Immune Response Resetting in Ongoing Sepsis.

Cure of severe infections, sepsis, and septic shock with antimicrobial drugs is a challenge because morbidity and mortality in these conditions are essentially caused by improper immune response. We have tested the hypothesis that repeated reactivation of established memory to pathogens may reset unfavorable immune responses. We have chosen for this purpose a highly stringent mouse model of polymicrobial sepsis by cecum ligation and puncture. Five weeks after priming with a diverse Ag pool, high-grade sepsis was induced in C57BL/6j mice that was lethal in 24 h if left untreated. Antimicrobial drug (imipenem) alone rescued 9.7% of the animals from death, but >5-fold higher cure rate could be achieved by combining imipenem and two rechallenges with the Ag pool (p < 0.0001). Antigenic stimulation fine-tuned the immune response in sepsis by contracting the total CD3+ T cell compartment in the spleen and disengaging the hyperactivation state in the memory T subsets, most notably CD8+ T cells, while preserving the recovery of naive subsets. Quantitative proteomics/lipidomics analyses revealed that the combined treatment reverted the molecular signature of sepsis for cytokine storm, and deregulated inflammatory reaction and proapoptotic environment, as well as the lysophosphatidylcholine/phosphatidylcholine ratio. Our results showed the feasibility of resetting uncontrolled hyperinflammatory reactions into ordered hypoinflammatory responses by memory reactivation, thereby reducing morbidity and mortality in antibiotic-treated sepsis. This beneficial effect was not dependent on the generation of a pathogen-driven immune response itself but rather on the reactivation of memory to a diverse Ag pool that modulates the ongoing response.

Lysophosphatidylcholine Induces NLRP3 Inflammasome-Mediated Foam Cell Formation and Pyroptosis in Human Monocytes and Endothelial Cells.

Foam cells are specialized lipid-loaded macrophages derived from monocytes and are a key pathological feature of atherosclerotic lesions. Lysophosphatidylcholine (LPC) is a major lipid component of the plasma membrane with a broad spectrum of proinflammatory activities and plays a key role in atherosclerosis. However, the role of LPC in lipid droplet (LD) biogenesis and the modulation of inflammasome activation is still poorly understood. In the present study, we investigated whether LPC can induce foam cell formation through an analysis of LD biogenesis and determined whether the cell signaling involved in this process is mediated by the inflammasome activation pathway in human endothelial cells and monocytes. Our results showed that LPC induced foam cell formation in both types of cells by increasing LD biogenesis via a NLRP3 inflammasome-dependent pathway. Furthermore, LPC induced pyroptosis in both cells and the activation of the inflammasome with IL-1ß secretion, which was dependent on potassium efflux and lysosomal damage in human monocytes. The present study described the IL-1ß secretion and foam cell formation triggered by LPC via an inflammasome-mediated pathway in human monocytes and endothelial cells. Our results will help improve our understanding of the relationships among LPC, LD biogenesis, and NLRP3 inflammasome activation in the pathogenesis of atherosclerosis.

Lysophosphatidylcholine induces cyclooxygenase-2-dependent IL-6 expression in human cardiac fibroblasts.

Lysophosphatidylcholine (LysoPC) has been shown to induce the expression of inflammatory proteins, including cyclooxygenase-2 (COX-2) and interleukin-6 (IL-6), associated with cardiac fibrosis. Here, we demonstrated that LysoPC-induced COX-2 and IL-6 expression was inhibited by silencing NADPH oxidase 1, 2, 4, 5; p65; and FoxO1 in human cardiac fibroblasts (HCFs). LysoPC-induced IL-6 expression was attenuated by a COX-2 inhibitor. LysoPC-induced responses were mediated via the NADPH oxidase-derived reactive oxygen species-dependent JNK1/2 phosphorylation pathway, leading to NF-κB and FoxO1 activation. In addition, we demonstrated that both FoxO1 and p65 regulated COX-2 promoter activity stimulated by LysoPC. Overexpression of wild-type FoxO1 and S256D FoxO1 enhanced COX-2 promoter activity and protein expression in HCFs. These results were confirmed by ex vivo studies, where LysoPC-induced COX-2 and IL-6 expression was attenuated by the inhibitors of NADPH oxidase, NF-κB, and FoxO1. Our findings demonstrate that LysoPC-induced COX-2 expression is mediated via NADPH oxidase-derived reactive oxygen species generation linked to the JNK1/2-dependent pathway leading to FoxO1 and NF-κB activation in HCFs. LysoPC-induced COX-2-dependent IL-6 expression provided novel insights into the therapeutic targets of the cardiac fibrotic responses.

