The Ca 2+ -independent phospholipases A 2 (iPLA 2 s) are part of a diverse family of PLA 2 s that hydrolyze the sn -2 substituent from membrane phospholipids to release a free fatty acid and a lysolipid ( 1, 2 ). These enzymes are ubiquitously expressed, and in contrast to secretory PLA 2 s (sPLA 2 s) and cytosolic PLA 2 s (cPLA 2 s), do not require Ca 2+ for either translocation or activity. Some of the fi rst descriptions of iPLA 2 activity were in the mid-to late-1980s with the identifi cation of a plasmalogen-selective PLA 2 in Abstract Among the family of phospholipases A 2 (PLA 2 s) are the Ca 2+ -independent PLA 2 s (iPLA 2 s) and they are designated group VI iPLA 2 s. In relation to secretory and cytosolic PLA 2 s, the iPLA 2 s are more recently described and details of their expression and roles in biological functions are rapidly emerging. The iPLA 2 s or patatin-like phospholipases (PNPLAs) are intracellular enzymes that do not require Ca 2+ for activity, and contain lipase (GXSXG) and nucleotide-binding (GXGXXG) consensus sequences. Though nine PNPLAs have been recognized, PNPLA8 (membraneassociated iPLA 2 ␥ ) and PNPLA9 (cytosol-associated iPLA 2 ␤ ) are the most widely studied and understood. The iPLA 2 s manifest a variety of activities in addition to phospholipase, are ubiquitously expressed, and participate in a multitude of biological processes, including fat catabolism, cell differentiation, maintenance of mitochondrial integrity, phospholipid remodeling, cell proliferation, signal transduction, and cell death. As might be expected, increased or decreased expression of iPLA 2 s can have profound effects on the metabolic state, CNS function, cardiovascular performance, and cell survival; therefore, dysregulation of iPLA 2 s can be a critical factor in the development of many diseases. This review is aimed at providing a general framework of the current understanding of the iPLA 2 s and discussion of the potential mechanisms of action of the iPLA 2 s and related involved lipid mediators. -Ramanadham, S

Edited by Jeffrey E. Pessin Alterations in endoplasmic reticulum (ER) calcium (Ca 2؉) levels diminish insulin secretion and reduce ␤-cell survival in both major forms of diabetes. The mechanisms responsible for ER Ca 2؉ loss in ␤ cells remain incompletely understood. Moreover, a specific role for either ryanodine receptor (RyR) or inositol 1,4,5-triphosphate receptor (IP 3 R) dysfunction in the pathophysiology of diabetes remains largely untested. To this end, here we applied intracellular and ER Ca 2؉ imaging techniques in INS-1 ␤ cells and isolated islets to determine whether diabetogenic stressors alter RyR or IP 3 R function. Our results revealed that the RyR is sensitive mainly to ER stress-induced dysfunction, whereas cytokine stress specifically alters IP 3 R activity. Consistent with this observation, pharmacological inhibition of the RyR with ryanodine and inhibition of the IP 3 R with xestospongin C prevented ER Ca 2؉ loss under ER and cytokine stress conditions, respectively. However, RyR blockade distinctly prevented ␤-cell death, propagation of the unfolded protein response (UPR), and dysfunctional glucose-induced Ca 2؉ oscillations in tunicamycin-treated INS-1 ␤ cells and mouse islets and Akita islets. Monitoring at the single-cell level revealed that ER stress acutely increases the frequency of intracellular Ca 2؉ transients that depend on both ER Ca 2؉ leakage from the RyR and plasma membrane depolarization. Collectively, these findings indicate that RyR dysfunction shapes ER Ca 2؉ dynamics in ␤ cells and regulates both UPR activation and cell death, suggesting that RyR-mediated loss of ER Ca 2؉ may be an early pathogenic event in diabetes. This work was supported by National Institutes of Health Grants R01 DK093954 and UC4 DK 104166 (to C. E-M.); Department of Veterans Affairs Merit Award I01BX001733 (to C. E-M.); and Sigma Beta Sorority, Ball Brothers Foundation, and George and Frances Ball Foundation gifts (to C. E-M.).

Store-operated Ca entry (SOCE) is a dynamic process that leads to refilling of endoplasmic reticulum (ER) Ca stores through reversible gating of plasma membrane Ca channels by the ER Ca sensor Stromal Interaction Molecule 1 (STIM1). Pathogenic reductions in β-cell ER Ca have been observed in diabetes. However, a role for impaired SOCE in this phenotype has not been tested. We measured the expression of SOCE molecular components in human and rodent models of diabetes and found a specific reduction in STIM1 mRNA and protein levels in human islets from donors with type 2 diabetes (T2D), islets from hyperglycemic streptozotocin-treated mice, and INS-1 cells (rat insulinoma cells) treated with proinflammatory cytokines and palmitate. Pharmacologic SOCE inhibitors led to impaired islet Ca oscillations and insulin secretion, and these effects were phenocopied by β-cell STIM1 deletion. STIM1 deletion also led to reduced ER Ca storage and increased ER stress, whereas STIM1 gain of function rescued β-cell survival under proinflammatory conditions and improved insulin secretion in human islets from donors with T2D. Taken together, these data suggest that the loss of STIM1 and impaired SOCE contribute to ER Ca dyshomeostasis under diabetic conditions, whereas efforts to restore SOCE-mediated Ca transients may have the potential to improve β-cell health and function.

