sphingosine-1-phosphate and Sepsis

Sphingosine 1-phosphate (S1P) is a bioactive metabolite of sphingomyelin. S1P activates a series of signaling cascades by acting on its receptors S1PR1-3 on endothelial cells (ECs), which plays an important role in endothelial barrier maintenance, anti-inflammation, antioxidant and angiogenesis, and thus is considered as a potential therapeutic biomarker for ischemic stroke, sepsis, idiopathic pulmonary fibrosis, cancers, type 2 diabetes and cardiovascular diseases. We presently review the levels of S1P in those vascular and vascular-related diseases. Plasma S1P levels were reduced in various inflammation-related diseases such as atherosclerosis and sepsis, but were increased in other diseases including type 2 diabetes, neurodegeneration, cerebrovascular damages such as acute ischemic stroke, Alzheimer's disease, vascular dementia, angina, heart failure, idiopathic pulmonary fibrosis, community-acquired pneumonia, and hepatocellular carcinoma. Then, we highlighted the molecular mechanism by which S1P regulated EC biology including vascular development and angiogenesis, inflammation, permeability, and production of reactive oxygen species (ROS), nitric oxide (NO) and hydrogen sulfide (H₂S), which might provide new ways for exploring the pathogenesis and implementing individualized therapy strategies for those diseases.

Topics: Diabetes Mellitus, Type 2; Endothelial Cells; Humans; Idiopathic Pulmonary Fibrosis; Inflammation; Ischemic Stroke; Lysophospholipids; Sepsis; Sphingosine

Sphingosine 1-phosphate (S1P) is an important immune modulator responsible for physiological cellular responses like lymphocyte development and function, positioning and emigration of T and B cells and cytokine secretion. Recent reports indicate that S1P does not only regulate immunity, but can also protect the function of organs by inducing disease tolerance. S1P also influences the replication of certain pathogens, and sphingolipids are also involved in pathogen recognition and killing. Certain carrier molecules for S1P like serum albumin and high density lipoproteins contribute to the regulation of S1P effects. They are able to associate with S1P and modulate its signaling properties. Similar to S1P, both carrier molecules are also decreased in sepsis patients and likely contribute to sepsis pathology and severity. In this review, we will introduce the concept of disease tolerance and the involvement of S1P. We will also discuss the contribution of S1P and its precursor sphingosine to host defense mechanisms against pathogens. Finally, we will summarize current data demonstrating the influence of carrier molecules for differential S1P signaling. The presented data may lead to new strategies for the prevention and containment of sepsis.

Topics: Animals; Humans; Immune Tolerance; Lysophospholipids; Sepsis; Signal Transduction; Sphingosine

Sphingosine 1-phosphate (S1P) is a multifaceted lipid signaling molecule that activates five specific G protein-coupled S1P receptors. Despite the fact that S1P is known as one of the strongest barrier-enhancing molecules for two decades, no medical application is available yet. The reason for this lack of translation into clinical practice may be the complex regulatory network of S1P signaling, metabolism and transportation.In this review, we will provide an overview about the physiology and the network of S1P signaling with the focus on endothelial barrier maintenance in inflammation. We briefly describe the physiological role of S1P and the underlying S1P signaling in barrier maintenance, outline differences of S1P signaling and metabolism in inflammatory diseases, discuss potential targets and compounds for medical intervention, and summarize our current knowledge regarding the role of S1P in the maintenance of specialized barriers like the blood-brain barrier and the placenta.

Topics: Humans; Inflammation; Lysophospholipids; Sepsis; Sphingosine

Sepsis is an acute life-threatening multiple organ failure caused by a dysregulated host response to infection. Endothelial dysfunction, particularly barrier disruption leading to increased vascular permeability, edema, and insufficient tissue oxygenation, is critical to sepsis pathogenesis. Sphingosine-1-phosphate (S1P) is a signaling lipid that regulates important pathophysiological processes including vascular endothelial cell permeability, inflammation, and coagulation. It is present at high concentrations in blood and lymph and at very low concentrations in tissues due to the activity of the S1P-degrading enzyme S1P-lyase in tissue cells. Recently, four preclinical observational studies determined S1P levels in serum or plasma of sepsis patients, and all found reduced S1P levels associated with the disease. Based on these findings, this review summarizes S1P-regulated processes pertaining to endothelial functions, discusses the possible use of S1P as a marker and possibilities how to manipulate S1P levels and S1P receptor activation to restore endothelial integrity, dampens the inflammatory host response, and improves organ function in sepsis.

Topics: Biomarkers; Endothelial Cells; Humans; Lysophospholipids; Sepsis; Signal Transduction; Sphingosine

Acute lung injury (ALI) attributable to sepsis or mechanical ventilation and subacute lung injury because of ionizing radiation (RILI) share profound increases in vascular permeability as a key element and a common pathway driving increased morbidity and mortality. Unfortunately, despite advances in the understanding of lung pathophysiology, specific therapies do not yet exist for the treatment of ALI or RILI, or for the alleviation of unremitting pulmonary leakage, which serves as a defining feature of the illness. A critical need exists for new mechanistic insights that can lead to novel strategies, biomarkers, and therapies to reduce lung injury. Sphingosine 1-phosphate (S1P) is a naturally occurring bioactive sphingolipid that acts extracellularly via its G protein-coupled S1P1-5 as well as intracellularly on various targets. S1P-mediated cellular responses are regulated by the synthesis of S1P, catalyzed by sphingosine kinases 1 and 2, and by the degradation of S1P mediated by lipid phosphate phosphatases, S1P phosphatases, and S1P lyase. We and others have demonstrated that S1P is a potent angiogenic factor that enhances lung endothelial cell integrity and an inhibitor of vascular permeability and alveolar flooding in preclinical animal models of ALI. In addition to S1P, S1P analogues such as 2-amino-2-(2-[4-octylphenyl]ethyl)-1,3-propanediol (FTY720), FTY720 phosphate, and FTY720 phosphonates offer therapeutic potential in murine models of lung injury. This translational review summarizes the roles of S1P, S1P analogues, S1P-metabolizing enzymes, and S1P receptors in the pathophysiology of lung injury, with particular emphasis on the development of potential novel biomarkers and S1P-based therapies for ALI and RILI.

