Serum and cerebrospinal fluid neurofilament light chain and glial fibrillary acid protein levels in early and advanced stages of cerebral amyloid Angiopathy.

BACKGROUND: Neurofilament light chain (NFL) is a biomarker for neuroaxonal damage and glial fibrillary acidic protein (GFAP) for reactive astrocytosis. Both processes occur in cerebral amyloid angiopathy (CAA), but studies investigating the potential of NFL and GFAP as markers for CAA are lacking. We aimed to investigate NFL and GFAP as biomarkers for neuroaxonal damage and astrocytosis in CAA. METHODS: For this cross-sectional study serum and cerebrospinal fluid (CSF) samples were collected between 2010 and 2020 from controls, (pre)symptomatic Dutch-type hereditary (D-CAA) mutation-carriers and participants with sporadic CAA (sCAA) from two prospective CAA studies at two University hospitals in the Netherlands. NFL and GFAP levels were measured with Simoa-assays. The association between NFL and GFAP levels and age, cognitive performance (MoCA), CAA-related MRI markers (CAA-CSVD-burden) and Aß40 and Aß42 levels in CSF were assessed with linear regression adjusted for confounders. The control group was divided in age < 55 and ≥55 years to match the specific groups. RESULTS: We included 187 participants: 28 presymptomatic D-CAA mutation-carriers (mean age 40 years), 29 symptomatic D-CAA participants (mean age 58 years), 59 sCAA participants (mean age 72 years), 33 controls < 55 years (mean age 42 years) and 38 controls ≥ 55 years (mean age 65 years). In presymptomatic D-CAA, only GFAP in CSF (7.7*103pg/mL vs. 4.4*103pg/mL in controls; P<.001) was increased compared to controls. In symptomatic D-CAA, both serum (NFL:26.2pg/mL vs. 12.5pg/mL; P=0.008, GFAP:130.8pg/mL vs. 123.4pg/mL; P=0.027) and CSF (NFL:16.8*102pg/mL vs. 7.8*102pg/mL; P=0.01 and GFAP:11.4*103pg/mL vs. 7.5*103pg/mL; P<.001) levels were higher than in controls and serum levels (NFL:26.2pg/mL vs. 6.7pg/mL; P=0.05 and GFAP:130.8pg/mL vs. 66.0pg/mL; P=0.004) were higher than in pre-symptomatic D-CAA. In sCAA, only NFL levels were increased compared to controls in both serum (25.6pg/mL vs. 12.5pg/mL; P=0.005) and CSF (20.0*102pg/mL vs 7.8*102pg/mL; P=0.008). All levels correlated with age. Serum NFL correlated with MoCA (P=0.008) and CAA-CSVD score (P<.001). NFL and GFAP in CSF correlated with Aß42 levels (P=0.01/0.02). CONCLUSIONS: GFAP level in CSF is an early biomarker for CAA and is increased years before symptom onset. NFL and GFAP levels in serum and CSF are biomarkers for advanced CAA.

Dental sleep medicine education among undergraduate dental students in France.

INTRODUCTION: Dental sleep medicine (DSM) is an emerging discipline that studies the oral and maxillofacial causes and consequences of sleep-related problems. DSM is of upmost importance given the major medical challenge it represents. Therefore, to verify whether the future generation of dentists would be ready to face this challenge, the main objective of this study was to assess the degree of involvement of the French dental schools in teaching DSM at the undergraduate level. MATERIALS AND METHODS: All 16 dental schools in France were asked to participate in the study by answering to an online survey concerning the DSM curriculum during the 2018-2019 academic year. The survey was addressed to the head of the dental school and/or to relevant course coordinators and contained 10 questions related to the type, content and amount of DSM teaching to undergraduate dental students. RESULTS: Nine of the 16 (56.2%) French dental schools responded to the questionnaire. All these nine reported the inclusion of DSM in their undergraduate curriculum. The total average hours dedicated to teaching DSM was 5.6 h (SD 4.2; range 1-15 h). Seven of the 9 dental schools spent most of their DSM curriculum teaching time in the fifth year. All of them reviewed obstructive sleep apnoea and sleep-related bruxism and covered some topics related to therapies for sleep-related breathing disorders, such as the use of oral appliance. CONCLUSION: The results of this survey showed that, although the average hourly volume is relatively high, the DSM teaching in French dental schools appeared to be non-standardised, heterogeneous and often lacunar. It is therefore essential to develop a common curriculum and implement it in all dental schools to provide undergraduate students a comprehensive and updated teaching in DSM.