Autoimmune β-cell death leads to type 1 diabetes, and with findings that Ca2+-independent phospholipase A2β (iPLA2β) activation contributes to β-cell death, we assessed the effects of iPLA2β inhibition on diabetes development. Administration of FKGK18, a reversible iPLA2β inhibitor, to NOD female mice significantly reduced diabetes incidence in association with 1) reduced insulitis, reflected by reductions in CD4+ T cells and B cells; 2) improved glucose homeostasis; 3) higher circulating insulin; and 4) β-cell preservation. Furthermore, FKGK18 inhibited production of tumor necrosis factor-α (TNF-α) from CD4+ T cells and antibodies from B cells, suggesting modulation of immune cell responses by iPLA2β-derived products. Consistent with this, 1) adoptive transfer of diabetes by CD4+ T cells to immunodeficient and diabetes-resistant NOD.scid mice was mitigated by FKGK18 pretreatment and 2) TNF-α production from CD4+ T cells was reduced by inhibitors of cyclooxygenase and 12-lipoxygenase, which metabolize arachidonic acid to generate bioactive inflammatory eicosanoids. However, adoptive transfer of diabetes was not prevented when mice were administered FKGK18-pretreated T cells or when FKGK18 administration was initiated with T-cell transfer. The present observations suggest that iPLA2β-derived lipid signals modulate immune cell responses, raising the possibility that early inhibition of iPLA2β may be beneficial in ameliorating autoimmune destruction of β-cells and mitigating type 1 diabetes development.

Type 1 diabetes (T1D) results from autoimmune destruction of islet ␤-cells, but the underlying mechanisms that contribute to this process are incompletely understood, especially the role of lipid signals generated by ␤-cells. Proinflammatory cytokines induce ER stress in ␤-cells and we previously found that the Ca 2ϩ -independent phospholipase A 2 ␤ (iPLA 2 ␤) participates in ER stress-induced ␤-cell apoptosis. In view of reports of elevated iPLA 2 ␤ in T1D, we examined if iPLA 2 ␤ participates in cytokine-mediated islet ␤-cell apoptosis. We find that the proinflammatory cytokine combination IL-1␤ϩIFN␥, induces: a) ER stress, mSREBP-1, and iPLA 2 ␤, b) lysophosphatidylcholine (LPC) generation, c) neutral sphingomyelinase-2 (NSMase2), d) ceramide accumulation, e) mitochondrial membrane decompensation, f) caspase-3 activation, and g) ␤-cell apoptosis. The presence of a sterol regulatory element in the iPLA 2 ␤ gene raises the possibility that activation of SREBP-1 after proinflammatory cytokine exposure contributes to iPLA 2 ␤ induction. The IL-1␤ϩIFN␥-induced outcomes (b-g) are all inhibited by iPLA 2 ␤ inactivation, suggesting that iPLA 2 ␤-derived lipid signals contribute to consequential islet ␤-cell death. Consistent with this possibility, ER stress and ␤-cell apoptosis induced by proinflammatory cytokines are exacerbatedinisletsfromRIP-iPLA 2 ␤-TgmiceandbluntedinisletsfromiPLA 2 ␤-KOmice.Theseobservations suggest that iPLA 2 ␤-mediated events participate in amplifying ␤-cell apoptosis due to proinflammatory cytokines and also that iPLA 2 ␤ activation may have a reciprocal impact on ER stress development. They raise thepossibilitythatiPLA 2 ␤inhibition,leadingtoameliorationsinERstress,apoptosis,andimmuneresponses resulting from LPC-stimulated immune cell chemotaxis, may be beneficial in preserving ␤-cell mass and delaying/preventing T1D evolution. Abbreviations: ATF6, activating transcription factor 6; S-BEL, bromoenol lactone suicide inhibitor of iPLA 2 ␤; CTK, IL-1␤ϩIFN␥; DAPI, 4Ј,6Ј-diamidino-2-phenylindole; DMSO, dimethyl sulfoxide; ER, endoplasmic reticulum; ESI, electrospray ionization; GRP78, 78 kDa glucose-regulated protein; GSH, glutathione; iNOS, inducible nitric oxide synthase; iPLA 2 ␤, ␤-isoform of group VIA calcium-independent phospholipase A 2 ; iPLA 2 ␤-KO, globally iPLA 2 ␤-deficient; IRE1, inositol-requiring enzyme 1; KO, knockout; LPC, lysophosphatidylcholine; ⌬⌿, mitochondrial membrane potential; MS, mass spectrometry; mSREBP-1, mature form of sterol regulatory element binding protein-1; NOD, non-obese diabetic; NSMase2, neutral sphingomyelinase-2; pPERK, phosphorylated form of ER-stress transducer pancreatic ER kinase; PLA 2 , phospholipase A 2 ; pNA, p-nitroaniline; RIP-iPLA 2 ␤-Tg, iPLA 2 ␤ overexpressed in only ␤-cells; ROS, reactive oxygen species; RT-qPCR, quantitative RT-PCR; SPT1, serine palmitoyl transferase; T1D, type 1 diabetes; TUNEL, terminal deoxynucleotidyl transferase dUTP nick end labeling; WT, wild-type.