DFMG attenuates the activation of macrophages induced by co­culture with LPC­injured HUVE­12 cells via the TLR4/MyD88/NF­κB signaling pathway.

7­difluoromethoxy­5,4'­dimethoxy­genistein (DFMG) is a novel active chemical entity, which modulates the function and signal transduction of endothelial cells and macrophages (MPs), and is essential in the prevention of atherosclerosis. In the present study, the activity and molecular mechanism of DFMG on MPs was investigated using a Transwell assay to construct a non­contact co­culture model. Human umbilical vein endothelial cells (HUVE­12), which were incubated with lysophosphatidylcholine (LPC), were seeded in the upper chambers, whereas PMA­induced MPs were grown in the lower chambers. The generation of reactive oxygen species (ROS) and the release of lactate dehydrogenase (LDH) were measured using the corresponding assay kits. The proliferation and migration were assessed using 3­(4,5­dimethylthiazol­2­yl)­2,5­diphenyltetrazolium bromide and wound healing assays, respectively. Foam cell formation was examined using oil red O staining and a total cholesterol assay. The protein expression levels of Toll­like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88) and nuclear factor (NF)­κB p65 were detected by western immunoblotting. The secretion of interleukin (IL)­1ß was examined using an enzyme­linked immunosorbent assay. It was found that LPC significantly increased the generation of ROS and the release of LDH in HUVE­12 cells. The LPC­injured HUVE­12 cells activated MPs under co­culture conditions and this process was inhibited by DFMG treatment. LPC upregulated the expression levels of TLR4, MyD88 and NF­κB p65, and the secretion of IL­1ß in the supernatant of the co­cultured HUVE­12 cells and MPs. These effects were reversed by the application of DFMG. Furthermore, CLI­095 and IL­1Ra suppressed the activation of MPs that was induced by co­culture with injured HUVE­12 cells. These effects were further enhanced by co­treatment with DFMG, and DFMG exhibited synergistic effects with a TLR4­specific inhibitor. Take together, these findings revealed that DFMG attenuated the activation of MP induced by co­culture with LPC­injured HUVE­12 cells. This process was mediated via inhibition of the TLR4/MyD88/NF­κB signaling pathway in HUVE­12 cells.

TRPM2 contributes to LPC-induced intracellular Ca2+ influx and microglial activation.

Microglia are the resident immune cells which become activated in some pathological conditions in central nervous system (CNS). Lysophosphatidylcholine (LPC), an endogenous inflammatory phospholipid, is implicated in immunomodulatory function of glial cells in the CNS. Although several studies uncovered that LPC induces intracellular Ca2+ influx and morphologic change in microglia, there is still no direct evidence showing change of phosphorylation of mitogen-activated protein kinase (MAPK) p38 (p-p38), a widely used microglia activation marker, by LPC. Furthermore, the cellular mechanism of LPC-induced microglia activation remains unknown. In this study, we found that LPC induced intracellular Ca2+ increase in primary cultured microglia, which was blocked in the presence of Gd3+, non-selective transient receptor potential (TRP) channel blocker. RT-PCR and whole cell patch clamp recordings revealed molecular and functional expression of TRP melastatin 2 (TRPM2) in microglia. Using western blotting, we also observed that LPC increased phosphorylation of p38 MAPK, and the increase of p-p38 expression is also reversed in TRPM2-knockout (KO) microglia. Moreover, LPC induced membrane trafficking of TRPM2 and intrathecal injection of LPC increased Iba-1 immunoreactivity in the spinal cord, which were significantly reduced in KO mice. In addition, LPC-induced intracellular Ca2+ increase and inward currents were abolished in TRPM2-KO microglia. Taken together, our results suggest that LPC induces intracellular Ca2+ influx and increases phosphorylation of p38 MAPK via TRPM2, which in turn activates microglia.