Patients with Obstructive Sleep Apnea and Cardiovascular Diseases: What, When, and Why Is Mandibular Advancement Device Treatment Required? A Short Review.

Obstructive sleep apnea is a potentially dangerous condition with significant risks of comorbidities if left untreated. It represents a cardiovascular risk factor in the general population, and a higher prevalence is observed in patients already suffering from cardiovascular diseases. The gold standard treatment, continuous positive airway pressure, is not always accepted or tolerated. The mandibular advancement device represents an alternative treatment that we propose to implement in our study. The objective here is to first present a brief review of the topic. Due to poor evidence in the field, we propose a pilot study to evaluate the effect of a mandibular advancement device in patients with cardiovascular disease who are not treated for their sleep pathology in order to improve their therapeutic management.

Cerebrospinal fluid p-tau231 as an early indicator of emerging pathology in Alzheimer's disease.

BACKGROUND: Phosphorylated tau (p-tau) epitopes in cerebrospinal fluid (CSF) are accurate biomarkers for a pathological and clinical diagnosis of Alzheimer's disease (AD) and are seen to be increased in preclinical stage of the disease. However, it is unknown if these increases transpire earlier, prior to amyloid-beta (Aß) positivity as determined by position emission tomography (PET), and if an ordinal sequence of p-tau epitopes occurs at this incipient phase METHODS: We measured CSF concentrations of p-tau181, p-tau217 and p-tau231 in 171 participants across the AD continuum who had undergone Aß ([18F]AZD4694) and tau ([18F]MK6240) position emission tomography (PET) and clinical assessment FINDINGS: All CSF p-tau biomarkers were accurate predictors of cognitive impairment but CSF p-tau217 demonstrated the largest fold-changes in AD patients in comparison to non-AD dementias and cognitively unimpaired individuals. CSF p-tau231 and p-tau217 predicted Aß and tau to a similar degree but p-tau231 attained abnormal levels first. P-tau231 was sensitive to the earliest changes of Aß in the medial orbitofrontal, precuneus and posterior cingulate before global Aß PET positivity was reached INTERPRETATION: We demonstrate that CSF p-tau231 increases early in development of AD pathology and is a principal candidate for detecting incipient Aß pathology for therapeutic trial application FUNDING: Canadian Institutes of Health Research (CIHR), Canadian Consortium of Neurodegeneration and Aging, Weston Brain Institute, Brain Canada Foundation, the Fonds de Recherche du Québec.

Cerebrospinal fluid hemoglobin levels as markers of blood contamination: relevance for α-synuclein measurement.

OBJECTIVES: Cerebrospinal fluid α-synuclein (CSF α-syn) represents a possible biomarker in Parkinson's disease (PD) diagnosis. CSF blood contamination can introduce a bias in α-syn measurement. To date, CSF samples with a red blood cells (RBC) count >50 RBC × 106/L or haemoglobin (Hb) concentration >200 µg/L are excluded from biomarker studies. However, investigations for defining reliable cut-off values are missing. METHODS: We evaluated the effect of blood contamination on CSF α-syn measurement by a systematic approach in a cohort of 42 patients with different neurological conditions who underwent lumbar puncture (LP) for diagnostic reasons. CSF samples were spiked with whole blood and serially diluted to 800, 400, 200, 100, 75, 50, 25, 5, 0 RBC × 106/L. CSF α-syn and Hb levels were measured by ELISA. RESULTS: In neat CSF, the average concentration of α-syn was 1,936 ± 636 ng/L. This value increased gradually in spiked CSF samples, up to 4,817 ± 1,456 ng/L (+149% α-syn variation) in samples with 800 RBC × 106/L. We established different cut-offs for discriminating samples with α-syn level above 5, 10, and 20% variation, corresponding to a Hb (RBC) concentration of 1,569 µg/L (37 RBC × 106/L), 2,082 µg/L (62 RBC × 106/L), and 3,118 µg/L (87 RBC × 106/L), respectively. CONCLUSIONS: Our data show the high impact of CSF blood contamination on CSF α-syn levels, highlighting the measurement of Hb concentration as mandatory when assessing CSF α-syn. The thresholds we calculated are useful to classify CSF samples for blood contamination, considering as reliable only those showing a Hb concentration <1,569 µg/L.

