Browsing by Author "Ramanadham, Sasanka"

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    A proteomic meta-analysis refinement of plasma extracellular vesicles
    (Springer Nature, 2023-11-28) Vallejo, Milene C.; Sarkar, Soumyadeep; Elliott, Emily C.; Henry, Hayden R.; Powell, Samantha M.; Diaz Ludovico, Ivo; You, Youngki; Huang, Fei; Payne, Samuel H.; Ramanadham, Sasanka; Sims, Emily K.; Metz, Thomas O.; Mirmira, Raghavendra G.; Nakayasu, Ernesto S.; Pediatrics, School of Medicine
    Extracellular vesicles play major roles in cell-to-cell communication and are excellent biomarker candidates. However, studying plasma extracellular vesicles is challenging due to contaminants. Here, we performed a proteomics meta-analysis of public data to refine the plasma EV composition by separating EV proteins and contaminants into different clusters. We obtained two clusters with a total of 1717 proteins that were depleted of known contaminants and enriched in EV markers with independently validated 71% true-positive. These clusters had 133 clusters of differentiation (CD) antigens and were enriched with proteins from cell-to-cell communication and signaling. We compared our data with the proteins deposited in PeptideAtlas, making our refined EV protein list a resource for mechanistic and biomarker studies. As a use case example for this resource, we validated the type 1 diabetes biomarker proplatelet basic protein in EVs and showed that it regulates apoptosis of β cells and macrophages, two key players in the disease development. Our approach provides a refinement of the EV composition and a resource for the scientific community.
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    Extracellular vesicles in β cell biology: Role of lipids in vesicle biogenesis, cargo, and intercellular signaling
    (Elsevier, 2022) Aguirre, Rebecca S.; Kulkarni, Abhishek; Becker, Matthew W.; Lei, Xiaoyong; Sarkar, Soumyadeep; Ramanadham, Sasanka; Phelps, Edward A.; Nakayasu, Ernesto S.; Sims, Emily K.; Mirmira, Raghavendra G.; Pediatrics, School of Medicine
    Background: Type 1 diabetes (T1D) is a complex autoimmune disorder whose pathogenesis involves an intricate interplay between β cells of the pancreatic islet, other islet cells, and cells of the immune system. Direct intercellular communication within the islet occurs via cell surface proteins and indirect intercellular communication has traditionally been seen as occurring via secreted proteins (e.g., endocrine hormones and cytokines). However, recent literature suggests that extracellular vesicles (EVs) secreted by β cells constitute an additional and biologically important mechanism for transmitting signals to within the islet. Scope of review: This review summarizes the general mechanisms of EV formation, with a particular focus on how lipids and lipid signaling pathways influence their formation and cargo. We review the implications of EV release from β cells for T1D pathogenesis, how EVs and their cargo might be leveraged as biomarkers of this process, and how EVs might be engineered as a therapeutic candidate to counter T1D outcomes. Major conclusions: Islet β cells have been viewed as initiators and propagators of the cellular circuit giving rise to autoimmunity in T1D. In this context, emerging literature suggests that EVs may represent a conduit for communication that holds more comprehensive messaging about the β cells from which they arise. As the field of EV biology advances, it opens the possibility that intervening with EV formation and cargo loading could be a novel disease-modifying approach in T1D.
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    Lipid mediators and biomarkers associated with type 1 diabetes development
    (American Society for Clinical Investigation, 2020-08-20) Nelson, Alexander J.; Stephenson, Daniel J.; Bone, Robert N.; Cardona, Christopher L.; Park, Margaret A.; Tusing, Ying G.; Lei, Xiaoyong; Kokotos, George; Graves, Christina L.; Mathews, Clayton E.; Kramer, Joanna; Hessner, Martin J.; Chalfant, Charles E.; Ramanadham, Sasanka; Pediatrics, School of Medicine
    Type 1 diabetes (T1D) is a consequence of autoimmune β cell destruction, but the role of lipids in this process is unknown. We previously reported that activation of Ca2+-independent phospholipase A2β (iPLA2β) modulates polarization of macrophages (MΦ). Hydrolysis of the sn-2 substituent of glycerophospholipids by iPLA2β can lead to the generation of oxidized lipids (eicosanoids), pro- and antiinflammatory, which can initiate and amplify immune responses triggering β cell death. As MΦ are early triggers of immune responses in islets, we examined the impact of iPLA2β-derived lipids (iDLs) in spontaneous-T1D prone nonobese diabetic mice (NOD), in the context of MΦ production and plasma abundances of eicosanoids and sphingolipids. We find that (a) MΦNOD exhibit a proinflammatory lipid landscape during the prediabetic phase; (b) early inhibition or genetic reduction of iPLA2β reduces production of select proinflammatory lipids, promotes antiinflammatory MΦ phenotype, and reduces T1D incidence; (c) such lipid changes are reflected in NOD plasma during the prediabetic phase and at T1D onset; and (d) importantly, similar lipid signatures are evidenced in plasma of human subjects at high risk for developing T1D. These findings suggest that iDLs contribute to T1D onset and identify select lipids that could be targeted for therapeutics and, in conjunction with autoantibodies, serve as early biomarkers of pre-T1D.
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    Polarization of Macrophages toward M2 Phenotype Is Favored by Reduction in iPLA2β (Group VIA Phospholipase A2)
    (American Society for Biochemistry and Molecular Biology, 2016-10-28) Ashley, Jason W.; Hancock, William D.; Nelson, Alexander J.; Bone, Robert N.; Tse, Hubert M.; Wohltmann, Mary; Turk, John; Ramanadham, Sasanka; Medicine, School of Medicine
    Macrophages are important in innate and adaptive immunity. Macrophage participation in inflammation or tissue repair is directed by various extracellular signals and mediated by multiple intracellular pathways. Activation of group VIA phospholipase A2 (iPLA2β) causes accumulation of arachidonic acid, lysophospholipids, and eicosanoids that can promote inflammation and pathologic states. We examined the role of iPLA2β in peritoneal macrophage immune function by comparing wild type (WT) and iPLA2β−/− mouse macrophages. Compared with WT, iPLA2β−/− macrophages exhibited reduced proinflammatory M1 markers when classically activated. In contrast, anti-inflammatory M2 markers were elevated under naïve conditions and induced to higher levels by alternative activation in iPLA2β−/− macrophages compared with WT. Induction of eicosanoid (12-lipoxygenase (12-LO) and cyclooxygenase 2 (COX2))- and reactive oxygen species (NADPH oxidase 4 (NOX4))-generating enzymes by classical activation pathways was also blunted in iPLA2β−/− macrophages compared with WT. The effects of inhibitors of iPLA2β, COX2, or 12-LO to reduce M1 polarization were greater than those to enhance M2 polarization. Certain lipids (lysophosphatidylcholine, lysophosphatidic acid, and prostaglandin E2) recapitulated M1 phenotype in iPLA2β−/− macrophages, but none tested promoted M2 phenotype. These findings suggest that (a) lipids generated by iPLA2β and subsequently oxidized by cyclooxygenase and 12-LO favor macrophage inflammatory M1 polarization, and (b) the absence of iPLA2β promotes macrophage M2 polarization. Reducing macrophage iPLA2β activity and thereby attenuating macrophage M1 polarization might cause a shift from an inflammatory to a recovery/repair milieu.