A role for phospholipid oxidation products as modulators of inflammatory reactions in atherogenesis

Berliner i wsp. wykazali w 1990 r., że LDL umiarkowanie oksydowane przez wydłużone przechowywanie lub reakcję z jonami żelaza charakteryzują się specyficznymi właściwościami, odmiennymi od natywnych i oksydowanych LDL. Lipidy te zostały nazwane minimalnie zmodyfikowanymi lub minimalnie oksydowanymi (MM -LDL). Mają one specyficzną aktywność biologiczną wyrażoną przez: indukcję oddziaływań monocytów i śródbłonka, zwiększenie ekspresji MCP-1, transmigracji monocytów oraz wzmagania ekspresji genów wczesnej odpowiedzi (JE, KC, c-myc) w komórkach fibroblastów mysich. Związek 1-palmitoil-2-archidonoil-sn-glicero- -3-fosfocholina (PAPC) to fosfolipid zawierający resztę arachidonową, szczególnie podatny na oksydację. Zidentyfikowane pochodne PAPC o aktywności biologicznej to: 1-palmitoil-2-(5-oksowaleryl)-sn-glicero-3 -fosfocholina (POVPC) i 1-palmitoil-2-glutaryl-sn-glicero-3-fosfocholina (PGPC) oraz epoksyizoprostan-PC. Komórki eukariotyczne pod wpływem stymulacji różnymi czynnikami lub oksydacji lipidów, w wyniku wzrostu wewnątrzkomórkowego stężenia wapnia i utraty asymetrii błony, uwalniają do przestrzeni pozakomórkowej pęcherzyki błonowe (MV). Pęcherzyki te są przenośnikami lipidów aktywnych biologicznie. Wydaje się, że oksydowane lipidy mogą hamować ostry stan zapalny, a nasilać przewlekły. Istnieją dowody, że stan zapalny indukowany przez LPS, zależny od NFқB, jest hamowany przez OxPAPC. Oksydowane fosfolipidy zwiększają syntezę EGR-1. Nasilona ekspresja EGR-1 jest obecna w blaszkach miażdżycowych, a indukują ją czynniki wzrostu, cytokiny, hipoksja, siły mechaniczne i czynniki uszkadzające. EGR-1 zwiększa natomiast ekspresję TF. Wykazano również, że oksydowane fosfolipidy poza zwiększaniem stężenia czynników prozapalnych nasilają ekspresję enzymów protekcyjnych, takich jak HO-1. Enzym ten jest kluczowym, regulacyjnym enzymem katabolizmu hemu, wykazujący działanie antyoksydacyjne. Podsumowując, wyjaśnienie mechanizmów i szlaków przekazywania sygnału indukowanych przez oksydowane lipidy, regulujących odpowiedź zapalną w ścianie naczynia, będzie prowadzić do nowych sposobów interwencji leczniczej w chorobach związanych z przewlekłym odczynem zapalnym.In 1990 Berliner et al. demonstrated that LDL that had been mildly oxidized by prolonged storage or using iron had unique properties different from those of native and oxidized LDL (oxLDL). This LDL was called minimally oxidized or minimally modified LDL (MM-LDL) and shown to have unique bioactivity such as: induction of monocyte-endothelial interactions, expression of MCP-1, monocyte transmigration. 1-palmitoyl -2-archidonoyl-sn-glycero-3-phosphocholine (PAPC) is an arachidonic acid-containing phospholipid especially prone to oxidation. Biologically active oxidized derivatives of PAPC were identified as 1-palmitoyl-2-(5 -oxovaleryl)-sn-glycero-3-phosphocholine (POVPC) and 1-palmitoyl-2-glutaryl-sn-glycero-3-phosphocholine (PGPC) and epoxyisoprostane PC. Upon stimulation with various agonists or lipid oxidation, eukaryotic cells release membrane vesicles (MV) into the extracellular space due to increase in intracellular calcium and loss of membrane asymmetry. Membrane vesicles are also carriers of the bioactive lipids. Oxidized lipids may inhibit acute, but promote chronic inflammation. There is evidence that LPS-induced NFkB-mediated inflammation is down regulated by OxPAPC. Oxidized phospholipids increase synthesis of EGR-1. EGR-1 is known to be up regulated by growth factors, cytokines, hypoxia, physical forces and injurious stimuli, and high levels of EGR-1 were found in atherosclerotic lesions. EGR-1 up-regulates expression of TF. In contrast to the up-regulation of proinflammatory genes, oxidized phospholipids were also shown to induce the expression of protective enzymes such as HO-l, which is the rate-limiting enzyme in heme-catabolism and has antioxidative capacity. Thus, identification of mechanisms and signaling pathways induced by oxidized lipids that modulate inflammatory response in the vascular wall will lead to novel strategies of therapeutic intervention in chronic inflammatory diseases

