementia, semiquantitative assessment of the severity of cerebral amyloid angiopathy, semiquantitative assessment of the severity of cerebrovascular disease, and evidence of ischemic damage in the brain. RESULTS: Of 218 subjects with dementia, 175 (80.3%) had Alzheimer's disease alone or in combination with other lesions that might contribute to cognitive impairment, such as cerebrovascular disease and diffuse Lewy body disease (DLBD), 14 (6.4%) had frontotemporal dementia, and seven (3.2%) had isolated DLBD. The most common cause of death in participants with dementia was pneumonia (n = 57, 66.3%), followed by cardiovascular disease (n = 14, 16.3%). Eighteen subjects (20.9%) had lung disease, and 16 (18.6%) had evidence of an old or recent myocardial infarction. Clinically undiagnosed neoplasms included colonic adenocarcinoma, metastatic pulmonary neuroendocrine carcinoma, meningioma, and Schwannoma. CONCLUSION: Significant comorbidities were discovered at autopsy in individuals with dementia. Understanding the causes of death and associated comorbidities in individuals with various subtypes of dementia is important in the assessment of end-of-life care in these individuals.

A shift in microglial ß-amyloid binding in Alzheimer's disease is associated with cerebral amyloid angiopathy.

Alzheimer's disease (AD) and cerebral amyloid angiopathy (CAA) are two common pathologies associated with ß-amyloid (Aß) accumulation and inflammation in the brain; neither is well understood. The objective of this study was to evaluate human post-mortem brains from AD subjects with purely parenchymal pathology, and those with concomitant CAA (and age-matched controls) for differential expression of microglia-associated Aß ligands thought to mediate Aß clearance and the association of these receptors with complement activation. Homogenates of brain parenchyma and enriched microvessel fractions from occipital cortex were probed for levels of C3b, membrane attack complex (MAC), CD11b and α-2-macroglobulin and immunoprecipitation was used to immunoprecipitate (IP) CD11b complexed with C3b and Aß. Both C3b and MAC were significantly increased in CAA compared to AD-only and controls and IP showed significantly increased CD11b/C3b complexes with Aß in AD/CAA subjects. Confocal microscopy was used to visualize these interactions. MAC was remarkably associated with CAA-affected blood vessels compared to AD-only and control vessels. These findings are consistent with an Aß clearance mechanism via microglial CD11b that delivers Aß and C3b to blood vessels in AD/CAA, which leads to Aß deposition and propagation of complement to the cytolytic MAC, possibly leading to vascular fragility.

Intimal thickening of meningeal arteries after serial corticectomies for Rasmussen encephalitis.

Rasmussen encephalitis is a rare cause of intractable epilepsy in children. Between 2008 and 2010, 4 patients had second cortical resections performed after a primary corticectomy for Rasmussen encephalitis. In each case, we observed some degree of vessel wall change in leptomeningeal arteries, consisting of moderate to moderately severe intimal hyperplasia. The intervals between original resection and second operation ranged from 8 months to 10 years. Ages of the patients ranged from 9 to 12 years at their first resection and from 10 to 19 years at the time of revision. Four other Rasmussen encephalitis cases operated upon in the years 2006 to 2010 and 2 surgical revisions for severe cortical dysplasia, 1 for mild cortical dysplasia and 1 for recurrent dysembryoplastic neuroepithelial tumor, did not show significant vascular abnormalities (with surgical intervals of 10 months to 16 years). Leptomeningeal intimal hyperplasia appears to develop in the interval between repeated cortical resections for Rasmussen encephalitis, an inflammatory disorder. The pathogenesis of this vascular change may be related to meningeal inflammation in Rasmussen encephalitis. This finding in children undergoing surgical resection for Rasmussen encephalitis may itself lead to "secondary" ischemic change that contributes to worsening of epilepsy.

Antioxidant sestrin-2 redistribution to neuronal soma in human immunodeficiency virus-associated neurocognitive disorders.

Sestrin-2 is involved in p53-dependent antioxidant defenses and in the maintenance of metabolic homeostasis. We hypothesize that sestrin-2 expression is altered in the brains of subjects diagnosed with human immunodeficiency virus (HIV)-associated neurocognitive disorders (HAND) due to neuronal oxidative stress. We studied sestrin-2 immunoreactivity in 42 isocortex sections from HIV-1-infected subjects compared to 18 age-matched non-HIV controls and 19 advanced Alzheimer's disease (AD) cases. With HIV infection, the sestrin-2 immunoreactivity pattern shifted from neuropil predominance (N) to neuropil and neuronal-soma co-dominance (NS) and neuronal-soma predominance (S; P < 0.0001, Chi-square test for linear trend). Among HIV cases showing the NS or S pattern, HAND cases were preferentially associated with the S pattern (n = 10 of 20) compared to cognitively intact cases (n = 1 of 11; P = 0.047, Fisher's exact test). In AD brains, sestrin-2 immunoreactivity was mostly intense in the neuropil and co-localized with phospho-Tau immunoreactivity in a subset of neurofibrillary lesions. Phospho-Tau-immunoreactive neurofibrillary lesions were rare in HIV cases and their occurrence was not associated with HAND. Levels of isocortical 8-hydroxy-deoxyguanosine (marker of nucleic acid oxidation) immunoreactivity were not significantly altered in HAND cases compared to cognitively intact HIV cases. In conclusion, the sestrin-2 immunoreactivity redistribution to neuronal soma in HAND suggests unique involvement of sestrin-2 in the pathophysiology of HAND, which is different from the role of sestrin-2 in AD pathogenesis. Alternatively, the difference in sestrin-2 immunoreactivity distribution between HAND and AD may be related to different degrees of severity or stages of oxidative stress.