Topics: Acute Lung Injury; Animals; Anti-Inflammatory Agents; Biomarkers; Capillary Permeability; Fingolimod Hydrochloride; Humans; Lung; Lysophospholipids; Membrane Proteins; Nerve Tissue Proteins; Phosphotransferases (Alcohol Group Acceptor); Pneumonia; Propylene Glycols; Receptors, Lysosphingolipid; Sepsis; Sphingosine; Transferases (Other Substituted Phosphate Groups); Translational Research, Biomedical

The review will address the potential roles of apolipoprotein M (apoM) as a carrier protein and modulator of sphingosine-1-phosphate (S1P) bioactivity.. Recombinant apoM can bind small lipids such as retinoic acid, oxidized phospholipids, and S1P. Thus, the effects of apoM may be pleiotrophic. The S1P binding ability of apoM has biological impact. ApoM-bound S1P can activate S1P1 receptors on endothelial cells and deficiency of apoM abolishes the presence of S1P in HDL. In mice, the lack of apoM causes dysfunctional endothelial barrier function in the lungs. In humans, sepsis that is characterized by impaired endothelial function is associated with low plasma apoM.. Plasma apoM is mainly bound to HDL. The roles of apoM in atherosclerosis and lipoprotein metabolism have been given much attention. New in the field is the discovery of apoM as a chaperone for S1P. S1P is a bioactive lipid with effects on angiogenesis, lymphocyte trafficking, endothelial cell migration, and inflammation. A drug targeting the S1P-system (fingolimod) is now used for treatment of multiple sclerosis. It improves the blood-brain barrier and inhibits migration of lymphocytes into the brain. Further exploration of the apoM/S1P axis may uncover its potential as a biomarker and target for new treatments.

Topics: Animals; Apolipoproteins; Apolipoproteins M; Atherosclerosis; Endothelial Cells; Humans; Lipid Metabolism; Lipocalins; Lipoproteins, HDL; Lysophospholipids; Protein Binding; Sepsis; Sphingosine

Topics: Humans; Lysophospholipids; Receptor, PAR-1; Sepsis; Signal Transduction; Sphingosine; Thrombin

Sepsis is characterized physiologically by an aberrant systemic inflammatory response and microvascular dysfunction. While appropriate antibiotics and supportive care are essential in the management of the septic patient, therapies targeting specific aspects of the pathophysiology could have a significant impact on the morbidity and mortality associated with both sepsis and its sequlea, including acute lung injury (ALI). We have characterized several mediators of endothelial cell (EC) barrier function that may serve as novel therapies for sepsis-induced microvascular dysfunction including simvastatin, adenosine triphosphate (ATP), sphingosine 1-phosphate (S1P), and activated protein C (APC). Notably, APC is already available for the treatment of severe sepsis, however, to date its mechanism of action has been unclear. While distinct in many ways, we have found that these agonists have in common the ability to induce dynamic rearrangement of the EC actin cytoskeleton that corresponds to barrier protection. In addition, we have extended our in vitro findings to relevant animal models of endotoxin-induced acute lung injury and have confirmed beneficial effects of both simvastatin and S1P which are associated with evidence of decreased vascular permeability in this setting. Moreover, our data also indicate that APC effects in sepsis may be largely due to augmentation of EC barrier function affecting decreased microvascular permeability. We speculate that the administration of direct modulators of EC barrier function and microvascular permeability, such as those described here, may ultimately become the standard of care for the septic patient.

Topics: Adenosine Triphosphate; Animals; Capillaries; Capillary Permeability; Dogs; Humans; Lysophospholipids; Protein C; Sepsis; Simvastatin; Sphingosine; Venules

The aim of this study was to investigate the correlation between plasma sphingosine-1-phosphate (S1P) and ceramide concentrations in sepsis, and the possible mechanisms for altered expression.. Plasma S1P and ceramide concentrations were measured by HPLC-ESI-MS/MS. HLA-DR (human leukocyte antigen-DR) expression on peripheral blood mononuclear cells was examined by flow cytometry. Platelet sphingosine kinases 1/2 (SphK1/2) mRNA expression, protein content, and enzyme activities were determined by qRT-PCR, western blot, and commercial enzyme assay kits, respectively.. Compared with healthy and ICU controls, septic patients had significantly decreased plasma S1P but increased ceramide concentrations (P < 0.05). S1P concentration was negatively associated with the ceramide concentration in the septic patients (r = -0.36, P < 0.05). Linear regression analysis found that plasma S1P and ceramide were linked not only to sequential (sepsis-related) organ failure assessment (SOFA) score but also the HLA-DR expression on circulating monocytes. An receiver operating characteristic analysis, including S1P, ceramide, SOFA score and HLA-DR, showed integrated analysis of S1P and ceramide as the better powerful predictors of septic lethality with area under the curve value of 0.95. More importantly, we found the platelet SphKs activities and the expression levels of SphK1 were significantly decreased in septic patients (P < 0.05). Linear regression analysis revealed platelet SphKs activity was positively associated with the plasma S1P concentration of the septic patients (r = -0.41, P = 0.02).. Integrated analysis of plasma S1P and ceramide predict septic mortality with high accuracy. The decreased platelet SphK1 expression and subsequent reduced SphKs activity might be responsible for the decreased plasma S1P levels during sepsis.

Topics: Adult; Aged; Ceramides; Disease-Free Survival; Female; Humans; Lysophospholipids; Male; Middle Aged; Predictive Value of Tests; Sepsis; Sphingosine; Survival Rate

Sepsis is a leading cause of in-hospital mortality resulting from a dysregulated response to infection. Novel immunomodulatory therapies targeting macrophage metabolism have emerged as an important focus for current sepsis research. However, understanding the mechanisms underlying macrophage metabolic reprogramming and how they impact immune response requires further investigation. Here, we identify macrophage-expressed Spinster homolog 2 (Spns2), a major transporter of sphingosine-1-phosphate (S1P), as a crucial metabolic mediator that regulates inflammation through the lactate-reactive oxygen species (ROS) axis. Spns2 deficiency in macrophages significantly enhances glycolysis, thereby increasing intracellular lactate production. As a key effector, intracellular lactate promotes pro-inflammatory response by increasing ROS generation. The overactivity of the lactate-ROS axis drives lethal hyperinflammation during the early phase of sepsis. Furthermore, diminished Spns2/S1P signaling impairs the ability of macrophages to sustain an antibacterial response, leading to significant innate immunosuppression in the late stage of infection. Notably, reinforcing Spns2/S1P signaling contributes to balancing the immune response during sepsis, preventing both early hyperinflammation and later immunosuppression, making it a promising therapeutic target for sepsis.

