This chapter provides a detailed description of a method used to study temporal changes in the endoplasmic reticulum (ER) proteome of fibroblast cells exposed to ER stress agents (tunicamycin and thapsigargin). Differential stable isotope labeling by amino acids in cell culture (SILAC) is used in combination with crude ER fractionation, SDS-PAGE and LC-MS/MS to define altered protein expression in tunicamycin or thapsigargin treated cells versus untreated cells. Treated and untreated cells are harvested at different time points, mixed at a 1:1 ratio and processed for ER fractionation. Samples containing labeled and unlabeled proteins are separated by SDS-PAGE, bands are digested with trypsin and the resulting peptides analyzed by LC-MS/MS. Proteins are identified using Bioworks software and the Swiss-Prot database, whereas ratios of protein expression between treated and untreated cells are quantified using ZoomQuant software. Data visualization is facilitated by GeneSpring software.
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http://dx.doi.org/10.1007/978-1-60761-977-2_22 | DOI Listing |
Blood Adv
January 2025
The Ohio State University, Columbus, Ohio, United States.
Hemophagocytic lymphohistiocytosis (HLH) is a rare but aggressive and potentially lethal hyperinflammatory syndrome characterized by pathologic immune activation and excessive production of proinflammatory cytokines leading to tissue damage and multisystem organ failure. There is an urgent need for the discovery of novel targets and development of therapeutic strategies to treat this rare but deadly syndrome. Protein Arginine Methyltransferase 5 (PRMT5) mediates T cell-based inflammatory responses, making it a potential actionable target for the treatment of HLH.
View Article and Find Full Text PDFFree Radic Biol Med
January 2025
Department of Drug and Health Sciences, University of Catania, Catania, Italy; Unit of Neuropharmacology and Translational Neurosciences, Oasi Research Institute-IRCCS, Troina, Italy. Electronic address:
Alzheimer's disease (AD) is a neurodegenerative disorder characterized by cognitive decline and memory loss. A critical aspect of AD pathology is represented by oxidative stress, which significantly contributes to neuronal damage and death. Microglia and astrocytes, the primary glial cells in the brain, are crucial for managing oxidative stress and supporting neuronal function.
View Article and Find Full Text PDFJ Biotechnol
January 2025
Department of Chemical Engineering, University of Massachusetts Lowell, Lowell, MA 01854. Electronic address:
Recombinant adeno-associated viruses (rAAVs) comprise a promising viral vector for therapeutic gene delivery to treat disease. However, the current manufacturing capability of rAAVs must be improved to meet commercial demand. Previously published omics studies indicate that rAAV production through transient transfection triggers antiviral responses and endoplasmic reticulum stress responses in the host cell.
View Article and Find Full Text PDFPLoS One
January 2025
Departments of Microbiology, College of Medicine, Ewha Womans University, Seoul, Korea.
Mast cells, immune sentinels that respond to various stimuli in barrier organs, provide defense by expressing pattern recognition receptors, such as Toll-like receptors (TLRs). They may affect inflammatory responses and wound healing. Here, we investigated the effect of TLR2/6-stimulated mast cells on wound healing in keratinocytes.
View Article and Find Full Text PDFRSC Adv
January 2025
Pharmaceutical Medicinal Chemistry and Drug Design Department, Faculty of Pharmacy (Girls), Al-Azhar University Cairo 11754 Egypt
The current work focuses on the creation of novel derivatives of the quinazolinone ring system, with various substituted thiophene, thienopyrimidine, and thienopyridine scaffolds 3a,b-11. Employing the standard MTT assay, every target compound's antiproliferative efficacy was evaluated in comparison with doxorubicin against both normal WI-38 cells and various cancer cell lines. Derivatives 6, 8a, and 8b demonstrated the most potent activity, alongside their safety profiles against WI-38.
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