Targeted rescue of synaptic plasticity improves cognitive decline in sepsis-associated encephalopathy.

Sepsis-associated encephalopathy (SAE) is a frequent complication of severe systemic infection resulting in delirium, premature death, and long-term cognitive impairment. We closely mimicked SAE in a murine peritoneal contamination and infection (PCI) model. We found long-lasting synaptic pathology in the hippocampus including defective long-term synaptic plasticity, reduction of mature neuronal dendritic spines, and severely affected excitatory neurotransmission. Genes related to synaptic signaling, including the gene for activity-regulated cytoskeleton-associated protein (Arc/Arg3.1) and members of the transcription-regulatory EGR gene family, were downregulated. At the protein level, ARC expression and mitogen-activated protein kinase signaling in the brain were affected. For targeted rescue we used adeno-associated virus-mediated overexpression of ARC in the hippocampus in vivo. This recovered defective synaptic plasticity and improved memory dysfunction. Using the enriched environment paradigm as a non-invasive rescue intervention, we found improvement of defective long-term potentiation, memory, and anxiety. The beneficial effects of an enriched environment were accompanied by an increase in brain-derived neurotrophic factor (BDNF) and ARC expression in the hippocampus, suggesting that activation of the BDNF-TrkB pathway leads to restoration of the PCI-induced reduction of ARC. Collectively, our findings identify synaptic pathomechanisms underlying SAE and provide a conceptual approach to target SAE-induced synaptic dysfunction with potential therapeutic applications to patients with SAE.

Aß-Induced Alterations in Membrane Lipids Occur before Synaptic Loss Appears.

Loss of active synapses and alterations in membrane lipids are crucial events in physiological aging as well as in neurodegenerative disorders. Both are related to the abnormal aggregation of amyloid-beta (Aß) species, generally known as amyloidosis. There are two major known human Aß species: Aß(1-40) and Aß(1-42). However, which of these species have more influence on active synapses and membrane lipids is still poorly understood. Additionally, the time-dependent effect of Aß species on alterations in membrane lipids of hippocampal neurones and glial cells remains unknown. Therefore, our study contributes to a better understanding of the role of Aß species in the loss of active synapses and the dysregulation of membrane lipids in vitro. We showed that Aß(1-40) or Aß(1-42) treatment influences membrane lipids before synaptic loss appears and that the loss of active synapses is not dependent on the Aß species. Our lipidomic data analysis showed early changes in specific lipid classes such as sphingolipid and glycerophospholipid neurones. Our results underscore the potential role of lipids as a possible early diagnostic biomarker in amyloidosis-related disorders.

Low-dose hydrocortisone prolongs survival in a lethal sepsis model in adrenalectomized rats.

BACKGROUND: Controversial clinical findings of low-dose hydrocortisone supplementation in septic shock led us to investigate the impact of administration in lethal septic shock in adrenalectomized rats. MATERIALS AND METHODS: After preliminary experiments, to define the intravenous dose of hydrocortisone delivered in bilaterally adrenalectomized rats with serum cortisol level similar to sham rats, survival experiments were run in 75 rats after intraperitoneal challenge with Escherichia coli. Rats were treated with placebo, ertapenem, hydrocortisone, and a combination. Sacrifice experiments were run to measure gene transcripts in whole blood and in the liver and to assess cytokine stimulation of splenocytes and tissue overgrowth. RESULTS: The combination of hydrocortisone and ertapenem was superior to any single treatment and mandatory to achieve survival benefit. Splenocytes from infected rats had decreased production of tumor necrosis factor-alpha (TNFα); this was reversed with hydrocortisone treatment. Hydrocortisone increased the expression of TNF, Il1r2, and Hdac4 and decreased that of Dnmt3a. Bacterial burden of E. coli in kidney was decreased after hydrocortisone treatment. CONCLUSIONS: Low dose of hydrocortisone is a mandatory adjunctive to antimicrobial therapy in a rat model of septic shock after bilateral adrenalectomy. The mechanism of action is related to reversal of sepsis-induced immunosuppression through interaction with histone deacetylases and de novo DNA methyltransferases.

Acid Sphingomyelinase Inhibition Stabilizes Hepatic Ceramide Content and Improves Hepatic Biotransformation Capacity in a Murine Model of Polymicrobial Sepsis.

