scholarly journals Tracking the evolution of CNS remyelinating lesion in mice with neutral red dye

2019 ◽  
Vol 116 (28) ◽  
pp. 14290-14299 ◽  
Author(s):  
Maryna Baydyuk ◽  
David S. Cha ◽  
Jingwen Hu ◽  
Reiji Yamazaki ◽  
Evan M. Miller ◽  
...  
Keyword(s):  
Neutral Red ◽  
Mass Spectrometry Analysis ◽  
Focal Lesion ◽  
Cns Injury ◽  
Autoimmune Encephalomyelitis ◽  
Spectrometry Analysis ◽  
Immune Mediated ◽  
Demyelinating Lesions ◽  
Demyelinating Disorders ◽  
Red Dye

Animal models of central nervous system (CNS) demyelination, including toxin-induced focal demyelination and immune-mediated demyelination through experimental autoimmune encephalomyelitis (EAE), have provided valuable insights into the mechanisms of neuroinflammation and CNS remyelination. However, the ability to track changes in transcripts, proteins, and metabolites, as well as cellular populations during the evolution of a focal lesion, has remained challenging. Here, we developed a method to label CNS demyelinating lesions by the intraperitoneal injection of a vital dye, neutral red (NR), into mice before killing. We demonstrate that NR-labeled lesions can be easily identified on the intact spinal cord in both lysolecithin- and EAE-mediated demyelination models. Using fluorescence microscopy, we detected NR in activated macrophages/microglia and astrocytes, but not in oligodendrocytes present in lesions. Importantly, we successfully performed RT-qPCR, Western blot, flow cytometry, and mass spectrometry analysis of precisely dissected NR-labeled lesions at 5, 10, and 20 d postlesion (dpl) and found differential changes in transcripts, proteins, cell populations, and metabolites in lesions over the course of remyelination. Therefore, NR administration is a simple and powerful method to track and analyze the detailed molecular, cellular, and metabolic changes that occur within the lesion microenvironment over time following CNS injury. Furthermore, this method can be used to identify molecular and metabolic pathways that regulate neuroinflammation and remyelination and facilitate the development of therapies to promote repair in demyelinating disorders such as multiple sclerosis.

scholarly journals The proteome of remyelination is different from that of developmental myelination

2021 ◽  
Author(s):  
Joana Paes de Faria ◽  
Maria M Azevedo ◽  
Damaris Bausch-Fluck ◽  
Ana I Seixas ◽  
Helena S Domingues ◽  
...  
Keyword(s):  
Mass Spectrometry Analysis ◽  
Myelin Protein ◽  
Oligodendrocyte Progenitor Cells ◽  
Oligodendrocyte Differentiation ◽  
Spectrometry Analysis ◽  
Potential Contributor ◽  
The Difference ◽  
Demyelinating Disorders ◽  
Cytoskeleton Dynamics

Loss of myelin underlies the pathology of several neurological disorders of diverse etiology. CNS remyelination by adult oligodendrocyte progenitor cells (OPCs) can occur but it differs from developmental myelination carried out by neonatal OPCs. We asked whether the myelin proteome of remyelinated regions is changed. We compared the myelin proteome formed during development to the remyelination proteome attained after lysolecithin-induced demyelination in the mouse spinal cord. Mass-spectrometry analysis of iTRAQ labelled myelin protein lysates showed that the proteome of remyelination is different from that of developmental myelination, leading to profound changes in myelin protein content. Aside from known mediators of oligodendrocyte differentiation, we found proteome alterations included modulators of metabolism, cell signaling and actin cytoskeleton dynamics. Downregulating one candidate (FSCN1/Fascin1) was sufficient to partially hamper oligodendrocytes in-vitro. In summary, we identify the difference in the proteome of remyelinating oligodendrocytes as a novel potential contributor to the pathophysiology of demyelinating disorders, thus providing new potential therapeutic targets for future studies.

Spectrophotometry Measurement of Polynuclear Aromatic Hydrocarbons in Hamilton Harbour Sediments

1991 ◽  
Vol 26 (1) ◽  
pp. 1-16 ◽  
Author(s):  
T.P. Murphy ◽  
H. Brouwer ◽  
M.E. Fox ◽  
E. Nagy
Keyword(s):  
Aromatic Hydrocarbons ◽  
Total Concentration ◽  
Coal Tar ◽  
Sediment Cores ◽  
Polynuclear Aromatic Hydrocarbons ◽  
Mass Spectrometry Analysis ◽  
Gas Chromatography Mass Spectrometry ◽  
Near Surface ◽  
Spectrometry Analysis ◽  
Hamilton Harbour

Abstract Eighty-one sediment cores were collected to determine the extent of coal tar contamination in a toxic area of Hamilton Harbour. Over 800 samples were analyzed by a UV spectrophotometric technique that was standardized with gas chromatography/mass spectrometry analysis. The coal tar distribution was variable. The highest concentrations were near the Stelco outfalls and the Hamilton-Wentworth combined sewer outfalls. The total concentration of the 16 polynuclear aromatic hydrocarbons (PAHs) in 48,300 m3 of near-surface sediments exceeded 200 µg/g.

scholarly journals Effect of Ocular Hypertension on D-β-Aspartic Acid-Containing Proteins in the Retinas of Rats

