Emmanuelle Uro-Coste

Beneficial effect of human anti-amyloid-β active immunization on neurite morphology and tau pathology

Anti-amyloid-beta immunization leads to amyloid clearance in patients with Alzheimers disease, but the effect of vaccination on amyloid-beta-induced neuronal pathology has not been quantitatively examined. The objectives of this study were to address the effects of anti-amyloid-beta active immunization on neurite trajectories and the pathological hallmarks of Alzheimers disease in the human hippocampus. Hippocampal sections from five patients with Alzheimers disease enrolled in the AN1792 Phase 2a trial were compared with those from 13 non-immunized Braak-stage and age-matched patients with Alzheimers disease, and eight age-matched non-demented controls. Analyses included neurite curvature ratio as a quantitative measure of neuritic abnormalities, amyloid and tau loads, and a quantitative characterization of plaque-associated neuritic dystrophy and astrocytosis. Amyloid load and density of dense-core plaques were decreased in the immunized group compared to non-immunized patients (P < 0.01 and P < 0.001, respectively). The curvature ratio in non-immunized patients with Alzheimers disease was elevated compared to non-demented controls (P < 0.0001). In immunized patients, however, the curvature ratio was normalized when compared to non-immunized patients (P < 0.0001), and not different from non-demented controls. In the non-immunized patients, neurites close to dense-core plaques (within 50 microm) were more abnormal than those far from plaques (i.e. beyond 50 microm) (P < 0.0001). By contrast, in the immunized group neurites close to and far from the remaining dense-core plaques did not differ, and both were straighter compared to the non-immunized patients (P < 0.0001). Compared to non-immunized patients, dense-core plaques remaining after immunization had similar degree of astrocytosis (P = 0.6060), more embedded dystrophic neurites (P < 0.0001) and were more likely to have mitochondrial accumulation (P < 0.001). In addition, there was a significant decrease in the density of paired helical filament-1-positive neurons in the immunized group as compared to the non-immunized (P < 0.05), but not in the density of Alz50 or thioflavin-S positive tangles, suggesting a modest effect of anti-amyloid-beta immunization on tangle pathology. Clearance of amyloid plaques upon immunization with AN1792 effectively improves a morphological measure of neurite abnormality in the hippocampus. This improvement is not just attributable to the decrease in plaque load, but also occurs within the halo of the remaining dense-core plaques. However, these remaining plaques still retain some of their toxic potential. Anti-amyloid-beta immunization might also ameliorate the hippocampal tau pathology through a decrease in tau phosphorylation. These data agree with preclinical animal studies and further demonstrate that human anti-amyloid-beta immunization does not merely clear amyloid from the Alzheimers disease brain, but reduces some of the neuronal alterations that characterize Alzheimers disease.

Recessive TTN truncating mutations define novel forms of core myopathy with heart disease

Core myopathies (CM), the main non-dystrophic myopathies in childhood, remain genetically unexplained in many cases. Heart disease is not considered part of the typical CM spectrum. No congenital heart defect has been reported, and childhood-onset cardiomyopathy has been documented in only two CM families with homozygous mutations of the TTN gene. TTN encodes titin, a giant protein of striated muscles. Recently, heterozygous TTN truncating mutations have also been reported as a major cause of dominant dilated cardiomyopathy. However, relatively few TTN mutations and phenotypes are known, and titin pathophysiological role in cardiac and skeletal muscle conditions is incompletely understood. We analyzed a series of 23 families with congenital CM and primary heart disease using TTN M-line-targeted sequencing followed in selected patients by whole-exome sequencing and functional studies. We identified seven novel homozygous or compound heterozygous TTN mutations (five in the M-line, five truncating) in 17% patients. Heterozygous parents were healthy. Phenotype analysis identified four novel titinopathies, including cardiac septal defects, left ventricular non-compaction, Emery-Dreifuss muscular dystrophy or arthrogryposis. Additionally, in vitro studies documented the first-reported absence of a functional titin kinase domain in humans, leading to a severe antenatal phenotype. We establish that CM are associated with a large range of heart conditions of which TTN mutations are a major cause, thereby expanding the TTN mutational and phenotypic spectrum. Additionally, our results suggest titin kinase implication in cardiac morphogenesis and demonstrate that heterozygous TTN truncating mutations may not manifest unless associated with a second mutation, reassessing the paradigm of their dominant expression.