Liver dysfunction during sepsis is an independent risk factor leading to increased mortality rates. Specifically, dysregulation of hepatic biotransformation capacity, especially of the cytochrome P450 (CYP) system, represents an important distress factor during host response. The activity of the conserved stress enzyme sphingomyelin phosphodiesterase 1 (SMPD1) has been shown to be elevated in sepsis patients, allowing for risk stratification. Therefore, the aim of the present study was to investigate whether SMPD1 activity has an impact on expression and activity of different hepatic CYP enzymes using an animal model of polymicrobial sepsis. Polymicrobial sepsis was induced in SMPD1 wild-type and heterozygous mice and hepatic ceramide content as well as CYP mRNA, protein expression and enzyme activities were assessed at two different time points, at 24 h, representing the acute phase, and at 28 days, representing the post-acute phase of host response. In the acute phase of sepsis, SMPD1+/+ mice showed an increased hepatic C16- as well as C18-ceramide content. In addition, a downregulation of CYP expression and activities was detected. In SMPD1+/- mice, however, no noticeable changes of ceramide content and CYP expression and activities during sepsis could be observed. After 28 days, CYP expression and activities were normalized again in all study groups, whereas mRNA expression remained downregulated in SMPD+/+ animals. In conclusion, partial genetic inhibition of SMPD1 stabilizes hepatic ceramide content and improves hepatic monooxygenase function in the acute phase of polymicrobial sepsis. Since we were also able to show that the functional inhibitor of SMPD1, desipramine, ameliorates downregulation of CYP mRNA expression and activities in the acute phase of sepsis in wild-type mice, SMPD1 might be an interesting pharmacological target, which should be further investigated.

Increased lipogenesis in spite of upregulated hepatic 5'AMP-activated protein kinase in human non-alcoholic fatty liver.

AIMS: Molecular adaptations in human non-alcoholic fatty liver disease (NAFLD) are incompletely understood. This study investigated the main gene categories related to hepatic de novo lipogenesis and lipid oxidation capacity. METHODS: Liver specimens of 48 subjects were histologically classified according to steatosis severity. In-depth analyses were undertaken using real-time polymerase chain reaction, immunoblotting, and immunohistochemistry. Lipid profiles were analyzed by gas chromatography/flame ionization detection, and effects of key fatty acids were studied in primary human hepatocytes. RESULTS: Real-time polymerase chain reaction, immunoblotting, and immunohistochemistry indicated 5'AMP-activated protein kinase (AMPK) to be increased with steatosis score ≥ 2 (all P < 0.05), including various markers of de novo lipogenesis and lipid degradation (all P < 0.05). Regarding endoplasmic reticulum stress, X-Box binding protein-1 (XBP1) was upregulated in steatosis score ≥ 2 (P = 0.029) and correlated with plasma palmitate (r = 0.34; P = 0.035). Palmitate incubation of primary human hepatocytes increased XBP1 and downstream stearoyl CoA desaturase-1 mRNA expression (both P < 0.05). Moreover, plasma and liver tissue exposed a NAFLD-related lipid profile with reduced polyunsaturated/saturated fatty acid ratio, increased palmitate and palmitoleate, and elevated lipogenesis and desaturation indices with steatosis score ≥ 2 (all P < 0.05). CONCLUSION: In humans with advanced fatty liver disease, hepatic AMPK protein is upregulated, potentially in a compensatory manner. Moreover, pathways of lipid synthesis and degradation are co-activated in subjects with advanced steatosis. Palmitate may drive lipogenesis by activating XBP1-mediated endoplasmic reticulum stress and represent a target for future dietary or pharmacological intervention.

Adjustment of Dysregulated Ceramide Metabolism in a Murine Model of Sepsis-Induced Cardiac Dysfunction.

