Differential lipid signaling from CD4+ and CD8+ T cells contributes to type 1 diabetes development.

Introduction: We reported that Ca2+-independent phospholipase A2ß (iPLA2ß)-derived lipids (iDLs) contribute to type 1 diabetes (T1D) onset. As CD4+ and CD8+ T cells are critical in promoting ß-cell death, we tested the hypothesis that iDL signaling from these cells participates in T1D development. Methods: CD4+ and CD8+ T cells from wild-type non-obese diabetic (NOD) and NOD.iPLA2ß+/- (NOD.HET) mice were administered in different combinations to immunodeficient NOD.scid. Results: In mice receiving only NOD T cells, T1D onset was rapid (5 weeks), incidence 100% by 20 weeks, and islets absent. In contrast, onset was delayed 1 week and incidence reduced 40%-50% in mice receiving combinations that included NOD.HET T cells. Consistently, islets from these non-diabetic mice were devoid of infiltrate and contained insulin-positive ß-cells. Reduced iPLA2ß led to decreased production of proinflammatory lipids from CD4+ T cells including prostaglandins and dihydroxyeicosatrienoic acids (DHETs), products of soluble epoxide hydrolase (sEH), and inhibition of their signaling decreased (by 82%) IFNγ+CD4+ cells abundance. However, only DHETs production was reduced from CD8+ T cells and was accompanied by decreases in sEH and granzyme B. Discussion: These findings suggest that differential select iDL signaling in CD4+ and CD8+ T cells contributes to T1D development, and that therapeutics targeting such signaling might be considered to counter T1D.

Selective Reduction of Ca2+-Independent Phospholipase A2ß (iPLA2ß)-Derived Lipid Signaling from Macrophages Mitigates Type 1 Diabetes Development.

Type 1 diabetes (T1D) is a consequence of autoimmune destruction of ß-cells and macrophages (MΦ) have a central role in initiating processes that lead to ß-cell demise. We reported that Ca2+-independent phospholipase A2ß (iPLA2ß)-derived lipid (iDL) signaling contributes to ß-cell death. As MΦ express iPLA2ß, we assessed its role in T1D development. We find that selective reduction of myeloid-iPLA2ß in spontaneously diabetes-prone nonobese diabetic (NOD) mice (a) deceases proinflammatory eicosanoid production by MΦ, (b) favors anti-inflammatory (M2-like) MΦ phenotype, and (c) diminishes activated CD4+ and CD8+ T-cells phenotype in the pancreatic infiltrate, prior to T1D onset. These outcomes are associated with a significant reduction in T1D. Further, inhibition of select proinflammatory lipid signaling pathways reduces M1-like MΦ polarization and adoptive transfer of M2-like MΦ reduces NOD T1D incidence, suggesting a mechanism by which iDLs impact T1D development. These findings identify MΦ-iPLA2ß as a critical contributor to TID development and potential target to counter T1D onset.

Sphingolipid changes in mouse brain and plasma after mild traumatic brain injury at the acute phases.