Topics: Anion Transport Proteins; Humans; Immunosuppression Therapy; Lactates; Macrophages; Reactive Oxygen Species; Sepsis

Splenic B cells exhibit a high expression of the G protein-coupled sphingosine-1-phosphate (S1P) receptor type 4 (S1PR. In this study, S1PR. Loss of S1PR. These observations suggest that S1P signaling mediated by S1PR

Topics: Animals; Antibody Formation; Antigens; Germinal Center; Lysophospholipids; Mice; Sepsis

The high mortality seen in sepsis is caused by a systemic hypotension in part owing to a drastic increase in vascular permeability accompanied by a loss of pericytes. As has been shown previously, pericyte retention in the perivascular niche during sepsis can enhance the integrity of the vasculature and promote survival via recruitment of adhesion proteins such as VE-cadherin and N-cadherin. Sphingosine-1-phosphate (S1P) represents a lipid mediator regulating the deposition of the crucial adhesion molecule VE-cadherin at sites of interendothelial adherens junctions and of N-cadherin at endothelial-pericyte adherens junctions. Furthermore, in septic patients, S1P plasma levels are decreased and correlate with mortality in an indirectly proportional way. In the present study, we investigated the potential of S1P to ameliorate a lipopolysaccharide-induced septic hypercirculation in mice. Here we establish S1P as an antagonist of pericyte loss, vascular hyperpermeability, and systemic hypotension, resulting in an increased survival in mice. During sepsis S1P preserved VE-cadherin and N-cadherin deposition, mediated by a reduction of Src and cadherin phosphorylation. At least in part, this effect is mediated by a reduction of globular actin and a subsequent increase in nuclear translocation of MRTF-A (myocardin-related transcription factor A). These findings indicate that S1P may counteract pericyte loss and microvessel disassembly during sepsis and additionally emphasize the importance of pericyte-endothelial interactions to stabilize the vasculature.

Topics: Animals; Inflammation; Lipopolysaccharides; Lysophospholipids; Mice, Inbred C57BL; Pericytes; rhoA GTP-Binding Protein; Sepsis; Sphingosine; Trans-Activators

Acute ethanol intoxication increases the risk of sepsis and aggravates the symptoms of sepsis and lung injury. Therefore, this study aimed to explore whether sphingosine kinase 1 (SphK1)/sphingosine-1-phosphate (S1P)/S1P receptor 1 (S1PR1) signaling pathway functions in lung injury caused by acute ethanol intoxication-enhanced sepsis, as well as its underlying mechanism. The acute ethanol intoxication model was simulated by intraperitoneally administering mice with 32% ethanol solution, and cecal ligation and puncture (CLP) was used to construct the sepsis model. The lung tissue damage was observed by hematoxylin-eosin (H&E) staining, and the wet-to-dry (W/D) ratio was used to evaluate the degree of pulmonary edema. Inflammatory cell counting and protein concentration in bronchoalveolar lavage fluid (BALF) were, respectively, detected by hemocytometer and bicinchoninic acid (BCA) method. The levels of tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-1β, and IL-18 in BALF were detected by their commercial enzyme-linked immunosorbent assay (ELISA) kits. The myeloperoxidase (MPO) activity and expression of apoptosis-related proteins and SphK1/S1P/S1PR1 pathway-related proteins were, respectively, analyzed by MPO ELISA kit and Western blot analysis. The cell apoptosis in lung tissues was observed by TUNEL assay. Acute ethanol intoxication (EtOH) decreased the survival rate of mice and exacerbated the lung injury caused by sepsis through increasing pulmonary vascular permeability, neutrophil infiltration, release of inflammatory factors, and cell apoptosis. In addition, EtOH could activate the SphK1/S1P/S1PR1 pathway in CLP mice. However, PF-543, as a specific inhibitor of SphK1, could partially reverse the deleterious effects on lung injury of CLP mice. PF-543 alleviated lung injury caused by sepsis in acute ethanol intoxication rats by suppressing the SphK1/S1P/S1PR1 signaling pathway.

Topics: Alcoholic Intoxication; Animals; Apoptosis; Cytokines; Disease Models, Animal; Enzyme Inhibitors; Inflammation Mediators; Lung; Lung Injury; Lysophospholipids; Male; Methanol; Mice, Inbred C57BL; Neutrophil Infiltration; Oxidative Stress; Phosphotransferases (Alcohol Group Acceptor); Pneumonia; Pulmonary Edema; Pyrrolidines; Sepsis; Signal Transduction; Sphingosine; Sphingosine-1-Phosphate Receptors; Sulfones

In sepsis, the protection of the vascular endothelium is essential and the maintenance of its function is critical to prevent further deterioration. High-density lipoprotein (HDL)-associated sphingosine-1-phosphate (S1P) is a bioactive lipid in plasma and its role in sepsis has not been extensively studied. This study aimed to investigate the effects of HDL-S1P on sepsis in cellular and animal models, as well as human plasma samples.. We established an animal model of sepsis with different severities achieved by caecal ligation and puncture (CLP) and lipopolysaccharide (LPS) injection, and then explored the relationship between HDL-S1P and lung endothelial dysfunction in vivo. To determine the effects of HDL-S1P in the pulmonary endothelium of septic rats, we then injected HDL-S1P into septic rats to find out if it can reduce the lung injury caused by sepsis. Further, we explored the mechanism in vitro by studying the role of S1P-specific receptor agonists and inhibitors in LPS-stimulated human umbilical vein endothelial cells. We also explored the relationship between plasma HDL-S1P content and sepsis severity in septic patients by analysing their plasma samples.. HDL-S1P concentrations in plasma were negatively correlated with endothelial functional damage in sepsis, both in the animal model and in the septic patients in our study. In vivo, HDL-S1P injection significantly reduced pulmonary oedema and endothelial leakage in septic rats. In vitro, cell experiments showed that HDL-S1P effectively protected the proliferation and migration abilities of endothelial cells, which could be partly explained by its biased activation of the S1P receptor 1.. Our study preliminary explored the function of HDL-S1P in sepsis in cellular and animal models, as well as human subjects. The results indicate HDL-S1P protected endothelial functions in septic patients. Thus, it has therapeutic potential and can be used for the clinical treatment of sepsis.