Cardiac dysfunction, in particular of the left ventricle, is a common and early event in sepsis, and is strongly associated with an increase in patients' mortality. Acid sphingomyelinase (SMPD1)-the principal regulator for rapid and transient generation of the lipid mediator ceramide-is involved in both the regulation of host response in sepsis as well as in the pathogenesis of chronic heart failure. This study determined the degree and the potential role to which SMPD1 and its modulation affect sepsis-induced cardiomyopathy using both genetically deficient and pharmacologically-treated animals in a polymicrobial sepsis model. As surrogate parameters of sepsis-induced cardiomyopathy, cardiac function, markers of oxidative stress as well as troponin I levels were found to be improved in desipramine-treated animals, desipramine being an inhibitor of ceramide formation. Additionally, ceramide formation in cardiac tissue was dysregulated in SMPD1+/+ as well as SMPD1-/- animals, whereas desipramine pretreatment resulted in stable, but increased ceramide content during host response. This was a result of elevated de novo synthesis. Strikingly, desipramine treatment led to significantly improved levels of surrogate markers. Furthermore, similar results in desipramine-pretreated SMPD1-/- littermates suggest an SMPD1-independent pathway. Finally, a pattern of differentially expressed transcripts important for regulation of apoptosis as well as antioxidative and cytokine response supports the concept that desipramine modulates ceramide formation, resulting in beneficial myocardial effects. We describe a novel, protective role of desipramine during sepsis-induced cardiac dysfunction that controls ceramide content. In addition, it may be possible to modulate cardiac function during host response by pre-conditioning with the Food and Drug Administration (FDA)-approved drug desipramine.

Acid Sphingomyelinase Promotes Endothelial Stress Response in Systemic Inflammation and Sepsis.

The pathophysiology of sepsis involves activation of acid sphingomyelinase (SMPD1) with subsequent generation of the bioactive mediator ceramide. We herein evaluated the hypothesis that the enzyme exerts biological effects in endothelial stress response. Plasma-secreted sphingomyelinase activity, ceramide generation and lipid raft formation were measured in human microcirculatory endothelial cells (HMEC-1) stimulated with serum obtained from sepsis patients. Clustering of receptors relevant for signal transduction was studied by immuno staining. The role of SMPD1 for macrodomain formation was tested by pharmacological inhibition. To confirm the involvement of the stress enzyme, direct inhibitors (amino bisphosphonates) and specific downregulation of the gene was tested with respect to ADAMTS13 expression and cytotoxicity. Plasma activity and amount of SMPD1 were increased in septic patients dependent on clinical severity. Increased breakdown of sphingomyelin to ceramide in HMECs was observed following stimulation with serum from sepsis patients in vitro. Hydrolysis of sphingomyelin, clustering of receptor complexes, such as the CD95L/Fas-receptor, as well as formation of ceramide enriched macrodomains was abrogated using functional inhibitors (desipramine and NB6). Strikingly, the stimulation of HMECs with serum obtained from sepsis patients or mixture of proinflammatory cytokines resulted in cytotoxicity and ADAMTS13 downregulation which was abrogated using desipramine, amino bisphosphonates and genetic inhibitors. SMPD1 is involved in the dysregulation of ceramide metabolism in endothelial cells leading to macrodomain formation, cytotoxicity and downregulation of ADAMTS13 expression. Functional inhibitors, such as desipramine, are capable to improve endothelial stress response during sepsis and might be considered as a pharmacological treatment strategy to favor the outcome.

An Integrated Clinico-transcriptomic Approach Identifies a Central Role of the Heme Degradation Pathway for Septic Complications after Trauma.

OBJECTIVE: The present study was aimed to identify mechanisms linked to complicated courses and adverse events after severe trauma by a systems biology approach. SUMMARY BACKGROUND DATA: In severe trauma, overwhelming systemic inflammation can result in additional damage and the development of complications, including sepsis. METHODS: In a prospective, longitudinal single-center study, RNA samples from circulating leukocytes from patients with multiple injury (injury severity score ≥17 points; n = 81) were analyzed for dynamic changes in gene expression over a period of 21 days by whole-genome screening (discovery set; n = 10 patients; 90 samples) and quantitative RT-PCR (validation set; n = 71 patients, 517 samples). Multivariate correlational analysis of transcripts and clinical parameters was used to identify mechanisms related to sepsis. RESULTS: Transcriptome profiling of the discovery set revealed the strongest changes between patients with either systemic inflammation or sepsis in gene expression of the heme degradation pathway. Using quantitative RT-PCR analyses (validation set), the key components haptoglobin (HP), cluster of differentiation (CD) 163, heme oxygenase-1 (HMOX1), and biliverdin reductase A (BLVRA) showed robust changes following trauma. Upregulation of HP was associated with the severity of systemic inflammation and the development of sepsis. Patients who received allogeneic blood transfusions had a higher incidence of nosocomial infections and sepsis, and the amount of blood transfusion as source of free heme correlated with the expression pattern of HP. CONCLUSIONS: These findings indicate that the heme degradation pathway is associated with increased susceptibility to septic complications after trauma, which is indicated by HP expression in particular.