BACKGROUND: Traumatic brain injury (TBI) causes neuroinflammation and can lead to long-term neurological dysfunction, even in cases of mild TBI (mTBI). Despite the substantial burden of this disease, the management of TBI is precluded by an incomplete understanding of its cellular mechanisms. Sphingolipids (SPL) and their metabolites have emerged as key orchestrators of biological processes related to tissue injury, neuroinflammation, and inflammation resolution. No study so far has investigated comprehensive sphingolipid profile changes immediately following TBI in animal models or human cases. In this study, sphingolipid metabolite composition was examined during the acute phases in brain tissue and plasma of mice following mTBI. METHODS: Wildtype mice were exposed to air-blast-mediated mTBI, with blast exposure set at 50-psi on the left cranium and 0-psi designated as Sham. Sphingolipid profile was analyzed in brain tissue and plasma during the acute phases of 1, 3, and 7 days post-TBI via liquid-chromatography-mass spectrometry. Simultaneously, gene expression of sphingolipid metabolic markers within brain tissue was analyzed using quantitative reverse transcription-polymerase chain reaction. Significance (P-values) was determined by non-parametric t-test (Mann-Whitney test) and by Tukey's correction for multiple comparisons. RESULTS: In post-TBI brain tissue, there was a significant elevation of 1) acid sphingomyelinase (aSMase) at 1- and 3-days, 2) neutral sphingomyelinase (nSMase) at 7-days, 3) ceramide-1-phosphate levels at 1 day, and 4) monohexosylceramide (MHC) and sphingosine at 7-days. Among individual species, the study found an increase in C18:0 and a decrease in C24:1 ceramides (Cer) at 1 day; an increase in C20:0 MHC at 3 days; decrease in MHC C18:0 and increase in MHC C24:1, sphingomyelins (SM) C18:0, and C24:0 at 7 days. Moreover, many sphingolipid metabolic genes were elevated at 1 day, followed by a reduction at 3 days and an absence at 7-days post-TBI. In post-TBI plasma, there was 1) a significant reduction in Cer and MHC C22:0, and an increase in MHC C16:0 at 1 day; 2) a very significant increase in long-chain Cer C24:1 accompanied by significant decreases in Cer C24:0 and C22:0 in MHC and SM at 3 days; and 3) a significant increase of C22:0 in all classes of SPL (Cer, MHC and SM) as well as a decrease in Cer C24:1, MHC C24:1 and MHC C24:0 at 7 days. CONCLUSIONS: Alterations in sphingolipid metabolite composition, particularly sphingomyelinases and short-chain ceramides, may contribute to the induction and regulation of neuroinflammatory events in the early stages of TBI, suggesting potential targets for novel diagnostic, prognostic, and therapeutic strategies in the future.

Sphingolipid biosynthetic inhibitor L-Cycloserine prevents oxidative-stress-mediated death in an in vitro model of photoreceptor-derived 661W cells.

Oxidative stress plays a pivotal role in the pathogenesis of several neurodegenerative diseases. Retinal degeneration causes irreversible death of photoreceptor cells, ultimately leading to vision loss. Under oxidative stress, the synthesis of bioactive sphingolipid ceramide increases, triggering apoptosis in photoreceptor cells and leading to their death. This study investigates the effect of L-Cycloserine, a small molecule inhibitor of ceramide biosynthesis, on sphingolipid metabolism and the protection of photoreceptor-derived 661W cells from oxidative stress. The results demonstrate that treatment with L-Cycloserine, an inhibitor of Serine palmitoyl transferase (SPT), markedly decreases bioactive ceramide and associated sphingolipids in 661W cells. A nontoxic dose of L-Cycloserine can provide substantial protection of 661W cells against H2O2-induced oxidative stress by reversing the increase in ceramide level observed under oxidative stress conditions. Analysis of various antioxidant, apoptotic and sphingolipid pathway genes and proteins also confirms the ability of L-Cycloserine to modulate these pathways. Our findings elucidate the generation of sphingolipid mediators of cell death in retinal cells under oxidative stress and the potential of L-Cycloserine as a therapeutic candidate for targeting ceramide-induced degenerative diseases by inhibiting SPT. The promising therapeutic prospect identified in our findings lays the groundwork for further validation in in-vivo and preclinical models of retinal degeneration.

Ceramide-1-phosphate is a regulator of Golgi structure and is co-opted by the obligate intracellular bacterial pathogen Anaplasma phagocytophilum.