Lysophosphatidylcholine plays critical role in allergic airway disease manifestation.

Phospholipase A2 (sPLA2), pivotal for allergic and inflammatory response, hydrolyses phosphatidylcholine (PC) to lysophosphatidylcholine (LPC). In present study, the role of LPC in allergic airway disease manifestation was studied using mouse model. Balb/c mice were immunized using cockroach extract (CE) and LPC release was blocked by sPLA2 inhibitor. Airway hyperresponse (AHR), lung-histology, total and differential leukocyte count (TLC&DLC), Th2 type cytokines, sPLA2 activity and LPC levels in bronchoalveolar lavage fluid (BALF) were measured. Exogenous LPC was given to the mice with or without CE sensitization, to demonstrate its role in allergic airway disease manifestation. Anti-CD1d antibody was given to study the involvement of natural killer T (NKT) cells in LPC induced response. AHR, lung-inflammation, TLC, DLC, Th2 type cytokines, sPLA2 activity and LPC levels were increased on CE challenge. sPLA2 activity and LPC release was blocked by sPLA2-inhibitor, which decreased AHR, and inflammatory parameters. Exogenous LPC with or without CE sensitization increased above parameters. CE challenge or LPC exposure increased LY49C(+)TCRß(+) NKT cells in BALF and spleen, which was reduced by anti-CD1d antibody, accompanied with reduction in AHR and allergic airway inflammation parameters. Conclusively, LPC induces allergic airway disease manifestation and it does so probably via CD1d-restricted LY49C(+)TCRß(+) NKT cells.

Clonal Immunoglobulin against Lysolipids in the Origin of Myeloma.

Antigen-driven selection has been implicated in the pathogenesis of monoclonal gammopathies. Patients with Gaucher's disease have an increased risk of monoclonal gammopathies. Here we show that the clonal immunoglobulin in patients with Gaucher's disease and in mouse models of Gaucher's disease-associated gammopathy is reactive against lyso-glucosylceramide (LGL1), which is markedly elevated in these patients and mice. Clonal immunoglobulin in 33% of sporadic human monoclonal gammopathies is also specific for the lysolipids LGL1 and lysophosphatidylcholine (LPC). Substrate reduction ameliorates Gaucher's disease-associated gammopathy in mice. Thus, long-term immune activation by lysolipids may underlie both Gaucher's disease-associated gammopathies and some sporadic monoclonal gammopathies.

Recognition of lysophosphatidylcholine by type II NKT cells and protection from an inflammatory liver disease.

Lipids presented by the MHC class I-like molecule, CD1d, are recognized by NK T (NKT) cells, which can be broadly categorized into two subsets. The well-characterized type I NKT cells express a semi-invariant TCR and can recognize both α- and ß-linked glycolipids, whereas type II NKT cells are less well studied, express a relatively diverse TCR repertoire, and recognize ß-linked lipids. Recent structural studies have shown a distinct mode of recognition of a self-glycolipid sulfatide bound to CD1d by a type II NKT TCR. To further characterize Ag recognition by these cells, we have used the structural data and screened other small molecules able to bind to CD1d and activate type II NKT cells. Using plate-bound CD1d and APC-based Ag presentation assay, we found that phospholipids such as lysophosphatidylcholine (LPC) can stimulate the sulfatide-reactive type II NKT hybridoma Hy19.3 in a CD1d-dependent manner. Using plasmon resonance studies, we found that this type II NKT TCR binds with CD1d-bound LPC with micromolar affinities similar to that for sulfatide. Furthermore, LPC-mediated activation of type II NKT cells leads to anergy induction in type I NKT cells and affords protection from Con A-induced hepatitis. These data indicate that, in addition to self-glycolipids, self-lysophospholipids are also recognized by type II NKT cells. Because lysophospholipids are involved during inflammation, our findings have implications for not only understanding activation of type II NKT cells in physiological settings, but also for the development of immune intervention in inflammatory diseases.