Preconditioned suppression of prolyl-hydroxylases attenuates renal injury but increases mortality in septic murine models.

BACKGROUND: Septic conditions contribute to tissue hypoxia, potentially leading to multiple organ failure, including acute kidney injury. The regulation of cellular adaptation to low oxygen levels is regulated by hypoxia-inducible transcription factors (HIFs). While the role of HIFs in ischaemia/reperfusion is more studied, their function in sepsis-induced renal injury is not well characterized. In this study, we investigated whether pharmacological activation of HIFs by suppression of prolyl-hydroxylases (PHDs) protects against septic acute kidney injury. METHODS: Two models of sepsis-caecal ligation and punction and peritoneal contamination and infection-were induced on 12-week-old C57BL6/J mice. Pharmacological inhibition of PHDs, leading to HIF activation, was achieved by intraperitoneal application of 3,4-dihydroxybenzoate (3,4-DHB) before sepsis. A quantitative real-time reverse transcription polymerase chain reaction, immunohistology and enzyme-linked immunosorbent assays were utilized to detect gene expression, renal protein levels and renal functional parameters, respectively. Tissue morphology was analysed by periodic acid-Schiff reaction. Early kidney injury was estimated by kidney injury molecule-1 analyses. Apoptosis was detected in situ by terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling stain. The systemic effect of 3,4-DHB pretreatment in sepsis was analysed by 72-h survival studies. RESULTS: Pharmacological activation of HIFs before sepsis induction attenuated sepsis-related vacuolization and dilation of the proximal tubules, reduced tubular apoptosis and correlated to lower T-cell infiltration in renal tissue compared with the non-treated septic animals. PHD suppression elevated the basal renal HIF-1α expression and basal plasma concentrations of HIF targets erythropoietin and vascular endothelial growth factor. Whereas it preserved renal structure in both models, it improved renal function in a model-dependent manner. Moreover, inhibition of PHDs led to increased mortality in both models. Analysis of liver function showed increased organ destruction with massive glycogen loss and hepatocyte's apoptosis due to 3,4-DHB administration before sepsis induction. CONCLUSIONS: In summary, the pharmacological activation of HIFs by 3,4-DHB administration, although it showed renoprotective effects in sepsis-related kidney injury, induced more severe problems in other organs such as the liver during sepsis, leading to increased mortality.

CORM-EDE1: A Highly Water-Soluble and Nontoxic Manganese-Based photoCORM with a Biogenic Ligand Sphere.

[Mn(CO)5Br] reacts with cysteamine and 4-amino-thiophenyl with a ratio of 2:3 in refluxing tetrahydrofuran to the complexes of the type [{(OC)3Mn}2(µ-SCH2CH2NH3)3]Br2 (1, CORM-EDE1) and [{(OC)3Mn}2(µ-SC6H4-4-NH3)3]Br2 (2, CORM-EDE2). Compound 2 precipitates during refluxing of the tetrahydrofuran solution as a yellow solid whereas 1 forms a red oil that slowly solidifies. Recrystallization of 2 from water yields the HBr-free complex [{(OC)3Mn}2(µ-S-C6H4-4-NH2)2(µ-SC6H4-4-NH3)] (3). The n-propylthiolate ligand (which is isoelectronic to the bridging thiolate of 1) leads to the formation of the di- and tetranuclear complexes [(OC)4Mn(µ-S-nPr)2]2 and [(OC)3Mn(µ-S-nPr)]4. CORM-EDE1 possesses ideal properties to administer carbon monoxide to biological and medicinal tissues upon irradiation (photoCORM). Isolated crystalline CORM-EDE1 can be handled at ambient and aerobic conditions. This complex is nontoxic, highly soluble in water, and indefinitely stable therein in the absence of air and phosphate buffer. CORM-EDE1 is stable as frozen stock in aqueous solution without any limitations, and these stock solutions maintain their CO release properties. The reducing dithionite does not interact with CORM-EDE1, and therefore, the myoglobin assay represents a valuable tool to study the release kinetics of this photoCORM. After CO liberation, the formation of MnHPO4 in aqueous buffer solution can be verified.