Many intracellular pathogens structurally disrupt the Golgi apparatus as an evolutionarily conserved promicrobial strategy. Yet, the host factors and signaling processes involved are often poorly understood, particularly for Anaplasma phagocytophilum, the agent of human granulocytic anaplasmosis. We found that A. phagocytophilum elevated cellular levels of the bioactive sphingolipid, ceramide-1-phosphate (C1P), to promote Golgi fragmentation that enables bacterial proliferation, conversion from its non-infectious to infectious form, and productive infection. A. phagocytophilum poorly infected mice deficient in ceramide kinase, the Golgi-localized enzyme responsible for C1P biosynthesis. C1P regulated Golgi morphology via activation of a PKCα/Cdc42/JNK signaling axis that culminates in phosphorylation of Golgi structural proteins, GRASP55 and GRASP65. siRNA-mediated depletion of Cdc42 blocked A. phagocytophilum from altering Golgi morphology, which impaired anterograde trafficking of trans-Golgi vesicles into and maturation of the pathogen-occupied vacuole. Cells overexpressing phosphorylation-resistant versions of GRASP55 and GRASP65 presented with suppressed C1P- and A. phagocytophilum-induced Golgi fragmentation and poorly supported infection by the bacterium. By studying A. phagocytophilum, we identify C1P as a regulator of Golgi structure and a host factor that is relevant to disease progression associated with Golgi fragmentation.IMPORTANCECeramide-1-phosphate (C1P), a bioactive sphingolipid that regulates diverse processes vital to mammalian physiology, is linked to disease states such as cancer, inflammation, and wound healing. By studying the obligate intracellular bacterium Anaplasma phagocytophilum, we discovered that C1P is a major regulator of Golgi morphology. A. phagocytophilum elevated C1P levels to induce signaling events that promote Golgi fragmentation and increase vesicular traffic into the pathogen-occupied vacuole that the bacterium parasitizes. As several intracellular microbial pathogens destabilize the Golgi to drive their infection cycles and changes in Golgi morphology is also linked to cancer and neurodegenerative disorder progression, this study identifies C1P as a potential broad-spectrum therapeutic target for infectious and non-infectious diseases.

Mouse Model of Nitrogen Mustard Ocular Surface Injury Characterization and Sphingolipid Signaling.

Vesicating chemicals like sulfur mustard (SM) or nitrogen mustard (NM) can cause devastating damage to the eyes, skin, and lungs. Eyes, being the most sensitive, have complicated pathologies that can manifest immediately after exposure (acute) and last for years (chronic). No FDA-approved drug is available to be used as medical counter measures (MCMs) against such injuries. Understanding the pathological mechanisms in acute and chronic response of the eye is essential for developing effective MCMs. Here, we report the clinical and histopathological characterization of a mouse model of NM-induced ocular surface injury (entire surface) developed by treating the eye with 2% (w/v) NM solution for 5 min. Unlike the existing models of specific injury, our model showed severe ocular inflammation, including the eyelids, structural deformity of the corneal epithelium and stroma, and diminished visual and retinal functions. We also observed alterations of the inflammatory markers and their expression at different phases of the injury, along with an activation of acidic sphingomyelinase (aSMase), causing an increase in bioactive sphingolipid ceramide and a reduction in sphingomyelin levels. This novel ocular surface mouse model recapitulated the injuries reported in human, rabbit, and murine SM or NM injury models. NM exposure of the entire ocular surface in mice, which is similar to accidental or deliberate exposure in humans, showed severe ocular inflammation and caused irreversible alterations to the corneal structure and significant vision loss. It also showed an intricate interplay between inflammatory markers over the injury period and alteration in sphingolipid homeostasis in the early acute phase.

Fluoxetine restrains allergic inflammation by targeting an FcɛRI-ATP positive feedback loop in mast cells.

There is a clinical need for new treatment options addressing allergic disease. Selective serotonin reuptake inhibitors (SSRIs) are a class of antidepressants that have anti-inflammatory properties. We tested the effects of the SSRI fluoxetine on IgE-induced function of mast cells, which are critical effectors of allergic inflammation. We showed that fluoxetine treatment of murine or human mast cells reduced IgE-mediated degranulation, cytokine production, and inflammatory lipid secretion, as well as signaling mediated by the mast cell activator ATP. In a mouse model of systemic anaphylaxis, fluoxetine reduced hypothermia and cytokine production. Fluoxetine was also effective in a model of allergic airway inflammation, where it reduced bronchial responsiveness and inflammation. These data show that fluoxetine suppresses mast cell activation by impeding an FcɛRI-ATP positive feedback loop and support the potential repurposing of this SSRI for use in allergic disease.

Skewing cPLA2α activity toward oxoeicosanoid production promotes neutrophil N2 polarization, wound healing, and the response to sepsis.