Stress-induced apoptosis is not mediated by endolysosomal ceramide

A major lipid‐signaling pathway in mammalian cells implicates the generation of ceramide from the ubiquitous sphingolipid sphingomyelin (SM). Hydrolysis of SM by a sphingomyeli‐nase present in acidic compartments has been reported to mediate, via the production of cer‐amide, the apoptotic cell death triggered by stress‐inducing agents. In the present study, we investigated whether the ceramide formed within or accumulated in lysosomes indeed triggers apopto‐sis. A series of observations strongly suggests that ceramide involved in stress‐induced apoptosis is not endolysosomal: 1) Although short‐chain cer‐amides induced apoptosis, loading cells with natural ceramide through receptor‐mediated endocy‐tosis did not result in cell death. 2) Neither TNF‐α nor anti‐CD95 induced the degradation to ceramide of a natural SM that had been first introduced selectively into acidic compartments. 3) Stimulation of SV40‐transformed fibroblasts by TNF‐α or CD40 ligand resulted in apoptosis equally well in cells derived from control individuals and from patients affected with Farber disease, having a genetic defect of acid ceramidase activity leading to lysosomal accumulation of cer‐amide. Also, induction of apoptosis using anti‐CD95 (Fas) or anti‐CD40 antibodies, TNF‐α, daunorubicin, and ionizing radiation was similar in control and Farber disease lymphoid cells. In all cases, apoptosis was preceded by a comparable increase of intracellular ceramide levels. 4) Retro‐viral‐mediated gene transfer and overexpression of acid ceramidase in Farber fibroblasts, which led to complete metabolic correction of the ceramide catabolic defect, did not affect the cell response to TNF‐α and CD40 ligand.—Ségui, B., Bezombes, C., Uro‐Coste, E., Medin, J. A., Andrieu‐Abadie, N., Augé, N., Brouchet, A., Laurent, G., Salvayre, R., Jaffrézou, J.‐P., Levade, T. Stress‐induced apoptosis is not mediated by endolysosomal ceramide. FASEB J. 14, 36–47(2000)

Beyond PrPres Type 1/Type 2 Dichotomy in Creutzfeldt-Jakob Disease

Sporadic Creutzfeldt-Jakob disease (sCJD) cases are currently subclassified according to the methionine/valine polymorphism at codon 129 of the PRNP gene and the proteinase K (PK) digested abnormal prion protein (PrPres) identified on Western blotting (type 1 or type 2). These biochemically distinct PrPres types have been considered to represent potential distinct prion strains. However, since cases of CJD show co-occurrence of type 1 and type 2 PrPres in the brain, the basis of this classification system and its relationship to agent strain are under discussion. Different brain areas from 41 sCJD and 12 iatrogenic CJD (iCJD) cases were investigated, using Western blotting for PrPres and two other biochemical assays reflecting the behaviour of the disease-associated form of the prion protein (PrPSc) under variable PK digestion conditions. In 30% of cases, both type 1 and type 2 PrPres were identified. Despite this, the other two biochemical assays found that PrPSc from an individual patient demonstrated uniform biochemical properties. Moreover, in sCJD, four distinct biochemical PrPSc subgroups were identified that correlated with the current sCJD clinico-pathological classification. In iCJD, four similar biochemical clusters were observed, but these did not correlate to any particular PRNP 129 polymorphism or western blot PrPres pattern. The identification of four different PrPSc biochemical subgroups in sCJD and iCJD, irrespective of the PRNP polymorphism at codon 129 and the PrPres isoform provides an alternative biochemical definition of PrPSc diversity and new insight in the perception of Human TSE agents variability.

Long-term Outcomes of CLIPPERS (Chronic Lymphocytic Inflammation With Pontine Perivascular Enhancement Responsive to Steroids) in a Consecutive Series of 12 Patients