Preserved Expression of mRNA Coding von Willebrand Factor-Cleaving Protease ADAMTS13 by Selenite and Activated Protein C.

In sepsis, the severity-dependent decrease of von Willebrand factor (VWF)-inactivating protease, a disintegrin and metalloproteinase with thrombospondin motifs 13 (ADAMTS13), results in platelet aggregation and consumption, leading to sepsis-associated thrombotic microangiopathy (TMA) and organ failure. Previous reports assessing its functional deficiency have pinpointed involvement of autoantibodies or mutations to propagate thrombotic thrombocytopenic purpura (TTP). However, mechanisms of acquired ADAMTS13 deficiency during host response remain unclear. To enhance understanding of ADAMTS13 deficiency in sepsis, we evaluated changes in expression of mRNA coding ADAMTS13 during septic conditions using primary cellular sources of the protease. We hypothesized that proinflammatory cytokines and constituents of serum from septic patients affect the transcriptional level of ADAMTS13 in vitro, and previously recommended therapeutic agents as adjunctive therapy for sepsis interact therewith. Cultured hepatic stellate cells (HSCs), endothelial cells (HMEC) and human precision-cut liver slices as an ex vivo model were stimulated with sepsis prototypic cytokines, bacterial endotoxin and pooled serum obtained from septic patients. Stimulation resulted in a significant decrease in ADAMTS13 mRNA between 10% and 80% of basal transcriptional rates. Costimulation of selenite or recombinant activated protein C (APC) with serum prevented ADAMTS13 decrease in HSCs and increased ADAMTS13 transcripts in HMEC. In archived clinical samples, the activity of ADAMTS13 in septic patients treated with APC (n = 5) increased with an accompanying decrease in VWF propeptide as surrogate for improved endothelial function. In conclusion, proinflammatory conditions of sepsis repress mRNA coding ADAMTS13 and the ameliorating effect by selenite and APC may support the concept for identification of beneficial mechanisms triggered by these drugs at a molecular level.

Imbalance of von Willebrand factor and its cleaving protease ADAMTS13 during systemic inflammation superimposed on advanced cirrhosis.

BACKGROUND & AIMS: Systemic inflammation in advanced cirrhosis represents a spectrum ranging from subclinical pathological bacterial translocation and immune activation to overt bacterial infection and sepsis. We hypothesized that systemic inflammation in cirrhosis is accompanied by a failure of ADAMTS13 to control the prothrombotic function of von Willebrand factor (VWF), which is increased in portal hypertension and hepatic fibrosis. METHODS: Patients with Child A cirrhosis (n = 25), Child B/C cirrhosis without clinical features of systemic inflammation (n = 31), and Child B/C cirrhosis with overt bacterial infections or systemic inflammatory response syndrome (n = 24) were analysed for ADAMTS13 and associated parameters and were followed to determine transplant-free survival. RESULTS: Plasma concentration and activity of ADAMTS13 were decreased in patients with systemic inflammation. Furthermore, ADAMTS13 inversely correlated with the extent of bacterial translocation and the severity of acute-phase reaction. As a function of reduced ADAMTS13 activity and increased VWF antigen, plasma from patients with superimposed inflammation strongly aggregated the platelet receptor glycoprotein Ib in presence of ristocetin. VWF:RCo correlated with higher concentrations of leucocytes and lipopolysaccharide-binding protein, organ dysfunction, augmented turnover of cross-linked intravascular fibrin, and the occurrence of acute kidney injury during follow-up. VWF:RCo of 390% or more predicted transplant-free survival in univariate analysis [HR = 8.24 (3.30-20.54)] and after adjustment for MELD [HR = 3.58 (1.30-9.88)]. However, adverse outcome was not associated with the accumulation of high-molecular weight VWF multimers. CONCLUSIONS: Systemic inflammation complicating advanced cirrhosis is accompanied by reduced activity of ADAMTS13 promoting a prothrombotic function of VWF, which can be employed to predict clinical outcome.

Improvement of prognostic performance in severely injured patients by integrated clinico-transcriptomics: a translational approach.