Uncontrolled inflammation is linked to poor outcomes in sepsis and wound healing, both of which proceed through distinct inflammatory and resolution phases. Eicosanoids are a class of bioactive lipids that recruit neutrophils and other innate immune cells. The interaction of ceramide 1-phosphate (C1P) with the eicosanoid biosynthetic enzyme cytosolic phospholipase A2 (cPLA2) reduces the production of a subtype of eicosanoids called oxoeicosanoids. We investigated the effect of shifting the balance in eicosanoid biosynthesis on neutrophil polarization and function. Knockin mice expressing a cPLA2 mutant lacking the C1P binding site (cPLA2αKI/KI mice) showed enhanced and sustained neutrophil infiltration into wounds and the peritoneum during the inflammatory phase of wound healing and sepsis, respectively. The mice exhibited improved wound healing and reduced susceptibility to sepsis, which was associated with an increase in anti-inflammatory N2-type neutrophils demonstrating proresolution behaviors and a decrease in proinflammatory N1-type neutrophils. The N2 polarization of cPLA2αKI/KI neutrophils resulted from increased oxoeicosanoid biosynthesis and autocrine signaling through the oxoeicosanoid receptor OXER1 and partially depended on OXER1-dependent inhibition of the pentose phosphate pathway (PPP). Thus, C1P binding to cPLA2α suppresses neutrophil N2 polarization, thereby impairing wound healing and the response to sepsis.

Bioactive lipid mediators in plasma are predictors of preeclampsia irrespective of aspirin therapy.

There are few early biomarkers to identify pregnancies at risk of preeclampsia (PE) and abnormal placental function. In this cross-sectional study, we utilized targeted ultra-performance liquid chromatography-ESI MS/MS and a linear regression model to identify specific bioactive lipids that serve as early predictors of PE. Plasma samples were collected from 57 pregnant women prior to 24-weeks of gestation with outcomes of either PE (n = 26) or uncomplicated term pregnancies (n = 31), and the profiles of eicosanoids and sphingolipids were evaluated. Significant differences were revealed in the eicosanoid, (±)11,12 DHET, as well as multiple classes of sphingolipids; ceramides, ceramide-1-phosphate, sphingomyelin, and monohexosylceramides; all of which were associated with the subsequent development of PE regardless of aspirin therapy. Profiles of these bioactive lipids were found to vary based on self-designated race. Additional analyses demonstrated that PE patients can be stratified based on the lipid profile as to PE with a preterm birth linked to significant differences in the levels of 12-HETE, 15-HETE, and resolvin D1. Furthermore, subjects referred to a high-risk OB/GYN clinic had higher levels of 20-HETE, arachidonic acid, and Resolvin D1 versus subjects recruited from a routine, general OB/GYN clinic. Overall, this study shows that quantitative changes in plasma bioactive lipids detected by ultra-performance liquid chromatography-ESI-MS/MS can serve as an early predictor of PE and stratify pregnant people for PE type and risk.

Ceramide Kinase Inhibition Drives Ferroptosis and Sensitivity to Cisplatin in Mutant KRAS Lung Cancer by Dysregulating VDAC-Mediated Mitochondria Function.

Ceramide kinase (CERK) is the mammalian lipid kinase from which the bioactive sphingolipid, ceramide-1-phosphate (C1P), is derived. CERK has been implicated in several promalignant phenotypes with little known as to mechanistic underpinnings. In this study, the mechanism of how CERK inhibition decreases cell survival in mutant (Mut) KRAS non-small cell lung cancer (NSCLC), a major lung cancer subtype, was revealed. Specifically, NSCLC cells possessing a KRAS mutation were more responsive to inhibition, downregulation, and genetic ablation of CERK compared with those with wild-type (WT) KRAS regarding a reduction in cell survival. Inhibition of CERK induced ferroptosis in Mut KRAS NSCLC cells, which required elevating VDAC-regulated mitochondria membrane potential (MMP) and the generation of cellular reactive oxygen species (ROS). Importantly, through modulation of VDAC, CERK inhibition synergized with the first-line NSCLC treatment, cisplatin, in reducing cell survival and in vivo tumor growth. Further mechanistic studies indicated that CERK inhibition affected MMP and cell survival by limiting AKT activation and translocation to mitochondria, and thus, blocking VDAC phosphorylation and tubulin recruitment. IMPLICATIONS: Our findings depict how CERK inhibition may serve as a new key point in combination therapeutic strategy for NSCLC, specifically precision therapeutics targeting NSCLC possessing a KRAS mutation.

Hydroxychloroquine Causes Early Inner Retinal Toxicity and Affects Autophagosome-Lysosomal Pathway and Sphingolipid Metabolism in the Retina.