Core Alzheimer's disease cerebrospinal fluid biomarker assays are not affected by aspiration or gravity drip extraction methods.

BACKGROUND: CSF biomarkers are well-established for routine clinical use, yet a paucity of comparative assessment exists regarding CSF extraction methods during lumbar puncture. Here, we compare in detail biomarker profiles in CSF extracted using either gravity drip or aspiration. METHODS: Biomarkers for ß-amyloidopathy (Aß1-42, Aß1-40), tauopathy (total tau), or synapse pathology (BACE1, Neurogranin Trunc-p75, α-synuclein) were assessed between gravity or aspiration extraction methods in a sub-population of the Australian Imaging, Biomarkers and Lifestyle (AIBL) study (cognitively normal, N = 36; mild cognitive impairment, N = 8; Alzheimer's disease, N = 6). RESULTS: High biomarker concordance between extraction methods was seen (concordance correlation > 0.85). Passing Bablock regression defined low beta coefficients indicating high scalability. CONCLUSIONS: Levels of these commonly assessed CSF biomarkers are not influenced by extraction method. Results of this study should be incorporated into new consensus guidelines for CSF collection, storage, and analysis of biomarkers.

How specific are the conformation-specific α-synuclein antibodies? Characterization and validation of 16 α-synuclein conformation-specific antibodies using well-characterized preparations of α-synuclein monomers, fibrils and oligomers with distinct structures and morphology.

Increasing evidence suggests that alpha-synuclein (α-syn) oligomers are obligate intermediates in the pathway involved in α-syn fibrillization and Lewy body (LB) formation, and may also accumulate within LBs in Parkinson's disease (PD) and other synucleinopathies. Therefore, the development of tools and methods to detect and quantify α-syn oligomers has become increasingly crucial for mechanistic studies to understand their role in PD, and to develop new diagnostic methods and therapies for PD and other synucleinopathies. The majority of these tools and methods rely primarily on the use of aggregation state-specific or conformation-specific antibodies. Given the impact of the data and knowledge generated using these antibodies on shaping the foundation and directions of α-syn and PD research, it is crucial that these antibodies are thoroughly characterized, and their specificity or ability to capture diverse α-syn species is tested and validated. Herein, we describe an antibody characterization and validation pipeline that allows a systematic investigation of the specificity of α-syn antibodies using well-defined and well-characterized preparations of various α-syn species, including monomers, fibrils, and different oligomer preparations that are characterized by distinct morphological, chemical and secondary structure properties. This pipeline was used to characterize 18 α-syn antibodies, 16 of which have been reported as conformation- or oligomer-specific antibodies, using an array of techniques, including immunoblot analysis (slot blot and Western blot), a digital ELISA assay using single molecule array technology and surface plasmon resonance. Our results show that i) none of the antibodies tested are specific for one particular type of α-syn species, including monomers, oligomers or fibrils; ii) all antibodies that were reported to be oligomer-specific also recognized fibrillar α-syn; and iii) a few antibodies showed high specificity for oligomers and fibrils but did not bind to monomers. These findings suggest that the great majority of α-syn aggregate-specific antibodies do not differentiate between oligomers and fibrils, thus highlighting the importance of exercising caution when interpreting results obtained using these antibodies. Our results also underscore the critical importance of the characterization and validation of antibodies before their use in mechanistic studies and as diagnostic tools or therapeutic agents. This will not only improve the quality and reproducibility of research and reduce costs but will also reduce the number of therapeutic antibody failures in the clinic.