Topics: Aged, 80 and over; Animals; Apolipoproteins M; Cell Movement; Cell Proliferation; Endothelium; Epithelial Cells; Female; Gene Expression Regulation; Humans; Intercellular Adhesion Molecule-1; Lipoproteins, HDL; Lung Injury; Lysophospholipids; Male; Rats; Sepsis; Sphingosine

Topics: Humans; Lysophospholipids; Sepsis; Sphingosine

The hyperpermeability of gut-vascular barrier (GVB) plays a role in gut-derived sepsis. The goal of this study was to evaluate if berberine might improve hepatic apolipoprotein M (ApoM) generation and raise plasma ApoM level to protect the compromised GVB.. The compromised GVB was induced by sepsis. Hepatic ApoM mRNA and phosphoenolpyruvate carboxykinase (PEPCK) mRNA and plasma ApoM level were assayed by qRT-PCR and ELISA, respectively. The permeability of intestinal capillary in vivo and of rat intestinal microvascular endothelial cells (RIMECs) in vitro was assayed by FITC-dextran. The blood glucose was detected by a glucometer. Plasma insulin, TNF-α and IL-1β were assayed by ELISA. The plasmalemma vesicle-associated protein-1 (PV1), β-catenin and occludin in RIMECs were assayed by Western blot.. Sepsis decreased hepatic ApoM mRNA and plasma ApoM level, but raised hepatic PEPCK mRNA and plasma glucose, insulin, TNF-α, and IL-1β levels. The increased vascular endothelial permeability was abrogated by recombinant rat ApoM in vivo or ApoM-bound S1P in vitro. ApoM-bound S1P decreased PV1 but increased occludin and β-catenin expression in LPS-treated RIMECs. Berberine in a dose-dependent manner raised hepatic ApoM mRNA and plasma ApoM level, but decreased septic hyperglycemia, insulin resistance and plasma TNF-α and IL-1β levels. Berberine reduced sepsis-induced PEPCK and TLR4 mRNA overexpression in the liver.. This study demonstrated berberine inhibited TLR4-mediated hyperglycemia, insulin resistance and proinflammatory molecule production, thereby increasing ApoM gene expression and plasma ApoM. Berberine protected the damaged GVB via modulation of ApoM/S1P pathway.

Topics: Animals; Apolipoproteins M; Berberine; Capillary Permeability; Disease Models, Animal; Gastrointestinal Tract; Hep G2 Cells; Humans; Lysophospholipids; Male; Rats, Wistar; Sepsis; Signal Transduction; Sphingosine

FTY720 is a sphingosine 1 phosphate (S1P) receptor agonist approved for the treatment of multiple sclerosis, which is a chronic inflammatory autoimmune disorder. Sepsis is a complex syndrome associated with progressive endotoxemic developments, which finally leads to damage of multiple organs, including the heart. In critical patients, cardiovascular dysfunction due to sepsis is a major cause of death. Previous studies have shown an association between S1P and cardioprotection in the situation of ischemia reperfusion and myocardial infarction. Therefore, we will study the role of S1P towards endotoxic cardiomyocytes. Different doses of FTY720 were applied or not to endotoxic cardiomyocytes. The concentration of inflammatory cytokines, including tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, and IL-10 was measured by enzyme-linked immuno sorbent assay. Western blotting was used to analyze the downstream signaling pathways. We discovered that FTY720 reduced the levels of TNF-α and IL-6 through the NF-ΚB pathway, inhibited the expression of caspase-3, and activated both protein kinase B and extracellular signal-regulated kinase 1/2. Additionally, the activation of protein kinase B and extracellular signal-regulated kinase 1/2 could be inhibited by the S1P1 and S1P3 receptor antagonist vulcanized polyethylene23019. Therefore, we infer that S1P exerts a protective effect towards endotoxic cardiomyocytes by decreasing the levels of TNF-α and IL-6, regulating apoptotic and survival signaling pathway. The S1P1 and S1P3 receptors are involved in the prosurvival signal activation.

Topics: Animals; Apoptosis; Cell Line; Cytokines; Endotoxins; Fingolimod Hydrochloride; Lysophospholipids; MAP Kinase Signaling System; Myocytes, Cardiac; NF-kappa B; Rats; Sepsis; Sphingosine

We examined the efficacy of embryoid bodies from 6-day induced pluripotent stem cells an in vivo sepsis model. Injection of embryoid bodies to septic mice improved the condition of their lungs and significantly increased their survival rate. Although embryoid bodies secretedsphingosine-1-phosphate in vitro, its serum levels in mouse plasma were significantly reduced compared to that in the control (untreated mice receiving PBS). Low concentrations of sphingosine-1-phosphate protected endothelial cells, while high concentrations disrupted endothelial barrier integrity. Therefore, exogenous sphingosine-1-phosphate secreted by embryoid bodies during early stage of sepsis might down regulate endogenous production of sphingosine-1-phosphate. Inhibition of excessive sphingosine-1-phosphate release protects against endothelial injury and suppresses a vicious cycle of inflammatory reactions. The obtained results open new prospects in induced pluripotent stem cells-based therapy for sepsis.

Topics: Animals; Embryoid Bodies; Endothelial Cells; Hematopoietic Stem Cell Transplantation; Induced Pluripotent Stem Cells; Lysophospholipids; Male; Mice; Mice, Inbred C57BL; Peritonitis; Sepsis; Sphingosine

Endothelial barrier dysfunction is a hallmark in the pathogenesis of sepsis. Sphingosine-1-phosphate (S1P) has been proposed to be critically involved in the maintenance of endothelial barrier function predominately by activating S1P receptor-1 (S1P1). Previous studies have shown that the specific S1P1 agonist SEW2871 improves endothelial barrier function under inflammatory conditions. However, the effectiveness of SEW2871 and potential side effects remained largely unexplored in a clinically relevant model of sepsis. Therefore, this study aimed to evaluate the effects of SEW2871 in the Colon ascendens stent peritonitis (CASP) model.. Polymicrobial sepsis was induced in Sprague-Dawley rats using CASP model that enabled the monitoring of macro-hemodynamic parameters. Twelve hours after surgery, animals received either SEW2871 or sodium chloride. Mesenteric endothelial barrier function was evaluated 24 h after sepsis induction by intravital microscopy. Organ pathology was assessed in lungs. S1P levels, blood gas analyses, and blood values were measured at different time points. In parallel the effect of SEW2871 was evaluated in human dermal microvascular endothelial cells.. In vitro SEW2871 partially stabilized TNF-α-induced endothelial barrier breakdown. However, in vivo SEW2871 caused severe cardiac side effects in septic animals leading to an increased lethality. Sepsis-induced endothelial barrier dysfunction was not attenuated by SEW2871 as revealed by increased FITC-albumin extra-vasation, requirement of intravasal fluid replacement, and pulmonary edema. Interestingly, Sham-operated animals did not present any side effects after SEW2871 treatment.. Our study demonstrates that the application of SEW2871 causes severe cardiac side effects and cannot attenuate the inflammation-induced endothelial barrier breakdown in a clinically relevant sepsis model, suggesting that the time point of administration and the pro-inflammatory milieu play a pivotal role in the therapeutic benefit of SEW2871.