INTRODUCTION: Severe trauma triggers a systemic inflammatory response that contributes to secondary complications, such as nosocomial infections, sepsis or multi-organ failure. The present study was aimed to identify markers predicting complications and an adverse outcome of severely injured patients by an integrated clinico-transcriptomic approach. METHODS: In a prospective study, RNA samples from circulating leukocytes from severely injured patients (injury severity score ≥ 17 points; n = 104) admitted to a Level I Trauma Center were analyzed for dynamic changes in gene expression over a period of 21 days by quantitative RT-PCR. Transcriptomic candidates were selected based on whole genome screening of a representative discovery set (n = 10 patients) or known mechanisms of the immune response, including mediators of inflammation (IL-8, IL-10, TNF-α, MIF, C5, CD59, SPHK1), danger signaling (HMGB1, TLR2, CD14, IL-33, IL-1RL1), and components of the heme degradation pathway (HP, CD163, HMOX1, BLVRA, BLVRB). Clinical markers comprised standard physiological and laboratory parameters and scoring systems routinely determined in trauma patients. RESULTS: Leukocytes, thrombocytes and the expression of sphingosine kinase-1 (SPHK1), complement C5, and haptoglobin (HP) have been identified as markers with the best performance. Leukocytes showed a biphasic course with peaks on day 0 and day 11 after trauma, and patients with sepsis exhibited significantly higher leukocyte levels. Thrombocyte numbers showed a typical profile with initial thrombopenia and robust thrombocytosis in week 3 after trauma, ranging 2- to 3-fold above the upper normal value. 'Relative thrombocytopenia' was associated with multi-organ dysfunction, the development of sepsis, and mortality, the latter of which could be predicted within 3 days prior to the time point of death. SPHK1 expression at the day of admission indicated mortality with excellent performance. C5-expression on day 1 after trauma correlated with an increased risk for the development of nosocomial infections during the later course, while HP was found to be a marker for the development of sepsis. CONCLUSIONS: The combination of clinical and transcriptomic markers improves the prognostic performance and may represent a useful tool for individual risk stratification in trauma patients.

Hyperresponsiveness of mice deficient in plasma-secreted sphingomyelinase reveals its pivotal role in early phase of host response.

Plasma secretion of acid sphingomyelinase is a hallmark of cellular stress response resulting in the formation of membrane embedded ceramide-enriched lipid rafts and the reorganization of receptor complexes. Consistently, decompartmentalization of ceramide formation from inert sphingomyelin has been associated with signaling events and regulation of the cellular phenotype. Herein, we addressed the question of whether the secretion of acid sphingomyelinase is involved in host response during sepsis. We found an exaggerated clinical course in mice genetically deficient in acid sphingomyelinase characterized by an increased bacterial burden, an increased phagocytotic activity, and a more pronounced cytokine storm. Moreover, on a functional level, leukocyte-endothelial interaction was found diminished in sphingomyelinase-deficient animals corresponding to a distinct leukocytes' phenotype with respect to rolling and sticking as well as expression of cellular surface proteins. We conclude that hydrolysis of membrane-embedded sphingomyelin, triggered by circulating sphingomyelinase, plays a pivotal role in the first line of defense against invading microorganisms. This function might be essential during the early phase of infection leading to an adaptive response of remote cells and tissues.

Physical exercise induces specific adaptations resulting in reduced organ injury and mortality during severe polymicrobial sepsis.

OBJECTIVES: High physical activity levels are associated with wide-ranging health benefits, disease prevention, and longevity. In the present study, we examined the impact of regular physical exercise on the severity of organ injury and survival probability, as well as characteristics of the systemic immune and metabolic response during severe polymicrobial sepsis. DESIGN: Animal study. SETTING: University laboratory. SUBJECTS: Male C57BL/6N mice. INTERVENTIONS: Mice were trained for 6 weeks by treadmill and voluntary wheel running or housed normally. Polymicrobial sepsis in mice was induced by injection of fecal slurry. Subsequently, mice were randomized into the following groups: healthy controls, 6 hours postsepsis, and 24 hours postsepsis. MEASUREMENTS AND MAIN RESULTS: Blood and organ samples were collected and investigated by measuring clinical chemistry variables, cytokines, plasma metabolites, and bacterial clearance. Organ morphology and damage were characterized by histological staining. Physical exercise improved survival and the ability of bacterial clearance in blood and organs. The release of pro- and anti-inflammatory cytokines, including interleukin-6 and interleukin-10, was diminished in trained compared to untrained mice during sepsis. The sepsis-associated acute kidney tubular damage was less pronounced in pretrained animals. By metabolic profiling and regression analysis, we detected lysophosphatidylcholine 14:0, tryptophan, as well as pimelylcarnitine linked with levels of neutrophil gelatinase-associated lipocalin representing acute tubular injury (corrected R=0.910; p<0.001). We identified plasma lysophosphatidylcholine 16:0, lysophosphatidylcholine 17:0, and lysophosphatidylcholine 18:0 as significant metabolites discriminating between trained and untrained mice during sepsis. CONCLUSIONS: Regular physical exercise reduces sepsis-associated acute kidney injury and death. As a specific mechanism of exercise-induced adaptation, we identified various lysophosphatidylcholines that might function as surrogate for improved outcome in sepsis.

