Marla Gearing

5-hmC-mediated epigenetic dynamics during postnatal neurodevelopment and aging

DNA methylation dynamics influence brain function and are altered in neurological disorders. 5-hydroxymethylcytosine (5-hmC), a DNA base that is derived from 5-methylcytosine, accounts for ∼40% of modified cytosine in the brain and has been implicated in DNA methylation–related plasticity. We mapped 5-hmC genome-wide in mouse hippocampus and cerebellum at three different ages, which allowed us to assess its stability and dynamic regulation during postnatal neurodevelopment through adulthood. We found developmentally programmed acquisition of 5-hmC in neuronal cells. Epigenomic localization of 5-hmC–regulated regions revealed stable and dynamically modified loci during neurodevelopment and aging. By profiling 5-hmC in human cerebellum, we found conserved genomic features of 5-hmC. Finally, we found that 5-hmC levels were inversely correlated with methyl-CpG–binding protein 2 dosage, a protein encoded by a gene in which mutations cause Rett syndrome. These data suggest that 5-hmC–mediated epigenetic modification is critical in neurodevelopment and diseases.

Oxidative Modifications and Down-regulation of Ubiquitin Carboxyl-terminal Hydrolase L1 Associated with Idiopathic Parkinson's and Alzheimer's Diseases

Alzheimers disease (AD) and Parkinsons disease (PD) are the two most common neurodegenerative diseases that occur either in relatively rare, familial forms or in common, sporadic forms. The genetic defects underlying several monogenic familial forms of AD and PD have recently been identified, however, the causes of other AD and PD cases, particularly sporadic cases, remain unclear. To gain insights into the pathogenic mechanisms involved in AD and PD, we used a proteomic approach to identify proteins with altered expression levels and/or oxidative modifications in idiopathic AD and PD brains. Here, we report that the protein level of ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1), a neuronal de-ubiquitinating enzyme whose mutation has been linked to an early-onset familial PD, is down-regulated in idiopathic PD as well as AD brains. By using a combination of two-dimensional gel electrophoresis and mass spectrometry, we have identified three human brain UCH-L1 isoforms, a full-length form and two amino-terminally truncated forms. Our proteomic analyses reveal that the full-length UCH-L1 is a major target of oxidative damage in AD and PD brains, which is extensively modified by carbonyl formation, methionine oxidation, and cysteine oxidation. Furthermore, immunohistochemical studies show that prominent UCH-L1 immunostaining is associated with neurofibrillary tangles and that the level of soluble UCH-L1 protein is inversely proportional to the number of tangles in AD brains. Together, these results provide evidence supporting a direct link between oxidative damage to the neuronal ubiquitination/de-ubiquitination machinery and the pathogenesis of sporadic AD and PD.

Primary age-related tauopathy (PART): a common pathology associated with human aging

We recommend a new term, “primary age-related tauopathy” (PART), to describe a pathology that is commonly observed in the brains of aged individuals. Many autopsy studies have reported brains with neurofibrillary tangles (NFTs) that are indistinguishable from those of Alzheimer’s disease (AD), in the absence of amyloid (Aβ) plaques. For these “NFT+/Aβ−” brains, for which formal criteria for AD neuropathologic changes are not met, the NFTs are mostly restricted to structures in the medial temporal lobe, basal forebrain, brainstem, and olfactory areas (bulb and cortex). Symptoms in persons with PART usually range from normal to amnestic cognitive changes, with only a minority exhibiting profound impairment. Because cognitive impairment is often mild, existing clinicopathologic designations, such as “tangle-only dementia” and “tangle-predominant senile dementia”, are imprecise and not appropriate for most subjects. PART is almost universally detectable at autopsy among elderly individuals, yet this pathological process cannot be specifically identified pre-mortem at the present time. Improved biomarkers and tau imaging may enable diagnosis of PART in clinical settings in the future. Indeed, recent studies have identified a common biomarker profile consisting of temporal lobe atrophy and tauopathy without evidence of Aβ accumulation. For both researchers and clinicians, a revised nomenclature will raise awareness of this extremely common pathologic change while providing a conceptual foundation for future studies. Prior reports that have elucidated features of the pathologic entity we refer to as PART are discussed, and working neuropathological diagnostic criteria are proposed.

