13 research outputs found
Additional file 1 of A mutant methionyl-tRNA synthetase-based toolkit to assess induced-mesenchymal stromal cell secretome in mixed-culture disease models
Additional file 1: Fig. S1. Uncropped agarose gel from Figure 1B
Levels of CD325a-syn EVs in PD and correlation with UPDRS scores.
A) Levels of total a-syn EVs in PD and correlation with UPDRS scores; B) UPDRS scores and median levels of CD325a-syn EVs in PD; C) Levels of total a-syn EVs in PD and correlation with UPDRS scores; D) UPDRS scores and median levels of a-syn EVs in PD; E) Levels of total a-syn EVs in PD and correlation with UPDRS scores in replication cohort; F) UPDRS scores and median levels of a-syn EVs in PD in replication cohort.</p
Table_1_Adult-Onset Leukoencephalopathy With Axonal Spheroids and Pigmented Glia: Review of Clinical Manifestations as Foundations for Therapeutic Development.docx
A comprehensive review of published literature was conducted to elucidate the genetics, neuropathology, imaging findings, prevalence, clinical course, diagnosis/clinical evaluation, potential biomarkers, and current and proposed treatments for adult-onset leukoencephalopathy with axonal spheroids and pigmented glia (ALSP), a rare, debilitating, and life-threatening neurodegenerative disorder for which disease-modifying therapies are not currently available. Details on potential efficacy endpoints for future interventional clinical trials in patients with ALSP and data related to the burden of the disease on patients and caregivers were also reviewed. The information in this position paper lays a foundation to establish an effective clinical rationale and address the clinical gaps for creation of a robust strategy to develop therapeutic agents for ALSP, as well as design future clinical trials, that have clinically meaningful and convergent endpoints.</p
Fig 4 -
A) Levels of total a-syn EVs in PD and correlation with PAR activity in PD. B) UPDRS scores and PAR activity in PD; C) Levels of total a-syn EVs in PD and correlation with PAR activity in PD in replication cohort; D) UPDRS scores and PAR activity in PD in replication cohort.</p
Western blot of plasma-derived EVs comparing healthy donors and PD patients.
Total aSyn and aSyn S129 are detectable only on PD samples. No difference has been detected between total aSyn and aSyn S129.</p
Levels of α-synuclein extracellular vesicles distinguish Parkinson disease (PD) patients from age-matched healthy individuals, Alzheimer disease (AD) patients, and patients having dementia with Lewy bodies (DLB).
Comparison between levels of a-syn EVs in normal, AD, DLB, and PD patients. A) Levels of a-syn positive EVs in the four groups; B) Levels of CD32 a-syn EVs in the four groups C) Percentage of CD32 a-syn in the four groups; D) Receiving Operator Curve of identifying a-syn positive EVs compared to controls: sensitivity of 92.98% and a specificity of 78.95% at a cut-off of 20,700 α-synuclein-EVs per microliter of blood. *** indicates the first batch of samples where CD235a was used.</p
Nanoscale flow cytometric detection of aSyn- and CD235a-positive extracellular vesicles in platelet-free plasma.
A) Representative scatterplot and histogram of a mixture of nanosized beads (110 to 1,300 nm). B) Scatterplots of nanoscale flow cytometric detection of aSyn- and CD235a-positive EVs from platelet-free plasma. C) Antibody titration curves for alpha-synuclein and CD235a antibodies using platelet-free plasma (N = 3 technical replicates). Arrows indicate optimal concentrations for each antibody. (DOCX)</p
Sensitive ELISA-based detection method for the mitophagy marker p-S65-Ub in human cells, autopsy brain, and blood samples
Mitochondrial dysfunction is an early, imminent event in neurodegenerative disorders including Parkinson disease (PD) and Alzheimer disease (AD). The enzymatic pair PINK1 and PRKN/Parkin recognize and transiently label damaged mitochondria with ubiquitin (Ub) phosphorylated at Ser65 (p-S65-Ub) as a signal for degradation via the autophagy-lysosome system (mitophagy). Despite its discovery in cell culture several years ago, robust and quantitative detection of altered mitophagy in vivo has remained challenging. Here we developed a sandwich ELISA targeting p-S65-Ub with the goal to assess mitophagy levels in mouse brain and in human clinical and pathological samples. We characterized five total Ub and four p-S65-Ub antibodies by several techniques and found significant differences in their ability to recognize phosphorylated Ub. The most sensitive antibody pair detected recombinant p-S65-Ub chains in the femtomolar to low picomolar range depending on the poly-Ub chain linkage. Importantly, this ELISA was able to assess very low baseline mitophagy levels in unstressed human cells and in brains from wild-type and prkn knockout mice as well as elevated p-S65-Ub levels in autopsied frontal cortex from AD patients vs. control cases. Moreover, the assay allowed detection of p-S65-Ub in blood plasma and was able to discriminate between PINK1 mutation carriers and controls. In summary, we developed a robust and sensitive tool to measure mitophagy levels in cells, tissue, and body fluids. Our data strongly support the idea that the stress-activated PINK1-PRKN mitophagy pathway is constitutively active in mice and humans under unstimulated, physiological and elevated in diseased, pathological conditions. Abbreviations: Ab: antibody; AD: Alzheimer disease; AP: alkaline phosphatase; CV: coefficient of variation; ECL: electrochemiluminescence; KO: knockout; LoB: Limit of Blank; LoD: Limit of Detection; LoQ: Limit of Quantification; MSD: meso scale discovery; PD: Parkinson disease; p-S65-PRKN: phosphorylated PRKN at serine 65; p-S65-Ub: phosphorylated ubiquitin at serine 65; Std.Dev.: standard deviation; Ub: ubiquitin; WT: wild type</p
DataSheet1_Plasma PolyQ-ATXN3 Levels Associate With Cerebellar Degeneration and Behavioral Abnormalities in a New AAV-Based SCA3 Mouse Model.pdf
Spinocerebellar ataxia type 3 (SCA3) is a dominantly inherited cerebellar ataxia caused by the expansion of a polyglutamine (polyQ) repeat in the gene encoding ATXN3. The polyQ expansion induces protein inclusion formation in the neurons of patients and results in neuronal degeneration in the cerebellum and other brain regions. We used adeno-associated virus (AAV) technology to develop a new mouse model of SCA3 that recapitulates several features of the human disease, including locomotor defects, cerebellar-specific neuronal loss, polyQ-expanded ATXN3 inclusions, and TDP-43 pathology. We also found that neurofilament light is elevated in the cerebrospinal fluid (CSF) of the SCA3 animals, and the expanded polyQ-ATXN3 protein can be detected in the plasma. Interestingly, the levels of polyQ-ATXN3 in plasma correlated with measures of cerebellar degeneration and locomotor deficits in 6-month-old SCA3 mice, supporting the hypothesis that this factor could act as a biomarker for SCA3.</p