Topics: Animals; Disease Models, Animal; Humans; Lysophospholipids; Male; Oxadiazoles; Rats; Rats, Sprague-Dawley; Receptors, Lysosphingolipid; Sepsis; Sphingosine; Sphingosine-1-Phosphate Receptors; Thiophenes; Tumor Necrosis Factor-alpha

High-density lipoprotein (HDL) has been epidemiologically shown to be associated with the outcome of sepsis. One potential mechanism is that HDL possesses pleiotropic effects, such as anti-apoptosis, some of which can be ascribed to sphingosine 1-phosphate (S1P) carried on HDL via apolipoprotein M (apoM). Therefore, the aim of this study was to elucidate the roles of apoM/S1P in the consequent lethal conditions of sepsis, such as multiple organ failure caused by severe inflammation and/or disseminated intravascular coagulation.. In mice treated with lipopolysaccharide (LPS), both plasma apoM levels and the expression of apoM in the liver and kidney were suppressed. The overexpression of apoM improved the survival rate and ameliorated the elevated plasma alanine aminotransferase (ALT) and creatinine levels, while the knockout or knockdown of apoM deteriorated these parameters in mice treated with LPS. Treatment with VPC23019, an antagonist against S1P receptor 1 and 3, or LY294002, a PI3K inhibitor, partially reversed these protective properties arising from the overexpression of apoM. The overexpression of apoM inhibited the elevation of plasma plasminogen activator inhibitor-1, restored the phosphorylation of Akt, and induced anti-apoptotic changes in the liver, kidney and heart.. These results suggest that apoM possesses protective properties against LPS-induced organ injuries and could potentially be introduced as a novel therapy for the severe conditions that are consequent to sepsis.

Topics: Alanine Transaminase; Animals; Apolipoproteins M; Clustered Regularly Interspaced Short Palindromic Repeats; Creatinine; Disease Models, Animal; Disseminated Intravascular Coagulation; Human Umbilical Vein Endothelial Cells; Humans; Inflammation; Lipopolysaccharides; Lipoproteins, HDL; Lysophospholipids; Mice; Mice, Inbred C57BL; Mice, Knockout; Multiple Organ Failure; Phosphoserine; Receptors, Lysosphingolipid; Sepsis; Sphingosine

Endothelial dysfunction contributes to sepsis outcome. Metabolic phenotypes associated with endothelial dysfunction are not well characterised in part due to difficulties in assessing endothelial metabolism in situ. Here, we describe the construction of iEC2812, a genome scale metabolic reconstruction of endothelial cells and its application to describe metabolic changes that occur following endothelial dysfunction. Metabolic gene expression analysis of three endothelial subtypes using iEC2812 suggested their similar metabolism in culture. To mimic endothelial dysfunction, an in vitro sepsis endothelial cell culture model was established and the metabotypes associated with increased endothelial permeability and glycocalyx loss after inflammatory stimuli were quantitatively defined through metabolomics. These data and transcriptomic data were then used to parametrize iEC2812 and investigate the metabotypes of endothelial dysfunction. Glycan production and increased fatty acid metabolism accompany increased glycocalyx shedding and endothelial permeability after inflammatory stimulation. iEC2812 was then used to analyse sepsis patient plasma metabolome profiles and predict changes to endothelial derived biomarkers. These analyses revealed increased changes in glycan metabolism in sepsis non-survivors corresponding to metabolism of endothelial dysfunction in culture. The results show concordance between endothelial health and sepsis survival in particular between endothelial cell metabolism and the plasma metabolome in patients with sepsis.

Topics: Biomarkers; Cell Line; Chromatography, High Pressure Liquid; Endothelial Cells; Fatty Acids; gamma-Aminobutyric Acid; Glycocalyx; Human Umbilical Vein Endothelial Cells; Humans; Interferon-gamma; Kynurenine; Lipopolysaccharides; Lysophospholipids; Metabolome; Models, Biological; Nitric Oxide; Permeability; Polysaccharides; Prostaglandin D2; Sepsis; Sphingosine; Survival Analysis; Tryptophan

Sepsis mouse models revealed thymus atrophy, characterised by decreased thymus weight and loss of thymocytes due to apoptosis. Mice suffered from lymphopenia, a lack of T cells in the periphery, which attenuates their ability to fight against recurring and secondary infections during sepsis progression. Key players in thymus atrophy are IL-6, which is directly involved in thymus involution, and the sphingosine-1-phosphate - sphingosine-1-phosphate receptor 1 signaling, influencing thymocytes emigration. In healthy individuals a sphingosine-1-phosphate (S1P) gradient from lymphoid organs to the circulatory system serves as signal for mature T cell egress. In the present study we investigated, whether inhibition of S1P generation improves thymus involution. In sepsis, induced by cecal ligation and puncture (CLP), S1P in the thymus increased, while it decreased in serum, thus disrupting the naturally occurring S1P gradient. As a potential source of S1P we identified increased numbers of apoptotic cells in the thymic cortex of septic mice. Pharmacological inhibition of the S1P generating sphingosine kinases, by 4- [[4-(4-Chlorophenyl)-2-thiazolyl]amino]phenol (SK I-II), administered directly following CLP, prevented thymus atrophy. This was reflected by lymphocytosis, diminished apoptosis, decreased IL-6 expression, and an unaltered thymus weight. In addition SK I-II-treatment preserved the S1P balance and prevented S1P-dependent internalization of the sphingosine-1-phosphate receptor 1. Our data suggest that inhibition of sphingosine kinase and thus, S1P generation during sepsis restores thymic T cell egress, which might improve septic outcome.