A Novel Tau Antibody Detecting the First Amino-Terminal Insert Reveals Conformational Differences Among Tau Isoforms.

As human Tau undergoes pathologically relevant post-translational modifications when expressed in yeast, the use of humanized yeast models for the generation of novel Tau monoclonal antibodies has previously been proven to be successful. In this study, human Tau2N4R-ΔK280 purified from yeast was used for the immunization of mice and subsequent selection of high affinity Tau-specific monoclonal antibodies. The characterization of four novel antibodies in different Tau model systems yielded a phosphorylation-dependent antibody (15A10), an antibody directed to the first microtubule-binding repeat domain (16B12), a carboxy-terminal antibody (20G10) and an antibody targeting an epitope on the hinge of the first and second amino-terminal insert (18F12). The latter was found to be conformation-dependent, suggesting structural differences between the Tau splicing isoforms and allowing insight in the roles played by the amino-terminal inserts. As this monoclonal antibody also has the capacity to detect tangle-like structures in different transgenic Tau mice and neurofibrillary tangles in brain sections of patients diagnosed with Alzheimer's disease, we also tested the diagnostic potential of 18F12 in a pilot study and found this monoclonal antibody to have the ability to discriminate Alzheimer's disease patients from control individuals based on increased Tau levels in the cerebrospinal fluid.

[OSA in adults: Role of the mandibular advancement device (MAD)]. / Troubles respiratoires obstructifs du sommeil de l'adulte : place de l'orthèse d'avancée mandibulaire.

Mandibular advancement devices are an alternative to continuous positive airway pressure for patients with mild or moderate obstructive sleep apnea/ hypopnea syndrome (OSA). Oral appliances advance the mandible and tongue, enlarge and stabilize the upper airways during sleep. Clinical examination is used to select candidates for oral appliances and a particular appliance for a given patient. Titration of the advancement will optimize its effectiveness by resolving subjective clinical symptoms. Polygraphic or ventilatory polysomnography controls the effectiveness of the treatment (IAH reduction, increase of oxygen saturation and improvement of the main symptoms). Clinical follow-up is required to assess longterm outcomes, side effects and treatment adherence. Oral appliances are effective in the short-term provided they achieved a 50% reduction in IAH and complete symptom resolution.

Automation on an Open-Access Platform of Alzheimer's Disease Biomarker Immunoassays.

The lack of (inter-)laboratory standardization has hampered the application of universal cutoff values for Alzheimer's disease (AD) cerebrospinal fluid (CSF) biomarkers and their transfer to general clinical practice. The automation of the AD biomarker immunoassays is suggested to generate more robust results than using manual testing. Open-access platforms will facilitate the integration of automation for novel biomarkers, allowing the introduction of the protein profiling concept. A feasibility study was performed on an automated open-access platform of the commercial immunoassays for the 42-amino-acid isoform of amyloid-ß (Aß1-42), Aß1-40, and total tau in CSF. Automated Aß1-42, Aß1-40, and tau immunoassays were performed within predefined acceptance criteria for bias and imprecision. Similar accuracy was obtained for ready-to-use calibrators as for reconstituted lyophilized kit calibrators. When compared with the addition of a standard curve in each test run, the use of a master calibrator curve, determined before and applied to each batch analysis as the standard curve, yielded an acceptable overall bias of -2.6% and -0.9% for Aß1-42 and Aß1-40, respectively, with an imprecision profile of 6.2% and 8.4%, respectively. Our findings show that transfer of commercial manual immunoassays to fully automated open-access platforms is feasible, as it performs according to universal acceptance criteria.

Recommendations for cerebrospinal fluid collection for the analysis by ELISA of neurogranin trunc P75, α-synuclein, and total tau in combination with Aß(1-42)/Aß(1-40).