BACKGROUND Chronic lymphocytic inflammation with pontine perivascular enhancement responsive to steroids (CLIPPERS) is a central nervous system inflammatory disease. OBJECTIVE To describe the disease course of CLIPPERS. DESIGN A nationwide study was implemented to collect clinical, magnetic resonance imaging, cerebrospinal fluid, and brain biopsy specimen characteristics of patients with CLIPPERS. SETTING Academic research. PATIENTS Twelve patients with CLIPPERS. MAIN OUTCOME MEASURES The therapeutic management of CLIPPERS was evaluated. RESULTS Among 12 patients, 42 relapses were analyzed. Relapses lasted a mean duration of 2.5 months, manifested frequent cerebellar ataxia and diplopia, and were associated with a mean Expanded Disability Status Scale (EDSS) score of 4. Besides typical findings of CLIPPERS, magnetic resonance imaging showed brainstem mass effect in 5 patients, extensive myelitis in 3 patients, and closed ring enhancement in 1 patient. Inconstant oligoclonal bands were found on cerebrospinal fluid investigation in 4 patients, with an increased T-cell ratio of CD4 to CD8. Among 7 available brain biopsy specimens, staining was positive for perivascular CD4 T lymphocytes in 5 samples. Thirty-eight of 42 relapses were treated with pulse corticosteroid therapy, which led to improvement, with a mean residual EDSS score of 1.9 (range, 0-7). In 1 patient with untreated relapses, scores on the EDSS progressively increased to a score of 10 at death. Among 5 patients without long-term corticosteroid therapy, the mean annualized relapse rate was 0.5 (range, 0.25-2.8). Among 7 patients taking oral corticosteroids, no relapses occurred in those whose daily dose was 20 mg or higher. No progressive course of CLIPPERS was observed. Four patients with a final EDSS score of 4 or higher had experienced previous severe relapses (EDSS score, ≥5) and brainstem and spinal cord atrophy. CONCLUSIONS CLIPPERS is a relapsing-remitting disorder without progressive forms. Long-term disability is correlated with the severity of previous relapses. Further studies are needed to confirm that prolonged corticosteroid therapy prevents further relapses.

Lysosomal sphingomyelinase is not solicited for apoptosis signaling

Stress‐induced activation of an acidic sphingomyelinase leading to generation of ceramide, an important lipid mediator, has been associated with apoptosis; however, the implication of this hydrolase has been questioned. The present study aimed at re‐evaluating the role of this lysosomal enzyme in apoptosis initiated by different apoptotic inducers. The sensitivity of a series of acid sphingomyelinase‐deficient cell lines derived from Niemann‐Pick disease patients to stress‐induced apoptosis was investigated. We have now shown that stress stimuli, such as anthracyclines, ionizing radiation, and Fas ligation trigger similar apoptotic hallmarks in normal and acid sphingomyelinase‐deficient cell lines. Retrovirus‐mediated gene correction of enzyme deficiency in Niemann‐Pick cells does not modify response to apoptosis. Ceramide production is comparable in normal and Niemann‐Pick cells, and increased activity of neutral sphingomyelinase is observed. Thus, our findings cast serious doubts that lysosomal sphingomyelinase activation is responsible for stress‐induced apoptosis of cultured cells.

Genetic and expression profiles of cerebellar liponeurocytomas

Cerebellar liponeurocytoma, a rare, newly identified CNS neoplasm of adults, is characterized by advanced neuronal/neurocytic and focal lipomatous differentiation, low proliferative potential and a favorable clinical prognosis. Despite the different age distribution and benign biological behavior, the cerebellar liponeurocytoma shares several features with the cerebellar medulloblastoma, which may include an origin from the periventricular matrix of the fourth ventricle or the external granular layer of the cerebellum. To establish the genetic profile of cerebellar liponeurocytomas, we have formed an international consortium and collected tumor samples from 20 patients. DNA sequencing revealed TP53 missense mutations in 4 (20%) of 20 cerebellar liponeurocytomas, a frequency higher than in medulloblastomas. There was no case with PTCH, APC, or β‐catenin mutations, each of which may be present in subsets of medulloblastomas. Isochromosome 17q, a genetic hallmark of classic medulloblastomas, was not observed in any of the cases investigated by FISH analysis. cDNA array analyses were carried out on 4 cerebellar liponeurocytomas, 4 central neurocytomas, and 4 classic medulloblastomas. Cluster analysis of the cDNA expression data of 1176 genes grouped cerebellar liponeurocytomas close to central neurocytomas, but distinct from medulloblastomas. These results suggest cerebellar liponeurocytoma as a distinct tumor entity that is genetically different from medulloblastoma. Furthermore, the cDNA expression array data suggest a relationship to central neurocytomas, but the presence of TP53 mutations, which are absent in central neurocytomas, suggests that their genetic pathways are different.

Astrocytes accumulate 4-hydroxynonenal adducts in murine scrapie and human Creutzfeldt-Jakob disease.