Topics: Aminophenols; Animals; Apoptosis; Atrophy; Cecum; Disease Models, Animal; Interleukin-6; Lymphocytosis; Lymphopenia; Lysophospholipids; Male; Mice; Mice, Inbred C57BL; Phosphotransferases (Alcohol Group Acceptor); Receptors, Lysosphingolipid; Sepsis; Sphingosine; Thiazoles; Thymocytes; Thymus Gland

Sphingosine 1-phosphate (S1P) is an important regulator of vascular integrity and immune cell migration, carried in plasma by high-density lipoprotein (HDL)-associated apolipoprotein M (apoM) and by albumin. In sepsis, the protein and lipid composition of HDL changes dramatically. The aim of this study was to evaluate changes in S1P and its carrier protein apoM during sepsis. For this purpose, plasma samples from both human sepsis patients and from an experimental Escherichia coli sepsis model in baboons were used. In the human sepsis cohort, previously studied for apoM, plasma demonstrated disease-severity correlated decreased S1P levels, the profile mimicking that of plasma apoM. In the baboons, a similar disease-severity dependent decrease in plasma levels of S1P and apoM was observed. In the lethal E. coli baboon sepsis, S1P decreased already within 6-8 hrs, whereas the apoM decrease was seen later at 12-24 hrs. Gel filtration chromatography of plasma from severe human or baboon sepsis on Superose 6 demonstrated an almost complete loss of S1P and apoM in the HDL fractions. S1P plasma concentrations correlated with the platelet count but not with erythrocytes or white blood cells. The liver mRNA levels of apoM and apoA1 decreased strongly upon sepsis induction and after 12 hr both were almost completely lost. In conclusion, during septic challenge, the plasma levels of S1P drop to very low levels. Moreover, the liver synthesis of apoM decreases severely and the plasma levels of apoM are reduced. Possibly, the decrease in S1P contributes to the decreased endothelial barrier function observed in sepsis.

Topics: Animals; Apolipoproteins; Apolipoproteins M; Blood Platelets; Case-Control Studies; Chromatography, Gel; Colony Count, Microbial; Erythrocytes; Escherichia coli; Humans; Kidney; Leukocytes; Lipocalins; Lysophospholipids; Papio; RNA, Messenger; Sepsis; Sphingosine; Transcription, Genetic

Serum levels of the lipid mediator sphingosine-1-phosphate (S1P) are reduced in septic patients and are inversely associated with disease severity. We show that serum S1P is reduced in human sepsis and in murine models of sepsis. We then investigated whether pharmacological or genetic approaches that alter serum S1P may attenuate cardiac dysfunction and whether S1P signaling might serve as a novel theragnostic tool in sepsis. Mice were challenged with lipopolysaccharide and peptidoglycan (LPS/PepG). LPS/PepG resulted in an impaired systolic contractility and reduced serum S1P. Administration of the immunomodulator FTY720 increased serum S1P, improved impaired systolic contractility and activated the phosphoinositide 3-kinase (PI3K)-pathway in the heart. Cardioprotective effects of FTY720 were abolished following administration of a S1P receptor 2 (S1P2) antagonist or a PI3K inhibitor. Sphingosine kinase-2 deficient mice had higher endogenous S1P levels and the LPS/PepG-induced impaired systolic contractility was attenuated in comparison with wild-type mice. Cardioprotective effects of FTY720 were confirmed in polymicrobial sepsis. We show here for the first time that the impaired left ventricular systolic contractility in experimental sepsis is attenuated by FTY720. Mechanistically, our results indicate that activation of S1P2 by increased serum S1P and the subsequent activation of the PI3K-Akt survival pathway significantly contributes to the observed cardioprotective effect of FTY720.

Topics: Animals; Cardiotonic Agents; Disease Models, Animal; Fingolimod Hydrochloride; Heart; Humans; Inflammation; Lipopolysaccharides; Lysophospholipids; Male; Mice; Mice, Inbred C57BL; Myocardial Contraction; Myocardium; Peptidoglycan; Phosphatidylinositol 3-Kinases; Phosphorylation; Pilot Projects; Receptors, Lysosphingolipid; Sepsis; Sphingosine

Sepsis is a systemic inflammatory response to pathogens and a leading cause of hospital related mortality worldwide. Sphingosine 1-phosphate (S1P) regulates multiple cellular processes potentially involved in the pathogenesis of sepsis, including antigen presentation, lymphocyte egress, and maintenance of vascular integrity. We thus explored the impact of manipulating S1P signaling in experimental polymicrobial sepsis in mice. Administration of 4-deoxypyridoxine (DOP), an inhibitor of the S1P-degrading enzyme S1P-lyase, or of the sphingosine analog FTY720 that serves as an S1P receptor agonist after phosphorylation ameliorated morbidity, improved recovery from sepsis in surviving mice, and reduced sepsis-elicited hypothermia and body weight loss. Treated mice developed lymphopenia, leading to an accumulation of lymphocytes in peripheral lymph nodes, and reduced bacterial burden in liver, but not in blood. Sepsis-induced upregulation of mRNA expression of cytokines in spleen remained unchanged, but reduction of IL-6, TNF-α, MCP-1, and IL-10 in plasma was evident. DOP and FTY720 treatment significantly reduced levels of Evans blue leakage from blood into liver and lung, decreased hematocrit values, and lowered plasma levels of VEGF-A in septic mice. Collectively, our results indicate that modulation of S1P signaling showed a protective phenotype in experimental sepsis by modulating vascular and immune functions.