Hydroxychloroquine (HCQ) is an anti-malarial drug but also widely used to treat autoimmune diseases like arthritis and lupus. Although there have been multiple reports of the adverse effect of prolonged HCQ usage on the outer retina, leading to bull's-eye maculopathy, the effect of HCQ toxicity on the inner retina as well as on overall visual functions has not been explored in detail. Furthermore, lack of an established animal model of HCQ toxicity hinders our understanding of the underlying molecular mechanisms. Here, using a small clinical study, we confirmed the effect of HCQ toxicity on the inner retina, in particular the reduction in central inner retinal thickness, and established a mouse model of chronic HCQ toxicity that recapitulates the effects observed in human retina. Using the mouse model, we demonstrated that chronic HCQ toxicity results in loss of inner retinal neurons and retinal ganglion cells (RGC) and compromises visual functions. We further established that HCQ treatment prevents autophagosome-lysosome fusion and alters the sphingolipid homeostasis in mouse retina. Our results affirm the notion that HCQ treatment causes early damage to the inner retina and affects visual functions before leading to characteristic toxicity in the macular region of the outer retina, 'bull's-eye maculopathy.' We also provide insights into the underlying molecular mechanisms of HCQ retinal toxicity that may involve autophagy-lysosomal defects and alterations in sphingolipid metabolism.

Ceramide kinase regulates acute wound healing by suppressing 5-oxo-ETE biosynthesis and signaling via its receptor OXER1.

The sphingolipid, ceramide-1-phosphate (C1P), has been shown to promote the inflammatory phase and inhibit the proliferation and remodeling stages of wound repair via direct interaction with group IVA cytosolic phospholipase A2, a regulator of eicosanoid biosynthesis that fine-tunes the behaviors of various cell types during wound healing. However, the anabolic enzyme responsible for the production of C1P that suppresses wound healing as well as bioactive eicosanoids and target receptors that drive enhanced wound remodeling have not been characterized. Herein, we determined that decreasing C1P activity via inhibitors or genetic ablation of the anabolic enzyme ceramide kinase (CERK) significantly enhanced wound healing phenotypes. Importantly, postwounding inhibition of CERK enhanced the closure rate of acute wounds, improved the quality of healing, and increased fibroblast migration via a "class switch" in the eicosanoid profile. This switch reduced pro-inflammatory prostaglandins (e.g., prostaglandin E2) and increased levels of 5-hydroxyeicosatetraenoic acid and the downstream metabolite 5-oxo-eicosatetraenoic acid (5-oxo-ETE). Moreover, dermal fibroblasts from mice with genetically ablated CERK showed enhanced wound healing markers, while blockage of the murine 5-oxo-ETE receptor (oxoeicosanoid receptor 1) inhibited the enhanced migration phenotype of these cell models. Together, these studies reinforce the vital roles eicosanoids play in the wound healing process and demonstrate a novel role for CERK-derived C1P as a negative regulator of 5-oxo-ETE biosynthesis and the activation of oxoeicosanoid receptor 1 in wound healing. These findings provide foundational preclinical results for the use of CERK inhibitors to shift the balance from inflammation to resolution and increase the wound healing rate.

Systemic Elevation of n-3 Polyunsaturated Fatty Acids (n-3-PUFA) Is Associated with Protection against Visual, Motor, and Emotional Deficits in Mice following Closed-Head Mild Traumatic Brain Injury.