NFAT5 expression in bone marrow-derived cells enhances atherosclerosis and drives macrophage migration

Objective: We have previously shown that the transcription factor, nuclear factor of activated T-cells 5 (NFAT5), regulates vascular smooth muscle cell phenotypic modulation, but the role of NFAT5 in atherosclerosis is unknown. Our main objective was to determine if NFAT5 expression in bone marrow (BM)-derived cells altered atherosclerotic development and macrophage function. Methods and Results: NFAT5+/-ApoE-/- mice were generated for in vivo atherosclerosis studies. Following high fat diet feeding, en face analysis of the thoracic aorta established that genome-wide NFAT5 haploinsufficiency reduced atherosclerotic lesion formation by 73%. BM transplant studies revealed that transplantation of NFAT5+/-ApoE-/- marrow into NFAT5+/+ApoE-/- mice resulted in a similar 86% reduction in lesion formation. In vitro functional analysis of BM-derived macrophages demonstrated that NFAT5 is required for macrophage migration, which is a key event in the propagation of atherosclerosis. Conclusion: We have identified NFAT5 in BM-derived cells as a positive regulator of atherosclerotic lesion formation and macrophage function in the vasculature.open

Repurposing Anti-Inflammasome NRTIs for Improving Insulin Sensitivity and Reducing Type 2 Diabetes Development

Innate immune signaling through the NLRP3 inflammasome is activated by multiple diabetes-related stressors, but whether targeting the inflammasome is beneficial for diabetes is still unclear. Nucleoside reverse-transcriptase inhibitors (NRTI), drugs approved to treat HIV-1 and hepatitis B infections, also block inflammasome activation. Here, we show, by analyzing five health insurance databases, that the adjusted risk of incident diabetes is 33% lower in patients with NRTI exposure among 128,861 patients with HIV-1 or hepatitis B (adjusted hazard ratio for NRTI exposure, 0.673; 95% confidence interval, 0.638 to 0.710; P \u3c 0.0001; 95% prediction interval, 0.618 to 0.734). Meanwhile, an NRTI, lamivudine, improves insulin sensitivity and reduces inflammasome activation in diabetic and insulin resistance-induced human cells, as well as in mice fed with high-fat chow; mechanistically, inflammasome-activating short interspersed nuclear element (SINE) transcripts are elevated, whereas SINE-catabolizing DICER1 is reduced, in diabetic cells and mice. These data suggest the possibility of repurposing an approved class of drugs for prevention of diabetes

Nucleotides released by apoptotic cells act as a find-me signal to promote phagocytic clearance