Oxidative Damage of DJ-1 Is Linked to Sporadic Parkinson and Alzheimer Diseases

Mutations in DJ-1 cause an autosomal recessive, early onset familial form of Parkinson disease (PD). However, little is presently known about the role of DJ-1 in the more common sporadic form of PD and in other age-related neurodegenerative diseases, such as Alzheimer disease (AD). Here we report that DJ-1 is oxidatively damaged in the brains of patients with idiopathic PD and AD. By using a combination of two-dimensional gel electrophoresis and mass spectrometry, we have identified 10 different DJ-1 isoforms, of which the acidic isoforms (pI 5.5 and 5.7) of DJ-1 monomer and the basic isoforms (pI 8.0 and 8.4) of SDS-resistant DJ-1 dimer are selectively accumulated in PD and AD frontal cortex tissues compared with age-matched controls. Quantitative Western blot analysis shows that the total level of DJ-1 protein is significantly increased in PD and AD brains. Mass spectrometry analyses reveal that DJ-1 is not only susceptible to cysteine oxidation but also to previously unsuspected methionine oxidation. Furthermore, we show that DJ-1 protein is irreversibly oxidized by carbonylation as well as by methionine oxidation to methionine sulfone in PD and AD. Our study provides new insights into the oxidative modifications of DJ-1 and indicates association of oxidative damage to DJ-1 with sporadic PD and AD.

Common variants at 7p21 are associated with frontotemporal lobar degeneration with TDP-43 inclusions

Frontotemporal lobar degeneration (FTLD) is the second most common cause of presenile dementia. The predominant neuropathology is FTLD with TAR DNA-binding protein (TDP-43) inclusions (FTLD-TDP). FTLD-TDP is frequently familial, resulting from mutations in GRN (which encodes progranulin). We assembled an international collaboration to identify susceptibility loci for FTLD-TDP through a genome-wide association study of 515 individuals with FTLD-TDP. We found that FTLD-TDP associates with multiple SNPs mapping to a single linkage disequilibrium block on 7p21 that contains TMEM106B. Three SNPs retained genome-wide significance following Bonferroni correction (top SNP rs1990622, P = 1.08 × 10−11; odds ratio, minor allele (C) 0.61, 95% CI 0.53–0.71). The association replicated in 89 FTLD-TDP cases (rs1990622; P = 2 × 10−4). TMEM106B variants may confer risk of FTLD-TDP by increasing TMEM106B expression. TMEM106B variants also contribute to genetic risk for FTLD-TDP in individuals with mutations in GRN. Our data implicate variants in TMEM106B as a strong risk factor for FTLD-TDP, suggesting an underlying pathogenic mechanism.

The Consortium to Establish a Registry for Alzheimer's Disease (CERAD). Part X. Neuropathology Confirmation of the Clinical Diagnosis of Alzheimer's Disease

Article abstract-This report summarizes the neuropathologic findings in the first 106 autopsies of CERAD (Consortium to Establish a Registry for Alzheimers Disease) dementia patients diagnosed clinically as having Alzheimers disease (AD). In 92 (87%) of the 106 cases, neuropathologists confirmed Alzheimers disease (AD) as the primary dementing illness. Coexistent Parkinsons disease (PD) changes were present in 19 (21%) and vascular lesions of varying nature and size in 26 (28%) of these 92 AD cases. The 14 cases in which AD was not interpreted as the primary dementing illness can be divided into four major subgroups based on their neuropathology findings: PD and related pathology (n = 5), hippocampal sclerosis (n = 3), miscellaneous neurodegenerative and other disorders (n = 3), and no significant changes (n = 3). Despite the relatively high level of clinical diagnostic accuracy, further refinement of assessment batteries may facilitate distinction of non-AD dementias from AD. NEUROLOGY 1995;45: 461-466

Neuronal and glial pathological changes during epileptogenesis in the mouse pilocarpine model.

The rodent pilocarpine model of epilepsy exhibits hippocampal sclerosis and spontaneous seizures and thus resembles human temporal lobe epilepsy. Use of the many available mouse mutants to study this epilepsy model would benefit from a detailed neuropathology study. To identify new features of epileptogenesis, we characterized glial and neuronal pathologies after pilocarpine-induced status epilepticus (SE) in CF1 and C57BL/6 mice focusing on the hippocampus. All CF1 mice showed spontaneous seizures by 17-27 days after SE. By 6 h there was virtually complete loss of hilar neurons, but the extent of pyramidal cell death varied considerably among mice. In the mossy fiber pathway, neuropeptide Y (NPY) was persistently upregulated beginning 1 day after SE; NPY immunoreactivity in the supragranular layer after 31 days indicated mossy fiber sprouting. beta2 microglobulin-positive activated microglia, normally absent in brains without SE, became abundant over 3-31 days in regions of neuronal loss, including the hippocampus and the amygdala. Astrogliosis developed after 10 days in damaged areas. Amyloid precursor protein immunoreactivity in the thalamus at 10 days suggested delayed axonal degeneration. The mortality after pilocarpine injection was very high in C57BL/6 mice from Jackson Laboratories but not those from Charles River, suggesting that mutant mice in the C57BL/6(JAX) strain will be difficult to study in the pilocarpine model, although their neuropathology was similar to CF1 mice. Major neuropathological changes not previously studied in the rodent pilocarpine model include widespread microglial activation, delayed thalamic axonal death, and persistent NPY upregulation in mossy fibers, together revealing extensive and persistent glial as well as neuronal pathology.