Traumatic brain injury (TBI) causes neuroinflammation and neurodegeneration leading to various pathological complications such as motor and sensory (visual) deficits, cognitive impairment, and depression. N-3 polyunsaturated fatty acid (n-3 PUFA) containing lipids are known to be anti-inflammatory, whereas the sphingolipid, ceramide (Cer), is an inducer of neuroinflammation and degeneration. Using Fat1+-transgenic mice that contain elevated levels of systemic n-3 PUFA, we tested whether they are resistant to mild TBI-mediated sensory-motor and emotional deficits by subjecting Fat1-transgenic mice and their WT littermates to focal cranial air blast (50 psi) or sham blast (0 psi, control). We observed that visual function in WT mice was reduced significantly following TBI but not in Fat1+-blast animals. We also found Fat1+-blast mice were resistant to the decline in motor functions, depression, and fear-producing effects of blast, as well as the reduction in the area of oculomotor nucleus and increase in activated microglia in the optic tract in brain sections seen following blast in WT mice. Lipid and gene expression analyses confirmed an elevated level of the n-3 PUFA eicosapentaenoic acid (EPA) in the plasma and brain, blocking of TBI-mediated increase of Cer in the brain, and decrease in TBI-mediated induction of Cer biosynthetic and inflammatory gene expression in the brain of the Fat1+ mice. Our results demonstrate that suppression of ceramide biosynthesis and inflammatory factors in Fat1+-transgenic mice is associated with significant protection against the visual, motor, and emotional deficits caused by mild TBI. This study suggests that n-3 PUFA (especially, EPA) has a promising therapeutic role in preventing neurodegeneration after TBI.

Sustained local inhibition of thrombin preserves renal microarchitecture and function after onset of acute kidney injury.

Acute kidney injury (AKI) management remains mainly supportive as no specific therapeutic agents directed at singular signaling pathways have succeeded in clinical trials. Here, we report that inhibition of thrombin-driven clotting and inflammatory signaling with use of locally-acting thrombin-targeted perfluorocarbon nanoparticles (PFC NP) protects renal vasculature and broadly modulates diverse inflammatory processes that cause renal ischemia reperfusion injury. Each PFC NP was complexed with ~13,650 copies of the direct thrombin inhibitor, PPACK (proline-phenylalanine-arginine-chloromethyl-ketone). Mice treated after the onset of AKI with PPACK PFC NP exhibited downregulated VCAM-1, ICAM-1, PGD2 prostanoid, M-CSF, IL-6, and mast cell infiltrates. Microvascular architecture, tubular basement membranes, and brush border components were better preserved. Non-reperfusion was reduced as indicated by reduced red blood cell trapping and non-heme iron. Kidney function and tubular necrosis improved at 24 hours versus the untreated control group, suggesting a benefit for dual inhibition of thrombosis and inflammation by PPACK PFC NP.

Lipid mediators and biomarkers associated with type 1 diabetes development.

Type 1 diabetes (T1D) is a consequence of autoimmune ß cell destruction, but the role of lipids in this process is unknown. We previously reported that activation of Ca2+-independent phospholipase A2ß (iPLA2ß) modulates polarization of macrophages (MΦ). Hydrolysis of the sn-2 substituent of glycerophospholipids by iPLA2ß can lead to the generation of oxidized lipids (eicosanoids), pro- and antiinflammatory, which can initiate and amplify immune responses triggering ß cell death. As MΦ are early triggers of immune responses in islets, we examined the impact of iPLA2ß-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 iPLA2ß 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.

Placental Production of Eicosanoids and Sphingolipids in Women Who Developed Preeclampsia on Low-Dose Aspirin.

Low-dose aspirin, which selectively inhibits thromboxane synthesis, is now standard of care for the prevention of preeclampsia in at risk women, but some women still develop preeclampsia despite an aspirin regimen. To explore the "aspirin failures," we undertook a comprehensive evaluation of placental lipids to determine if abnormalities in non-aspirin sensitive lipids might help explain why some women on low-dose aspirin develop preeclampsia. We studied placentas from women with normal pregnancies and women with preeclampsia. Placental villous explants were cultured and media analyzed by mass spectrometry for aspirin-sensitive and non-aspirin-sensitive lipids. In women who developed severe preeclampsia and delivered preterm, there were significant elevations in non-aspirin-sensitive lipids with biologic actions that could cause preeclampsia. There were significant increases in 15- and 20-hydroxyeicosatetraenoic acids and sphingolipids: D-e-C18:0 ceramide, D-e-C18:0 sphingomyelin, D-e-sphingosine-1-phosphate, and D-e-sphinganine-1-phosphate. With regard to lipids sensitive to aspirin, there was no difference in placental production of thromboxane or prostacyclin, but prostaglandins were lower. There was no difference for isoprostanes, but surprisingly, anti-inflammatory omega 3 and 6 PUFAs were increased. In total, 10 of 30 eicosanoids and 5 of 42 sphingolipids were abnormal in women with severe early onset preeclampsia. Lipid changes in women with mild preeclampsia who delivered at term were of lesser magnitude with few significant differences. The placenta produces many aspirin-sensitive and non-aspirin-sensitive lipids. Abnormalities in eicosanoids and sphingolipids not sensitive to aspirin might explain why some aspirin-treated women develop preeclampsia.