Phagocytic removal of apoptotic cells occurs efficiently in vivo such that even in tissues with significant apoptosis, very few apoptotic cells are detectable1. This is thought to be due to the release of find-me signals by apoptotic cells that recruit motile phagocytes such as monocytes, macrophages, and dendritic cells, leading to the prompt clearance of the dying cells2. However, the identity and in vivo relevance of such find-me signals are not well understood. Here, through several lines of evidence, we identify extracellular nucleotides as a critical apoptotic cell find-me signal. We demonstrate the caspase-dependent release of ATP and UTP (in equimolar quantities) during the early stages of apoptosis by primary thymocytes and cell lines. Purified nucleotides at these concentrations were sufficient to induce monocyte recruitment comparable to apoptotic cell supernatants. Enzymatic removal of ATP and UTP (by apyrase or ectopic CD39 expression) abrogated the ability of apoptotic cell supernatants to recruit monocytes in vitro and in vivo. We then identified the ATP/UTP receptor P2Y2 as a critical sensor of nucleotides released by apoptotic cells using RNAi depletion studies in monocytes, and macrophages from P2Y2-null mice3. The in vivo relevance of nucleotides in apoptotic cell clearance was revealed by two approaches. First, in a murine air-pouch model, apoptotic cell supernatants induced a three-fold greater recruitment of monocytes and macrophages compared to supernatants from healthy cells; this recruitment was abolished by depletion of nucleotides and significantly decreased in P2Y2−/− mice. Second, clearance of apoptotic thymocytes was significantly impaired by either depletion of nucleotides or interference with P2Y receptor function (by pharmacological inhibition, or in P2Y2−/− mice). These results identify nucleotides as a critical find-me cue released by apoptotic cells to promote P2Y2-dependent phagocyte recruitment, and provide strong evidence for a clear relationship between a find-me signal and efficient corpse clearance in vivo

cGAS Drives Noncanonical-Inflammasome Activation in Age-Related Macular Degeneration

Geographic atrophy is a blinding form of age-related macular degeneration characterized by retinal pigmented epithelium (RPE) death; the RPE also exhibits DICER1 deficiency, resultant accumulation of endogenous Alu-retroelement RNA, and NLRP3-inflammasome activation. How the inflammasome is activated in this untreatable disease is largely unknown. Here we demonstrate that RPE degeneration in human-cell-culture and mouse models is driven by a noncanonical-inflammasome pathway that activates caspase-4 (caspase-11 in mice) and caspase-1, and requires cyclic GMP-AMP synthase (cGAS)-dependent interferon-β production and gasdermin D-dependent interleukin-18 secretion. Decreased DICER1 levels or Alu-RNA accumulation triggers cytosolic escape of mitochondrial DNA, which engages cGAS. Moreover, caspase-4, gasdermin D, interferon-β, and cGAS levels were elevated in the RPE in human eyes with geographic atrophy. Collectively, these data highlight an unexpected role of cGAS in responding to mobile-element transcripts, reveal cGAS-driven interferon signaling as a conduit for mitochondrial-damage-induced inflammasome activation, expand the immune-sensing repertoire of cGAS and caspase-4 to noninfectious human disease, and identify new potential targets for treatment of a major cause of blindness

Oxidized phospholipids inhibit phagocytosis and impair outcome in gram-negative sepsis in vivo

Oxidized phospholipids that are generated during inflammation exert anti-inflammatory properties and prevent death during murine endotoxemia. Oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholine (OxPAPC) inhibits the interaction of LPS with LPS-binding protein and CD14. In this study, we determined the functional properties of OxPAPC and potential interference with CD14 during abdominal sepsis caused by Escherichia coli. Administration of OxPAPC rendered mice highly susceptible to E. coli peritonitis, as indicated by an accelerated mortality and enhanced bacterial outgrowth and dissemination. CD14(-/-) mice also displayed increased mortality and bacterial outgrowth and OxPAPC did not further impair host defense in these animals. The mechanisms by which OxPAPC and CD14 deficiency impaired the immune response differed: whereas CD14(-/-) mice demonstrated a strongly reduced recruitment of phagocytes to the site of the infection, OxPAPC did not influence the influx of inflammatory cells but strongly diminished the phagocytosing capacity of neutrophils and macrophages by a CD14-independent mechanism. Furthermore, OxPAPC potently inhibited uptake of fluorospheres as well as receptor-mediated endocytosis and fluid-phase pinocytosis. These data suggest that oxidized phospholipids such as produced during inflammatory reactions may contribute to mortality during Gram-negative sepsis in vivo via impairment of the phagocytic properties of professional phagocyte