2019 ◽  
Vol 2019 ◽  
pp. 1-8 ◽  
Author(s):  
Takashi Kanamoto ◽  
Takashi Tachibana ◽  
Yasushi Kitaoka ◽  
Toshio Hisatomi ◽  
Yasuhiro Ikeda ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Ocular Hypertension ◽  
Aspartic Acid ◽  
Atp Synthase ◽  
Wistar Rats ◽  
Protein Band ◽  
Mass Spectrometry Analysis ◽  
Liquid Chromatography Mass Spectrometry ◽  
Spectrometry Analysis ◽  
Sds Page

Purpose. To investigate the effect of ocular hypertension-induced isomerization of aspartic acid in retinal proteins. Methods. Adult Wistar rats with ocular hypertension were used as an experimental model. D-β-aspartic acid-containing proteins were isolated by SDS-PAGE and western blot with an anti-D-β-aspartic acid antibody and identified by liquid chromatography-mass spectrometry analysis. The concentration of ATP was measured by ELISA. Results. D-β-aspartic acid was expressed in a protein band at around 44.5 kDa at much higher quantities in the retinas of rats with ocular hypertension than in those of normotensive rats. The 44.5 kDa protein band was mainly composed of α-enolase, S-arrestin, and ATP synthase subunits α and β, in both the ocular hypertensive and normotensive retinas. Moreover, increasing intraocular pressure was correlated with increasing ATP concentrations in the retinas of rats. Conclusion. Ocular hypertension affected the expression of proteins containing D-β-aspartic acid, including ATP synthase subunits, and up-regulation of ATP in the retinas of rats.

scholarly journals Phytochemicals of Minthostachys diffusa Epling and Their Health-Promoting Bioactivities

Foods ◽  
2020 ◽  
Vol 9 (2) ◽  
pp. 144
Author(s):  
Immacolata Faraone ◽  
Daniela Russo ◽  
Lucia Chiummiento ◽  
Eloy Fernandez ◽  
Alka Choudhary ◽  
...  
Keyword(s):  
Ethyl Acetate ◽  
Radical Scavenging ◽  
Ethyl Acetate Fraction ◽  
Mass Spectrometry Analysis ◽  
Antioxidant Power ◽  
Spectrometry Analysis ◽  
In Silico Docking ◽  
Aerial Parts ◽  
Health Promoting

The genus Minthostachys belonging to the Lamiaceae family, and is an important South American mint genus used commonly in folk medicine as an aroma in cooking. The phytochemical-rich samples of the aerial parts of Minthostachys diffusa Epling. were tested for pharmacological and health-promoting bioactivities using in vitro chemical and enzymatic assays. A range of radical scavenging activities of the samples against biological radicals such as nitric oxide and superoxide anion and against synthetic 2,2-diphenyl-1-picrylhydrazyl and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) radicals, the ferric reducing antioxidant power and the lipid peroxidation inhibition were determined and ranked using the ‘relative antioxidant capacity index’ (RACI). The ethyl acetate fraction showed the highest RACI of +1.12. Analysis of the various fractions’ inhibitory ability against enzymes involved in diabetes (α-amylase and α-glucosidase), and against enzymes associated with Parkinson’s or Alzheimer’s diseases (acetylcholinesterase and butyrylcholinesterase) also suggested that the ethyl acetate fraction was the most active. Liquid chromatography–tandem mass spectrometry analysis of the ethyl acetate fraction showed more than 30 polyphenolic compounds, including triterpenes. The inhibitory cholinesterase effects of the triterpenes identified from M. diffusa were further analysed by in silico docking of these compounds into 3D-structures of acetylcholinesterase and butyrylcholinesterase. This is the first study on pharmacological activities and phytochemical profiling of the aerial parts of M. diffusa, showing that this plant, normally used as food in South America, is also rich in health-promoting phytochemicals.

scholarly journals N-Terminomics Strategies for Protease Substrates Profiling

Molecules ◽  
2021 ◽  
Vol 26 (15) ◽  
pp. 4699
Author(s):  
Mubashir Mintoo ◽  
Amritangshu Chakravarty ◽  
Ronak Tilvawala
Keyword(s):  
Mass Spectrometry ◽  
Protease Activity ◽  
Viral Infections ◽  
Mass Spectrometry Analysis ◽  
Post Translational Modification ◽  
Spectrometry Analysis ◽  
Physiological Conditions ◽  
Inflammatory Conditions ◽  
Biological Mixtures

Proteases play a central role in various biochemical pathways catalyzing and regulating key biological events. Proteases catalyze an irreversible post-translational modification called proteolysis by hydrolyzing peptide bonds in proteins. Given the destructive potential of proteolysis, protease activity is tightly regulated. Dysregulation of protease activity has been reported in numerous disease conditions, including cancers, neurodegenerative diseases, inflammatory conditions, cardiovascular diseases, and viral infections. The proteolytic profile of a cell, tissue, or organ is governed by protease activation, activity, and substrate specificity. Thus, identifying protease substrates and proteolytic events under physiological conditions can provide crucial information about how the change in protease regulation can alter the cellular proteolytic landscape. In recent years, mass spectrometry-based techniques called N-terminomics have become instrumental in identifying protease substrates from complex biological mixtures. N-terminomics employs the labeling and enrichment of native and neo-N-termini peptides, generated upon proteolysis followed by mass spectrometry analysis allowing protease substrate profiling directly from biological samples. In this review, we provide a brief overview of N-terminomics techniques, focusing on their strengths, weaknesses, limitations, and providing specific examples where they were successfully employed to identify protease substrates in vivo and under physiological conditions. In addition, we explore the current trends in the protease field and the potential for future developments.

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