Topics: Animals; Capillary Permeability; Cells, Cultured; Cytokines; Fingolimod Hydrochloride; Immunomodulation; Lysophospholipids; Membrane Proteins; Mice; Mice, Inbred C57BL; Phosphoric Monoester Hydrolases; Pyridoxine; Receptors, Lysosphingolipid; Sepsis; Signal Transduction; Sphingosine; Vascular Endothelial Growth Factor A

Sphingosine-1-phosphate (S1P) is a signaling lipid that regulates pathophysiological processes involved in sepsis progression, including endothelial permeability, cytokine release, and vascular tone. The aim of this study was to investigate whether serum-S1P concentrations are associated with disease severity in patients with sepsis.. This single-center prospective-observational study includes 100 patients with systemic inflammatory response syndrome (SIRS) plus infection (n = 40), severe sepsis (n = 30), or septic shock (n = 30) and 214 healthy blood donors as controls. Serum-S1P was measured by mass spectrometry. Blood parameters, including C-reactive protein (CRP), procalcitonin (PCT), interleukin-6 (IL-6), lactate, and white blood cells (WBCs), were determined by routine assays. The Sequential Organ Failure Assessment (SOFA) score was generated and used to evaluate disease severity.. Serum-S1P concentrations were lower in patients than in controls (P < 0.01), and the greatest difference was between the control and the septic shock groups (P < 0.01). Serum-S1P levels were inversely correlated with disease severity as determined by the SOFA score (P < 0.01) as well as with IL-6, PCT, CRP, creatinine, lactate, and fluid balance. A receiver operating characteristic analysis for the presence or absence of septic shock revealed equally high sensitivity and specificity for S1P compared with the SOFA score. In a multivariate logistic regression model calculated for prediction of septic shock, S1P emerged as the strongest predictor (P < 0.001).. In patients with sepsis, serum-S1P levels are dramatically decreased and are inversely associated with disease severity. Since S1P is a potent regulator of endothelial integrity, low S1P levels may contribute to capillary leakage, impaired tissue perfusion, and organ failure in sepsis.

Topics: Adult; Female; Germany; Humans; Lysophospholipids; Male; Middle Aged; Multiple Organ Failure; Prospective Studies; Sepsis; Severity of Illness Index; Sphingosine

Increased vascular leakage leading to hypovolaemia and tissue oedema is common in severe sepsis. Hypovolaemia together with oedema formation may contribute to hypoxia and result in multiorgan failure and death. To improve treatment during sepsis, a potential therapeutic target may be to reduce the vascular leakage. Substances affecting the endothelial barrier are interesting in this respect, as it is suggested that increase in vascular leakage depends on reorganisation of the endothelial cells and breakdown of the endothelial barrier. The agonist of the bioactive lipid sphingosine-1-phosphate, FTY720, has been shown to modulate the integrity of the endothelium and reduce permeability both in vitro and in vivo. The aim of the present study was to determine if FTY720 could reduce the loss of plasma volume during experimental sepsis in rats.. Sepsis was induced by ligation and incision of the caecum in the rat. Plasma volume was determined before and 4.5 h after induction of sepsis by a dilution technique using (125) I-labelled albumin.. FTY720 in a dose of 0.2 mg/kg reduced the loss of plasma during sepsis by approximately 30% compared with vehicle, without any adverse effects on haemodynamic and physiological parameters. The increase in hematocrit and haemoglobin concentration was also found to be higher in the vehicle group.. FTY720 in a dose without haemodynamic side effects reduces loss of plasma volume during experimental sepsis most likely because of reduction in permeability and may therefore be beneficial in sepsis.

Topics: Animals; Capillary Leak Syndrome; Capillary Permeability; Cecum; Disease Models, Animal; Diuresis; Drug Evaluation, Preclinical; Edema; Endothelium, Vascular; Fingolimod Hydrochloride; Hematocrit; Hemodynamics; Hemoglobins; Intestinal Perforation; Lysophospholipids; Male; Plasma Volume; Propylene Glycols; Random Allocation; Rats; Rats, Sprague-Dawley; Sepsis; Sphingosine

The genetic mechanisms underlying the susceptibility to acute respiratory distress syndrome (ARDS) are poorly understood. We previously demonstrated that sphingosine 1-phosphate (S1P) and the S1P receptor S1PR3 are intimately involved in lung inflammatory responses and vascular barrier regulation. Furthermore, plasma S1PR3 protein levels were shown to serve as a biomarker of severity in critically ill ARDS patients. This study explores the contribution of single nucleotide polymorphisms (SNPs) of the S1PR3 gene to sepsis-associated ARDS. S1PR3 SNPs were identified by sequencing the entire gene and tagging SNPs selected for case-control association analysis in African- and ED samples from Chicago, with independent replication in a European case-control study of Spanish individuals. Electrophoretic mobility shift assays, luciferase activity assays, and protein immunoassays were utilized to assess the functionality of associated SNPs. A total of 80 variants, including 29 novel SNPs, were identified. Because of limited sample size, conclusive findings could not be drawn in African-descent ARDS subjects; however, significant associations were found for two promoter SNPs (rs7022797 -1899T/G; rs11137480 -1785G/C), across two ED samples supporting the association of alleles -1899G and -1785C with decreased risk for sepsis-associated ARDS. In addition, these alleles significantly reduced transcription factor binding to the S1PR3 promoter; reduced S1PR3 promoter activity, a response particularly striking after TNF-α challenge; and were associated with lower plasma S1PR3 protein levels in ARDS patients. These highly functional studies support S1PR3 as a novel ARDS candidate gene and a potential target for individualized therapy.

Topics: Base Sequence; Biomarkers; Case-Control Studies; Electrophoretic Mobility Shift Assay; Female; Genetic Association Studies; Genetic Predisposition to Disease; Genotype; Humans; Lysophospholipids; Male; Middle Aged; Molecular Sequence Data; Polymorphism, Single Nucleotide; Promoter Regions, Genetic; Receptors, Lysosphingolipid; Respiratory Distress Syndrome; Sepsis; Sequence Analysis, DNA; Sphingosine; Sphingosine-1-Phosphate Receptors

Activated protein C (APC) exerts anticoagulant effects via inactivation of factors Va and VIIIa and cytoprotective effects via protease activated receptor (PAR)1. Inhibition of endogenous APC in endotoxemia and sepsis results in exacerbation of coagulation and inflammation, with consequent enhanced lethality.. We here sought to dissect the distinct roles of the anticoagulant and cytoprotective functions of endogenous APC in severe Gram-negative pneumonia-derived sepsis (melioidosis).. We infected wild-type (WT) mice with Burkholderia pseudomallei, a common sepsis pathogen in southeast Asia, and treated them with antibodies inhibiting both the anticoagulant and cytoprotective functions of APC (MPC1609) or the anticoagulant functions of APC (MAPC1591) only. Additionally, we administered SEW2871 (stimulating the S1P1-pathway downstream from PAR1) to control and MPC1609-treated mice.. MPC1609, but not MAPC1591, significantly worsened survival, increased coagulation activation, facilitated bacterial growth and dissemination and enhanced the inflammatory response. The effects of MPC1609 could not be reversed by SEW2871, suggesting that S1P1 does not play a major role in this model.. These results suggest that the mere inhibition of the anticoagulant function of APC does not interfere with its protective role during Gram-negative pneumosepsis, suggesting a more prominent role for cytoprotective effects of APC .

