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Ca2+-independent phospholipase A2 (GVIA iPLA2) has gained increasing interest recently as it has been recognized as a participant in biological processes underlying diabetes development and autoimmune-based neurological disorders. The development of potent GVIA iPLA2 inhibitors is of great importance because only a few have been reported so far. We present a novel class of GVIA iPLA2 inhibitors based on the β-lactone ring. This functionality in combination with a four-carbon chain carrying a phenyl group at position-3 and a linear propyl group at position-4 of the lactone ring confers excellent potency. trans-3-(4-Phenylbutyl)-4-propyloxetan-2-one (GK563) was identified as being the most potent GVIA iPLA2 inhibitor ever reported ( XI(50) 0.0000021, IC50 1 nM) and also one that is 22 000 times more active against GVIA iPLA2 than GIVA cPLA2. It was found to reduce β-cell apoptosis induced by proinflammatory cytokines, raising the possibility that it can be beneficial in countering autoimmune diseases, such as type 1 diabetes.

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. 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. 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. [ABSTRACT FROM AUTHOR]

Type 1 diabetes (T1D) development, in part, is due to ER stress-induced β-cell apoptosis. Activation of the Ca2+-independent phospholipase A2 beta (iPLA2β) leads to the generation of pro-inflammatory eicosanoids, which contribute to β-cell death and T1D. ER stress induces iPLA2β-mediated generation of pro-apoptotic ceramides via neutral sphingomyelinase (NSMase). To gain a better understanding of the impact of iPLA2β on sphingolipids (SLs), we characterized their profile in β-cells undergoing ER stress. ESI/MS/MS analyses followed by ANOVA/Student's t-test were used to assess differences in sphingolipids molecular species in Vector (V) control and iPLA2β-overexpressing (OE) INS-1 and Akita (AK, spontaneous model of ER stress) and WT-littermate (AK-WT) β-cells. As expected, iPLA2β induction was greater in the OE and AK cells in comparison with V and WT cells. We report here that ER stress led to elevations in pro-apoptotic and decreases in pro-survival sphingolipids and that the inactivation of iPLA2β restores the sphingolipid species toward those that promote cell survival. In view of our recent finding that the SL profile in macrophages—the initiators of autoimmune responses leading to T1D—is not significantly altered during T1D development, we posit that the iPLA2β-mediated shift in the β-cell sphingolipid profile is an important contributor to β-cell death associated with T1D. [ABSTRACT FROM AUTHOR]

We previously reported that a bromoenol lactone (BEL) suicide inhibitor of the Group VIA Ca[sup 2+]-independent phospholipase A[sub 2] (iPLA[sub 2]β) suppresses ER stress-induced β-cell apoptosis, suggesting that iPLA[sub 2]β plays a role in this process. To gain a better understanding of the mechanism of involvement of iPLA[sub 2]β during ER stress, stable isotope labeling with amino acids in cell culture (SILAC) experiment were performed in empty-vector (V) transfected and iPLA[sub 2]β-overexpressing (OE) cells INS-1 β-cells. cells cultured in isotope-labeled (heavy) amino acids medium were treated with DMSO vehicle and Cells cultured in regular media were treated with thapsigargin (1 µM) to induce ER stress. After 24h, the vehicle- and thapsigargin-treated homogenate protein from each were combined in a 1:1 ratio, separated by SDS-PAGE (7.5%), digested with trypsin, and analyzed by LC/MS/MS mass spectrometry. ER stress induced no change in tubulin in either V or OE INS-1 cells, an increase in iPLA[sub 2]β in the OE cells, and dysregulation of several cytosolic-, mitochondrial-, and ER-associated proteins that participate in apoptosis in both V and OE INS-1 cells. While ubiquitin-like 1 activating enzyme E1A was up-regulated less in the OE than in the V INS-1 cells; GRP 78, Trap 1, Erp 61, hypoxia up-regulated 1, and UDP-glucose ceramide glucosyltransferase-like 1 were down-regulated more in the OE than in the V INS-1 cells. Thus, the SILAC protocol allowed identification of differentially-regulated proteins in β-cells undergoing ER stress, generation a dataset of proteins that might participate in ER stress-induced β-cell apoptosis, and assessment of the role of iPLA[sub 2]β in this process. Our findings indicate that induction of ER stress in β-cells leads to recruitment of the mitochondria and that both organelles, in concert with iPLA[sub 2]β, contribute to ER stress-induced β-cell apoptosis. [ABSTRACT FROM AUTHOR]

In type I diabetes, beta cell death can result from cytokine-induced nitric oxide (NO) production and apoptosis, iPLA2γ is a recently recognized Group VI phospholipase A2 that is expressed in mitochondria and other subcellular loci, and it has been suggested to participate in the maintenance of mitochondrial membrane integrity. Here we demonstrate that overexpression of iPLA2γ in INS-1 insulinoma cells (OE cells) protects against apoptosis induced by the NO donor S-nitroso-N-acetylpenicillamine (SNAP), as assessed by TUNEL staining up to 11.5±2.5% of control INS-1 cells experienced apoptosis after exposure to SNAP for 16 hr, but only 5.6±1.4% (p<0.01) of OE cells did so. In addition, fewer OE cells than control cells experienced collapse of the mitochondrial membrane potential after SNAP exposure. SNAP strongly induced expression of the ER stress marker CHOP in control INS-1 cells before onset of apoptosis, and these responses were greatly blunted in OE cells. No differences between control and OE cells were observed with respect to activation of the Akt or GSK3ß signaling pathways induced by SNAP exposure. These findings suggest that iPLA2γ might act to reduce both mitochondrial injury upon exposure to NO and the associated generation of signals that induce ER stress and that these actions of iPLA2γ do not involve the Akt or GSK3B signaling pathways. [ABSTRACT FROM AUTHOR]

Abstract The Ca2+-independent phospholipases, designated as group VI iPLA 2 s, also referred to as PNPLAs due to their shared homology with patatin, include the β, γ, δ, ε, ζ, and η forms of the enzyme. The iPLA 2 s are ubiquitously expressed, share a consensus GXSXG catalytic motif, and exhibit organelle/cell-specific localization. Among the iPLA 2 s, iPLA 2 β has received wide attention as it is recognized to be involved in membrane remodeling, cell proliferation, cell death, and signal transduction. Ongoing studies implicate participation of iPLA 2 β in a variety of disease processes including cancer, cardiovascular abnormalities, glaucoma, and peridonditis. This review will focus on iPLA 2 β and its links to male fertility, neurological disorders, metabolic disorders, and inflammation. [ABSTRACT FROM AUTHOR]