Scrapie-infected mice are considered a model for study in prion diseases, which are characterized by the progressive accumulation in the brain of an abnormal isoform (PrPsc) of the normal cellular prion protein (PrPc). Increasing data suggest that the neurodegenerative process in prion diseases may result, at least partially, from a defect in antioxidant function, but so far in vivo oxidative stress remains poorly documented. We report here that 4-hydroxynonenal, a lipid peroxidation by-product, forms protein adducts in brains of scrapie-infected mice and of Creutzfeldt-Jakob disease affected patients. In scrapie mice, studies on the progression of PrPsc accumulation, glial activation, ubiquitin deposition, and 4-HNE adduct formation allowed us to conclude the late occurrence of oxidative damage in the course of the disease. Massive 4-HNE accumulation was identified in astrocytes, but not in neurons or microglial cells. These findings suggest an important oxidative stress (and subsequent lipid peroxidation) in astrocytes, with possible consequences on their neuronal trophic function.

Contrast enhancement in 1p/19q-codeleted anaplastic oligodendrogliomas is associated with 9p loss, genomic instability, and angiogenic gene expression

BACKGROUND The aim of this study was to correlate MRI features and molecular characteristics in anaplastic oligodendrogliomas (AOs). METHODS The MRI characteristics of 50 AO patients enrolled in the French national network for high-grade oligodendroglial tumors were analyzed. The genomic profiles and IDH mutational statuses were assessed using high-resolution single-nucleotide polymorphism arrays and direct sequencing, respectively. The gene expression profiles of 25 1p/19q-codeleted AOs were studied on Affymetrix expression arrays. RESULTS Most of the cases were frontal lobe contrast-enhanced tumors (52%), but the radiological presentations of these cases were heterogeneous, ranging from low-grade glioma-like aspects (26%) to glioblastoma-like aspects (22%). The 1p/19q codeletion (n = 39) was associated with locations in the frontal lobe (P = .001), with heterogeneous intratumoral signal intensities (P = .003) and with no or nonmeasurable contrast enhancements (P = .01). The IDH wild-type AOs (n = 7) more frequently displayed ringlike contrast enhancements (P = .03) and were more frequently located outside of the frontal lobe (P = .01). However, no specific imaging pattern could be identified for the 1p/19q-codeleted AO or the IDH-mutated AO. Within the 1p/19q-codeleted AO, the contrast enhancement was associated with larger tumor volumes (P = .001), chromosome 9p loss and CDKN2A loss (P = .006), genomic instability (P = .03), and angiogenesis-related gene expression (P < .001), particularly for vascular endothelial growth factor A and angiopoietin 2. CONCLUSION In AOs, the 1p/19q codeletion and the IDH mutation are associated with preferential (but not with specific) imaging characteristics. Within 1p/19q-codeleted AO, imaging heterogeneity is related to additional molecular alterations, especially chromosome 9p loss, which is associated with contrast enhancement and larger tumor volume.

Reduced sphingosine kinase-1 and enhanced sphingosine 1-phosphate lyase expression demonstrate deregulated sphingosine 1-phosphate signaling in Alzheimer's disease

BackgroundThe accumulation of beta amyloid (Aβ) peptides, a hallmark of Alzheimer’s disease (AD) is related to mechanisms leading to neurodegeneration. Among its pleiotropic cellular effects, Aβ accumulation has been associated with a deregulation of sphingolipid metabolism. Sphingosine 1-phosphate (S1P) derived from sphingosine is emerging as a critical lipid mediator regulating various biological activities including cell proliferation, survival, migration, inflammation, or angiogenesis. S1P tissue level is low and kept under control through equilibrium between its synthesis mostly governed by sphingosine kinase-1 (SphK1) and its degradation by sphingosine 1-phosphate lyase (SPL). We have previously reported that Aβ peptides were able to decrease the activity of SphK1 in cell culture models, an effect that could be blocked by the prosurvival IGF-1/IGF-1R signaling.ResultsHerein, we report for the first time the expression of both SphK1 and SPL by immunohistochemistry in frontal and entorhinal cortices from 56 human AD brains. Immunohistochemical analysis revealed a decreased expression of SphK1 and an increased expression of SPL both correlated to amyloid deposits in the entorhinal cortex. Otherwise, analysis of brain tissue extracts showed a decrease of SphK1 expression in AD brains whereas SPL expression was increased. The content of IGF-1R, an activator of SphK1, was found decreased in AD brains as well as S1P1, the major receptor for S1P.ConclusionsCollectively, these results highlight the importance of S1P in AD suggesting the existence of a global deregulation of S1P signaling in this disease from its synthesis by SphK1 and degradation by SPL to its signaling by the S1P1 receptor.