Increased ceramide in brains with Alzheimer's and other neurodegenerative diseases.

Ceramide has been suggested to participate in the neuronal cell death that leads to Alzheimer's disease (AD), but its role is not yet well-understood. We compared the levels of six ceramide subspecies, which differ in the length of their fatty acid moieties, in brains from patients who suffered from AD, other neuropathological disorders, or both. We found elevated levels of Cer16, Cer18, Cer20, and Cer24 in brains from patients with any of the tested neural defects. Moreover, ceramide levels were highest in patients with more than one neuropathologic abnormality. Interestingly, the range of values was higher among brains with neural defects than in controls, suggesting that the regulation of ceramide synthesis is normally under tight control, and that this tight control may be lost during neurodegeneration. These changes, however, did not alter the ratio between the tested ceramide species. To explore the mechanisms underlying this dysregulation, we evaluated the expression of four genes connected to ceramide metabolism: ASMase, NSMase 2, GALC, and UGCG. The patterns of gene expression were complex, but overall, ASMase, NSMase 2, and GALC were upregulated in specimens from patients with neuropathologic abnormalities in comparison with age-matched controls. Such findings suggest these genes as attractive candidates both for diagnostic purposes and for intervening in neurodegenerative processes.

Correlation of hypointensities in susceptibility-weighted images to tissue histology in dementia patients with cerebral amyloid angiopathy: a postmortem MRI study.

Neuroimaging with iron-sensitive MR sequences [gradient echo T2* and susceptibility-weighted imaging (SWI)] identifies small signal voids that are suspected brain microbleeds. Though the clinical significance of these lesions remains uncertain, their distribution and prevalence correlates with cerebral amyloid angiopathy (CAA), hypertension, smoking, and cognitive deficits. Investigation of the pathologies that produce signal voids is necessary to properly interpret these imaging findings. We conducted a systematic correlation of SWI-identified hypointensities to tissue pathology in postmortem brains with Alzheimer's disease (AD) and varying degrees of CAA. Autopsied brains from eight AD patients, six of which showed advanced CAA, were imaged at 3T; foci corresponding to hypointensities were identified and studied histologically. A variety of lesions was detected; the most common lesions were acute microhemorrhage, hemosiderin residua of old hemorrhages, and small lacunes ringed by hemosiderin. In lesions where the bleeding vessel could be identified, ß-amyloid immunohistochemistry confirmed the presence of ß-amyloid in the vessel wall. Significant cellular apoptosis was noted in the perifocal region of recent bleeds along with heme oxygenase 1 activity and late complement activation. Acutely extravasated blood and hemosiderin were noted to migrate through enlarged Virchow­Robin spaces propagating an inflammatory reaction along the local microvasculature; a mechanism that may contribute to the formation of lacunar infarcts. Correlation of imaging findings to tissue pathology in our cases indicates that a variety of CAA-related pathologies produce MR-identified signal voids and further supports the use of SWI as a biomarker for this disease.

Increased activation of Iba1+ microglia in pediatric epilepsy patients with Rasmussen's encephalitis compared with cortical dysplasia and tuberous sclerosis complex.

Microgliosis is prominent in Rasmussen's encephalitis (RE), a disease with severe seizure activity. However, it is unclear if microglial activation is similar with different histopathologic substrates. Iba1-immunolabelled microglial activation was assessed in neocortex from pediatric epilepsy surgery patients with RE (n=8), cortical dysplasia (CD; n=6) and tuberous sclerosis complex (TSC; n=6). Microglial reactivity was increased, in severely affected RE areas (29% labeling) compared with minimally affected areas of RE cases (15%) and cases of TSC (14%) and CD (12%). There was no qualitative association of Iba1 immunolabelling with the presence of CD8(+) cytotoxic T-cells and no statistical association with clinical epilepsy variables, such as seizure duration or frequency. Iba1 appears to be an excellent marker for detecting extensive microglial activation in patients with RE. In addition, this study supports the notion that Iba1-labeled microglial activation is increased in patients with active RE, compared with cases of CD and TSC.