Topics: Animals; Antibodies, Monoclonal; Bacterial Load; Blood Coagulation; Burkholderia pseudomallei; Cytokines; Cytoprotection; Disease Models, Animal; Female; Inflammation; Inflammation Mediators; Liver; Lung; Lysophospholipids; Melioidosis; Mice; Mice, Inbred C57BL; Oxadiazoles; Protein C; Receptor, PAR-1; Sepsis; Signal Transduction; Sphingosine; Thiophenes; Time Factors

Sepsis and acute lung injury (ALI) have devastatingly high mortality rates. Both are associated with increased vascular leak, a process regulated by complex molecular mechanisms.. We hypothesized that integrin αvβ3 could be an important determinant of vascular leak and endothelial permeability in sepsis and ALI.. β3 subunit knockout mice were tested for lung vascular leak after endotracheal LPS, and systemic vascular leak and mortality after intraperitoneal LPS and cecal ligation and puncture. Possible contributory effects of β3 deficiency in platelets and other hematopoietic cells were excluded by bone marrow reconstitution experiments. Endothelial cells treated with αvβ3 antibodies were evaluated for sphingosine-1 phosphate (S1P)–mediated alterations in barrier function, cytoskeletal arrangement, and integrin localization.. β3 knockout mice had increased vascular leak and pulmonary edema formation after endotracheal LPS, and increased vascular leak and mortality after intraperitoneal LPS and cecal ligation and puncture. In endothelial cells, αvβ3 antibodies inhibited barrier-enhancing and cortical actin responses to S1P. Furthermore, S1P induced translocation of αvβ3 from discrete focal adhesions to cortically distributed sites through Gi- and Rac1-mediated pathways. Cortical αvβ3 localization after S1P was decreased by αvβ3 antibodies, suggesting that ligation of the αvβ3 with its extracellular matrix ligands is required to stabilize cortical αvβ3 focal adhesions.. Our studies identify a novel mechanism by which αvβ3 mitigates increased vascular leak, a pathophysiologic function central to sepsis and ALI. These studies suggest that drugs designed to block αvβ3 may have the unexpected side effect of intensifying sepsis- and ALI-associated vascular endothelial leak.

Topics: Actins; Acute Lung Injury; Animals; Disease Models, Animal; Endothelium, Vascular; Female; Integrin alphaVbeta3; Lysophospholipids; Mice; Mice, Knockout; Pulmonary Edema; Sepsis; Signal Transduction; Sphingosine; Vascular Diseases

The complement anaphylatoxin C5a has a pathogenetic role in endotoxin-induced lung inflammatory injury by regulating phagocytic cell migration and activation. Endotoxin and C5a activate the enzyme sphingosine kinase (Sphk) 1 to generate the signaling lipid sphingosine-1-phosphate (S1P), a critical regulator of phagocyte function. We assessed the function of Sphk1 and S1P in experimental lung inflammatory injury and determined their roles in anaphylatoxin receptor signaling and on the expression of the two C5a receptors, C5aR (CD88) and C5L2, on phagocytes. We report that Sphk1 gene deficient (Sphk1(-/-)) mice had augmented lung inflammatory response to endotoxin compared to wild type mice. Sphk1 was required for C5a-mediated reduction in cytokine and chemokine production by macrophages. Moreover, neutrophils from Sphk1(-/-) mice failed to upregulate the anaphylatoxin receptor C5L2 in response to LPS. Exogenous S1P restored C5L2 cell surface expression of Sphk1(-/-) mouse neutrophils to wild type levels but had no effect on cell surface expression of the other anaphylatoxin receptor, CD88. These results provide the first genetic evidence of the crucial role of Sphk1 in regulating the balance between expression of CD88 and C5L2 in phagocytes. S1P-mediated up-regulation of C5L2 is a novel therapeutic target for mitigating endotoxin-induced lung inflammatory injury.

Topics: Anaphylatoxins; Animals; Bone Marrow; Cytokines; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Lipopolysaccharides; Lysophospholipids; Macrophages; Mice; Mice, Inbred C57BL; Mice, Knockout; Neutrophils; Phosphorylation; Phosphotransferases (Alcohol Group Acceptor); Pneumonia; Receptor, Anaphylatoxin C5a; Receptors, Chemokine; Sepsis; Signal Transduction; Sphingosine

Activated protein C (APC), the only FDA-approved biotherapeutic drug for sepsis, possesses anticoagulant, antiinflammatory, and barrier-protective activities. However, the mechanisms underlying its anti-inflammatory functions are not well defined. Here, we report that the antiinflammatory activity of APC on macrophages is dependent on integrin CD11b/CD18, but not on endothelial protein C receptor (EPCR). We showed that CD11b/CD18 bound APC within specialized membrane microdomains/lipid rafts and facilitated APC cleavage and activation of protease-activated receptor-1 (PAR1), leading to enhanced production of sphingosine-1-phosphate (S1P) and suppression of the proinflammatory response of activated macrophages. Deletion of the gamma-carboxyglutamic acid domain of APC, a region critical for its anticoagulant activity and EPCR-dependent barrier protection, had no effect on its antiinflammatory function. Genetic inactivation of CD11b, PAR1, or sphingosine kinase-1, but not EPCR, abolished the ability of APC to suppress the macrophage inflammatory response in vitro. Using an LPS-induced mouse model of lethal endotoxemia, we showed that APC administration reduced the mortality of wild-type mice, but not CD11b-deficient mice. These data establish what we believe to be a novel mechanism underlying the antiinflammatory activity of APC in the setting of endotoxemia and provide clear evidence that the antiinflammatory function of APC is distinct from its barrier-protective function and anticoagulant activities.

Topics: 1-Carboxyglutamic Acid; Animals; Endotoxemia; Integrins; Lipopolysaccharides; Lysophospholipids; Membrane Microdomains; Mice; Mice, Inbred C57BL; Mice, Knockout; Phosphotransferases (Alcohol Group Acceptor); Protein C; Sepsis; Sphingosine