Regulation of Neuronal Survival Factor MEF2D by Chaperone-Mediated Autophagy

Chaperone-mediated autophagy controls the degradation of selective cytosolic proteins and may protect neurons against degeneration. In a neuronal cell line, we found that chaperone-mediated autophagy regulated the activity of myocyte enhancer factor 2D (MEF2D), a transcription factor required for neuronal survival. MEF2D was observed to continuously shuttle to the cytoplasm, interact with the chaperone Hsc70, and undergo degradation. Inhibition of chaperone-mediated autophagy caused accumulation of inactive MEF2D in the cytoplasm. MEF2D levels were increased in the brains of α-synuclein transgenic mice and patients with Parkinsons disease. Wild-type α-synuclein and a Parkinsons disease–associated mutant disrupted the MEF2D-Hsc70 binding and led to neuronal death. Thus, chaperone-mediated autophagy modulates the neuronal survival machinery, and dysregulation of this pathway is associated with Parkinsons disease.

Cerebral infarcts in patients with autopsy-proven Alzheimer's disease CERAD, part XVIII

Objective: To study the relation between cerebral infarction and clinical and neuropsychologic manifestations in patients with autopsy-proven Alzheimers disease (AD) enrolled in the Consortium to Establish a Registry for Alzheimers Disease (CERAD). Background: Prior studies report that subjects with neuropathologic evidence of AD and concomitant brain infarcts had poorer cognitive function and higher frequency of dementia than those with AD alone. Methods: Clinical and neuropsychologic manifestations of dementia were studied in 74 subjects with neuropathologic findings of AD alone and 32 with AD and concomitant cerebral infarcts or lacunar lesions. Results: The 32 patients with both AD and vascular lesions were significantly older at time of death (median age, 81 years) than the 74 patients with AD alone(76 years; p = 0.02). At the final follow-up visit, the severity of the dementia was greater in AD patients with vascular lesions(median Clinical Dementia Rating [CDR] = 3) than in those with AD alone (CDR= 2; p = 0.03). Patients with AD and vascular lesions performed significantly worse on verbal fluency, Boston Naming, and Mini-Mental State Examination (MMSE) tests. No differences between the groups were observed, however, in the semiquantitative measures of frequency of neuritic plaques or neurofibrillary tangles. Conclusions: The clinical-neuropathologic correlations in CERAD patients generally confirm those in prior studies, indicating that the presence of cerebral infarction in patients with AD is associated with greater overall severity of clinical dementia and poorer performance on specific tests of language and cognitive function.

Expanded GGGGCC repeat RNA associated with amyotrophic lateral sclerosis and frontotemporal dementia causes neurodegeneration

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) share phenotypic and pathologic overlap. Recently, an expansion of GGGGCC repeats in the first intron of C9orf72 was found to be a common cause of both illnesses; however, the molecular pathogenesis of this expanded repeat is unknown. Here we developed both Drosophila and mammalian models of this expanded hexanucleotide repeat and showed that expression of the expanded GGGGCC repeat RNA (rGGGGCC) is sufficient to cause neurodegeneration. We further identified Pur α as the RNA-binding protein of rGGGGCC repeats and discovered that Pur α and rGGGGCC repeats interact in vitro and in vivo in a sequence-specific fashion that is conserved between mammals and Drosophila. Furthermore, overexpression of Pur α in mouse neuronal cells and Drosophila mitigates rGGGGCC repeat-mediated neurodegeneration, and Pur α forms inclusions in the fly eye expressing expanded rGGGGCC repeats, as well as in cerebellum of human carriers of expanded GGGGCC repeats. These data suggest that expanded rGGGGCC repeats could sequester specific RNA-binding protein from their normal functions, ultimately leading to cell death. Taken together, these findings suggest that the expanded rGGGGCC repeats could cause neurodegeneration, and that Pur α may play a role in the pathogenesis of amyotrophic lateral sclerosis and frontotemporal dementia.