Emerging roles for human glycolipid transfer protein superfamily members in the regulation of autophagy, inflammation, and cell death.

Glycolipid transfer proteins (GLTPs) were first identified over three decades ago as ~24kDa, soluble, amphitropic proteins that specifically accelerate the intermembrane transfer of glycolipids. Upon discovery that GLTPs use a unique, all-α-helical, two-layer 'sandwich' architecture (GLTP-fold) to bind glycosphingolipids (GSLs), a new protein superfamily was born. Structure/function studies have provided exquisite insights defining features responsible for lipid headgroup selectivity and hydrophobic 'pocket' adaptability for accommodating hydrocarbon chains of differing length and unsaturation. In humans, evolutionarily-modified GLTP-folds have been identified with altered sphingolipid specificity, e. g. ceramide-1-phosphate transfer protein (CPTP), phosphatidylinositol 4-phosphate adaptor protein-2 (FAPP2) which harbors a GLTP-domain and GLTPD2. Despite the wealth of structural data (>40 Protein Data Bank deposits), insights into the in vivo functional roles of GLTP superfamily members have emerged slowly. In this review, recent advances are presented and discussed implicating human GLTP superfamily members as important regulators of: i) pro-inflammatory eicosanoid production associated with Group-IV cytoplasmic phospholipase A2; ii) autophagy and inflammasome assembly that drive surveillance cell release of interleukin-1ß and interleukin-18 inflammatory cytokines; iii) cell cycle arrest and necroptosis induction in certain colon cancer cell lines. The effects exerted by GLTP superfamily members appear linked to their ability to regulate sphingolipid homeostasis by acting in either transporter and/or sensor capacities. These timely findings are opening new avenues for future cross-disciplinary, translational medical research involving GLTP-fold proteins in human health and disease. Such avenues include targeted regulation of specific GLTP superfamily members to alter sphingolipid levels as a therapeutic means for combating viral infection, neurodegenerative conditions and circumventing chemo-resistance during cancer treatment.

Macrophage polarization is linked to Ca2+-independent phospholipase A2ß-derived lipids and cross-cell signaling in mice.

Phospholipases A2 (PLA2s) catalyze hydrolysis of the sn-2 substituent from glycerophospholipids to yield a free fatty acid (i.e., arachidonic acid), which can be metabolized to pro- or anti-inflammatory eicosanoids. Macrophages modulate inflammatory responses and are affected by Ca2+-independent phospholipase A2 (PLA2)ß (iPLA2ß). Here, we assessed the link between iPLA2ß-derived lipids (iDLs) and macrophage polarization. Macrophages from WT and KO (iPLA2ß-/-) mice were classically M1 pro-inflammatory phenotype activated or alternatively M2 anti-inflammatory phenotype activated, and eicosanoid production was determined by ultra-performance LC ESI-MS/MS. As a genotypic control, we performed similar analyses on macrophages from RIP.iPLA2ß.Tg mice with selective iPLA2ß overexpression in ß-cells. Compared with WT, generation of select pro-inflammatory prostaglandins (PGs) was lower in iPLA2ß-/- , and that of a specialized pro-resolving lipid mediator (SPM), resolvin D2, was higher; both changes are consistent with the M2 phenotype. Conversely, macrophages from RIP.iPLA2ß.Tg mice exhibited an opposite landscape, one associated with the M1 phenotype: namely, increased production of pro-inflammatory eicosanoids (6-keto PGF1α, PGE2, leukotriene B4) and decreased ability to generate resolvin D2. These changes were not linked with secretory PLA2 or cytosolic PLA2α or with leakage of the transgene. Thus, we report previously unidentified links between select iPLA2ß-derived eicosanoids, an SPM, and macrophage polarization. Importantly, our findings reveal for the first time that ß-cell iPLA2ß-derived signaling can predispose macrophage responses. These findings suggest that iDLs play critical roles in macrophage polarization, and we posit that they could be targeted therapeutically to counter inflammation-based disorders.