BACKGROUND: The pathophysiology of neurodegeneration is complex. Its diagnosis requires an early identification of sequential changes in several hallmarks in the brains of affected subjects. The presence of brain pathology can be visualized in the cerebrospinal fluid (CSF) by protein profiling. It is clear that the field of Alzheimer's disease (AD) will benefit from an integration of algorithms including CSF concentrations of individual proteins, especially as an aid in clinical decision-making or to improve patient enrolment in clinical trials. The protein profiling approach requires standard operating procedures for collection and storage of CSF which must be easy to integrate into a routine clinical lab environment. Our study provides recommendations for analysis of neurogranin trunc P75, α-synuclein, and tau, in combination with the ratio of ß-amyloid Aß(1-42)/Aß(1-40). METHODS: Protocols for CSF collection were compared with CSF derived from subjects with normal pressure hydrocephalus (n = 19). Variables included recipient type (collection, storage), tube volume, and addition of detergents at the time of collection. CSF biomarker analysis was performed with enzyme-linked immunosorbent assays (ELISAs). Data were analyzed with linear repeated measures and mixed effects models. RESULTS: Adsorption to recipients is lower for neurogranin trunc P75, α-synuclein, and tau (<10%), as compared to Aß(1-42). For neurogranin trunc P75 and total tau, there is still an effect on analyte concentrations as a function of the tube volume. Protocol-related differences for Aß(1-42) can be normalized at the (pre-)analytical level using the ratio Aß(1-42)/Aß(1-40), but not by using the ratio Aß(1-42)/tau. The addition of detergent at the time of collection eliminates differences due to adsorption. CONCLUSIONS: Our study recommends the use of low protein binding tubes for quantification in CSF (without additives) of all relevant CSF biomarkers. Pre-analytical factors have less effect on α-synuclein, neurogranin trunc P75, and total tau, as compared to Aß(1-42). The ratio of Aß(1-42)/Aß(1-40), but not Aß(1-42)/tau, can be used to adjust for pre-analytical differences in analyte concentrations. Our study does not recommend the inclusion of detergents at the time of collection of CSF. The present results provide an experimental basis for new recommendations for parallel analysis of several proteins using one protocol for collection and storage of CSF.

Supplementing lactating women with flaxseed oil does not increase docosahexaenoic acid in their milk.

BACKGROUND: Flaxseed oil is a rich source of 18:3n-3 (alpha-linolenic acid, or ALA), which is ultimately converted to 22:6n-3 (docosahexaenoic acid, or DHA), a fatty acid important for the development of the infant brain and retina. OBJECTIVE: The objective of this study was to determine the effect of flaxseed oil supplementation on the breast-milk, plasma, and erythrocyte contents of DHA and other n-3 fatty acids in lactating women. DESIGN: Seven women took 20 g flaxseed oil (10.7 g ALA) daily for 4 wk. Breast-milk and blood samples were collected weekly before, during, and after supplementation and were analyzed for fatty acid composition. RESULTS: Breast milk, plasma, and erythrocyte ALA increased significantly over time (P < 0.001) and after 2 and 4 wk of supplementation (P < 0.05). Over time, 20:5n-3 (eicosapentaenoic acid, or EPA) increased significantly in breast milk (P = 0.004) and in plasma (P < 0.001). In addition, plasma EPA increased significantly (P < 0.05) after 2 and 4 wk of supplementation. There were significant increases over time in breast-milk 22:5n-3 (docosapentaenoic acid, or DPA) (P < 0.02), plasma DPA (P < 0.001), and erythrocyte DPA (P < 0.01). No significant changes were observed in breast-milk, plasma, or erythrocyte DHA contents after flaxseed oil supplementation. CONCLUSIONS: Dietary flaxseed oil increased the breast-milk, plasma, and erythrocyte contents of the n-3 fatty acids ALA, EPA, and DPA but had no effect on breast-milk, plasma, or erythrocyte DHA contents.