A synthetic biotinylated peptide, BP21, inhibits the induction of mRNA expression of inflammatory substances by oxidized- and lyso-phosphatidylcholine.

Preclinical Research Oxidized low-density lipoprotein (ox-LDL) is implicated in many inflammatory diseases, e.g., type 2 diabetes, obesity, atherosclerosis, and metabolic syndrome. We previously reported that a synthetic biotinylated peptide, BP21, inhibits the bioactivity of ox-LDL via direct binding to ox-LDL. Here, we investigated the effect of BP21 on the mRNA expression of proinflammatory mediators induced by two major components of ox-LDL, oxidized- and lyso-phosphatidylcholine (ox-PC and LPC), in monocytes/macrophages (THP-1 cells) and adipocytes (3T3-L1 cells). In THP-1 cells, BP21 markedly reduced the mRNA expression of interleukin (IL)-6, adipocyte fatty acid-binding protein (aP2), tumor necrosis factor-α, and mitogen-activated protein kinase phosphatase-1, which are induced by one of the major bioactive components of ox-PC, 1-palmitoyl-2-(5'-oxo-valeroyl)-sn-glycero-3-phosphocholine (POVPC), and reduced the mRNA expression of IL-6, the ox-LDL-specific scavenger receptor CD36, and aP2 induced by LPC. In adipocytes, the mRNA expression of IL-1ß as an adipokine and aP2 is highly induced by ox-PC and LPC, and BP21 markedly reduced the mRNA expression of IL-1ß and aP2 induced by POVPC and LPC. Furthermore, BP21 specifically bound to LPC and POVPC in a dose-dependent manner. These results suggest that BP21 may be useful lead for the potential treatment and prevention of inflammatory diseases caused by ox-PC and LPC.

Impact of serial measurements of lysophosphatidylcholine on 28-day mortality prediction in patients admitted to the intensive care unit with severe sepsis or septic shock.

PURPOSE: The purpose of this study is to investigate the effect of serial lysophosphatidylcholine (LPC) measurement on 28-day mortality prediction in patients with severe sepsis or septic shock admitted to the medical intensive care unit (ICU). METHODS: This is a prospective observational study of 74 ICU patients in a tertiary hospital. Serum LPC, white blood cell, C-reactive protein, and procalcitonin (PCT) levels were measured at baseline (day 1 of enrollment) and day 7. The LPC concentrations were compared with inflammatory markers using their absolute levels and relative changes. RESULTS: The LPC concentration on day 7 was significantly lower in nonsurvivors than in survivors (68.45 ± 42.36 µmol/L and 99.76 ± 73.65 µmol/L; P = .04). A decreased LPC concentration on day 7 to its baseline as well as a sustained high concentration of PCT on day 7 at more than 50% of its baseline value was useful for predicting the 28-day mortality. Prognostic utility was substantially improved when combined LPC and PCT criteria were applied to 28-day mortality outcome predictions. Furthermore, LPC concentrations increased over time in patients with appropriate antibiotics but not in those with inappropriate antibiotics. CONCLUSIONS: Serial measurements of LPC help in the prediction of 28-day mortality in ICU patients with severe sepsis or septic shock.

Lysophosphatidylcholine perpetuates macrophage polarization toward classically activated phenotype in inflammation.

Pro-inflammatory macrophages are involved in vascular inflammation and serve as the major effector cells in the pathophysiology of atherosclerosis. Phosphatidylcholine (PC) is a major phospholipid moiety affixed to oxidized low-density lipoprotein (oxLDL) and thought to play important roles in the development of atherosclerosis. In this study we described that a bioactive lipid derivative, lysophosphatidylcholine (lysoPC), generated from hydrolysis of the PC moiety of oxidized LDL, promoted and stabilized a strong M1 phenotype in macrophage polarization. Another derivative, 9-hydroxyoctadecadienoic acid (9-HODE), did not show the similar biological function. Blockade of G protein coupled receptor, G2A, which mediates the signal transduction of lysoPC, diminished the effects of lysoPC on the macrophage polarization toward M1 phenotype. The results provide insights into the new mechanism on how oxidized LDL participates in tissue inflammation in atherosclerosis.

Modified lipoproteins provide lipids that modulate dendritic cell immune function.

Both physiological and pathological situations can result in biochemical changes of low-density lipoproteins (LDL). Because they can deliver signals to dendritic cells (DC), these modified lipoproteins now appear as regulators of the immune response. Among these modified lipoproteins, oxidized LDL (oxLDL) that accumulate during inflammatory conditions have been extensively studied. Numerous studies have shown that oxLDL induce the maturation of DC, enhancing their ability to activate IFNγ secretion by T cells. LDL treated by secreted phospholipase A(2) also promote DC maturation. Among the bioactive lipids generated by oxidation or phospholipase treatment of LDL, lysophosphatidylcholine (LPC) and some saturated fatty acids induce DC maturation whereas some unsaturated fatty acids or oxidized derivatives have opposite effects. Among other factors, the nuclear receptor peroxisome-proliferator activated receptor γ (PPARγ) plays a crucial role in this regulation. Non-modified lipoproteins also contribute to the regulation of DC function, suggesting that the balance between native and modified lipoproteins, as well as the biochemical nature of the LDL modifications, can regulate the activation threshold of DC. Here we discuss two pathological situations in which the impact of LDL modifications on inflammation and immunity could play an important role. During atherosclerosis, modified LDL accumulating in the arterial intima may interfere with DC maturation and function, promoting a Th1 immune response and a local inflammation favoring the development of the pathology. In patients chronically infected, the hepatitis C virus (HCV) interferes with lipoprotein metabolism resulting in the production of infectious modified lipoproteins. These lipo-viral-particles (LVP) are modified low-density lipoproteins containing viral material that can alter DC maturation and affect specific toll-like receptor signaling. In conclusion, lipoprotein modifications play an important role in the regulation of immunity by delivering signals of danger to DC and modulating their function.

Inhibitory effects of paeoniflorin on lysophosphatidylcholine-induced inflammatory factor production in human umbilical vein endothelial cells.

Lysophosphatidylcholine (LPC) plays an important role in atherosclerosis through initiation of endothelial inflammation response. Paeoniflorin (PEF), isolated from the dry root of Paeonia, has been reported to exert an anti-inflammatory effect, but the exact mechanism is not fully understood. The aim of this study was to investigate the inhibitory effects of PEF on LPC-induced inflammatory factor production and the underlying mechanisms. In human umbilical vein endothelial cells (HUVECs), different concentrations (1, 10 or 100 µmol/l) of PEF were added 2 h prior to exposure to LPC (10 mg/l) for 24 h. The results showed that PEF significantly inhibited LPC-induced inflammatory factor production. In addition, PEF was also able to suppress the enhanced high mobility group box-1 (HMGB1) expression and release, upregulated expression of receptor for advanced glycation end product (RAGE), Toll-like receptor (TLR)-2 and TLR-4, and increased nuclear factor-κB (NF-κB) activity induced by LPC. Our results suggest that PEF suppresses LPC-induced inflammatory factor production through inhibition of the HMGB1-RAGE/TLR-2/TLR-4-NF-κB pathway.

Lysophosphatidylcholine enhances the suppressive function of human naturally occurring regulatory T cells through TGF-ß production.

Naturally occurring CD4(+)CD25(+) regulatory T cells (nTregs) play a pivotal role in the maintenance of self-tolerance and immune homeostasis. To gain insight into the mechanism of action of nTregs in pathological and physiological immune responses, it is important to analyze bioactive molecules that modulate the maintenance and function of nTregs. From a library of bioactive lipids, we obtained lysophosphatidylcholine (LPC) as a molecule that enhanced the Foxp3 expression and suppressive function of human nTregs significantly in comparison with those of DMSO-treated nTregs (control). The expression levels of TGF-ß1 mRNA and protein in LPC-treated nTregs were significantly higher than those in control nTregs. After treatment with anti-TGF-ß1 antibody, the increases in Foxp3 expression and the suppressive properties of LPC-treated nTregs returned to the levels observed in control nTregs. These findings indicate that LPC enhances Foxp3 expression and the suppressive function of nTregs through TGF-ß1 produced by nTregs themselves. Experimental knockdown of G2A and GPR4 showed that this LPC-induced TGF-ß1 expression in nTregs was due to G2A signaling, and did not involve GPR4. Moreover, JNK was a major contributor to LPC-induced TGF-ß1 expression in nTregs, although LPC activated MAPKs including ERK1/2, p38 MAPK, and JNK via G2A. LPC is a bioactive lysolipid highly abundant in the circulation. Therefore, LPC may contribute to the maintenance and function of human nTregs in vivo.

Sodium dependence of lysophosphatidylcholine-induced caspase-1 activity and reactive oxygen species generation.

The proinflammatory cytokines interleukin (IL)-1ß and IL-18 play pivotal roles in neuroinflammatory diseases. Caspase-1-mediated proteolytic cleavage is required to convert the premature, biologically inactive cytokines to their biologically active forms capable of promoting tissue inflammation. Although caspases have been recognized as potential therapeutic targets in inflammatory diseases, mechanisms regulating caspase-1 activation are not fully understood. Here we demonstrate that the proinflammatory lipid lysophosphatidylcholine (LPC) initiates microglial caspase-1 activation in a Na(+)-dependent manner. LPC-induced caspase-1 activity was almost completely inhibited upon omission of extracellular Na(+), but was unaffected by inhibition of Na(+)/K(+)-ATPase with ouabain or by inhibition of Na(+)/H(+) antiport with amiloride. Inhibition of caspase-1-mediated IL-1ß processing by Na(+)-free medium led to reduced amounts of mature IL-1ß released from LPC-stimulated microglia. Furthermore, LPC-induced production of reactive oxygen species (ROS) was abolished by Na(+)-free medium, indicating Na(+) dependence of NADPH oxidase activity in LPC-stimulated microglia. Since ROS production was found to be crucial to caspase-1 activation in LPC-stimulated microglia, the Na(+) dependence of caspase-1 can be related to the Na(+) dependence of NADPH oxidase. In summary, it is suggested that in LPC-activated microglia, Na(+) influx is required for the production of NADPH oxidase-mediated ROS, which subsequently stimulate caspase-1 activity.

Schistosomal­derived lysophosphatidylcholine are involved in eosinophil activation and recruitment through Toll­like receptor-2-dependent mechanisms.

Parasite­derived lipids may play important roles in host­pathogen interactions and escape mechanisms. Herein, we evaluated the role of schistosomal­derived lipids in Toll­like receptor (TLR)-2 and eosinophil activation in Schistosoma mansoni infection. Mice lacking TLR2 exhibited reduced liver eosinophilic granuloma, compared with that of wild­type animals, following S. mansoni infection. Decreased eosinophil accumulation and eosinophil lipid body (lipid droplet) formation, at least partially due to reduced production of eotaxin, interleukin (IL)­5, and IL­13 in S. mansoni-infected TLR2-/- mice, compared with the corresponding production in wild­type mice, was noted. Although no differences were observed in survival rates during the acute schistosomal infection (up to 50 days), increased survival of TLR2-/- mice, compared with survival of wild­type mice, was observed during the chronic phase of infection. Schistosomal lipid extract­ and schistosomal­derived lysophosphatidylcholine (lyso­PC)-stimulated macrophages in vitro induced TLR2­dependent NF­kB activation and cytokine production. Furthermore, in vivo schistosomal lyso­PC administration induced eosinophil recruitment and cytokine production, in a mechanism largely dependent on TLR2. Taken together, our results suggest that schistosomal­derived lyso­PC may participate in cytokine production and eosinophil activation through a TLR2­dependent pathway in S. mansoni infection. Moreover, our results suggest that TLR2­dependent inflammatory reaction, cytokine production, and eosinophil recruitment and activation may contribute to the pathogenesis and lethality in the chronic phase of infection.

Single lysophosphatidylcholine components exhibit adjuvant activities in vitro and in vivo.

Improving vaccine immunogenicity by developing new adjuvant formulations has long been a goal of vaccinologists. It has previously been shown that a natural mix of lysophosphatidylcholine (LPC) from chicken eggs promotes mature dendritic cell (DC) generation in vitro and primes antigen-specific immune responses in mice. In the present study, we dissected the adjuvant potentials of five individual LPC components found in the chicken egg mixture. In vitro analyses of the impact of the individual components on the maturation of human DCs were performed by means of phenotypic analysis, chemokine secretion analysis, and analysis of the ability of mature DC to stimulate T lymphocytes. Two components, C16:0-LPC and C18:0-LPC, were identified to be capable of the upregulation of expression of CD86, HLA-DR, and CD40 on in vitro-cultured monocyte-derived DCs from healthy donors. Both induced the release of chemokines to high concentrations (macrophage inflammatory protein 1, monocyte chemoattractant protein 1) or moderate concentrations (interleukin-8 [IL-8], gamma interferon-inducible protein 10). In addition, C16:0-LPC engaged naïve T cells to produce gamma interferon. This suggests that C16:0-LPC and C18:0-LPC have the capacity to promote, at least in vitro, a Th1-oriented response. The intravenous injection of C16:0-LPC or C18:0-LPC into mice resulted in the detectable secretion of IL-6 and IL-5 in sera. Both LPC components were tested for their capacities to act as adjuvants for two selected immunogens: the hepatitis B virus surface antigen and the hepatitis C virus NS3 helicase. The secretion of specific IgG1 was observed with either or both C16:0-LPC and C18:0-LPC, depending on the immunogen tested, and was observed at an efficiency comparable to that of alum. These data identify C16:0-LPC and C18:0-LPC as the active components of the LPC natural mixture. Although discrepancies between the results of the in vitro and in vivo analyses existed, studies with animals suggest that these components can trigger significant and specific humoral-mediated immunity.

Recognition of lyso-phospholipids by human natural killer T lymphocytes.

Natural killer T (NKT) cells are a subset of T lymphocytes with potent immunoregulatory properties. Recognition of self-antigens presented by CD1d molecules is an important route of NKT cell activation; however, the molecular identity of specific autoantigens that stimulate human NKT cells remains unclear. Here, we have analyzed human NKT cell recognition of CD1d cellular ligands. The most clearly antigenic species was lyso-phosphatidylcholine (LPC). Diacylated phosphatidylcholine and lyso-phosphoglycerols differing in the chemistry of the head group stimulated only weak responses from human NKT cells. However, lyso-sphingomyelin, which shares the phosphocholine head group of LPC, also activated NKT cells. Antigen-presenting cells pulsed with LPC were capable of stimulating increased cytokine responses by NKT cell clones and by freshly isolated peripheral blood lymphocytes. These results demonstrate that human NKT cells recognize cholinated lyso-phospholipids as antigens presented by CD1d. Since these lyso-phospholipids serve as lipid messengers in normal physiological processes and are present at elevated levels during inflammatory responses, these findings point to a novel link between NKT cells and cellular signaling pathways that are associated with human disease pathophysiology.

Linking inflammation to natural killer T cell activation.

Immune activation is often associated with inflammation, but inflammation's role in the expansion of antigen-specific immune responses remains unclear. This primer focuses on recent findings that show how specific natural killer T cells are activated by inflammatory messengers, thus illuminating the cellular and molecular links between immunity and inflammation.