Early functional and transcriptomic changes in the myocardium predict outcome in a long-term rat model of sepsis.

Myocardial function is depressed in sepsis and is an important prognosticator in the human condition. Using echocardiography in a long-term fluid-resuscitated Wistar rat model of faecal peritonitis we investigated whether depressed myocardial function could be detected at an early stage of sepsis and, if so, whether the degree of depression could predict eventual outcome. At 6 h post-insult, a stroke volume <0.17 ml prognosticated 3-day mortality with positive and negative predictive values of 93 and 80%, respectively. Subsequent fluid loading studies demonstrated intrinsic myocardial depression with poor-prognosis animals tolerating less fluid than either good-prognosis or sham-operated animals. Cardiac gene expression analysis at 6 h detected 527 transcripts significantly up- or down-regulated by the septic process, including genes related to inflammatory and cell cycle pathways. Predicted mortality was associated with significant differences in transcripts of genes expressing proteins related to the TLR2/MyD88 (Toll-like receptor 2/myeloid differentiation factor 88) and JAK/STAT (Janus kinase/signal transducer and activator of transcription) inflammatory pathways, ß-adrenergic signalling and intracellular calcium cycling. Our findings highlight the presence of myocardial depression in early sepsis and its prognostic significance. Transcriptomic analysis in heart tissue identified changes in signalling pathways that correlated with clinical dysfunction. These pathways merit further study to both better understand and potentially modify the disease process.

Hepatic induction of cholesterol biosynthesis reflects a remote adaptive response to pneumococcal pneumonia.

Community-acquired pneumonia presents a spectrum of clinical phenotypes, from lobar pneumonia to septic shock, while mechanisms underlying progression are incompletely understood. In a transcriptomic and metabolomic study across tissues, we examined serotype-specific regulation of signaling and metabolic pathways in C57BL/6 mice intratracheally instilled with either serotype 19F Streptococcus pneumoniae (S19; causing lobar pneumonia), or serotype 2 S. pneumoniae (S2; causing septic pneumococcal disease,) or vehicle (Todd-Hewitt broth). Samples of lung, liver, and blood were collected at 6 and 24 h postinfection and subjected to microarray analysis and mass spectrometry. Results comprise a preferential induction of cholesterol biosynthesis in lobar pneumonia at low-infection doses (10(5) colony forming units/mouse) leading to increased plasma cholesterol (vehicle: 1.8±0.12 mM, S2: 2.3±0.10 mM, S19: 2.9±0.15 mM; P<0.05, comparing S19 to vehicle and S2). This induction was pneumolysin dependent, as a pneumolysin-deficient strain of serotype 19F failed to induce cholesterol biosynthesis (S19ΔPLY: 1.9±0.03 mM). Preincubation of pneumolysin with purified cholesterol or plasma from hypercholesterolemic mice prior to intratracheal instillation protected against lung barrier dysfunction and alveolar macrophage necrosis. Cholesterol may attenuate disease severity by neutralizing pneumolysin in the alveolar compartment and thus prevent septic disease progression.

Mechanisms and functional consequences of liver failure substantially differ between endotoxaemia and faecal peritonitis in rats.

BACKGROUND: Many of the concepts describing molecular mechanisms of sepsis-induced liver failure are derived from endotoxin models. However, the biological significance of such models is questionable as the complexity of clinical sepsis and associated organ failure is only partially replicated. AIMS: Comparison of cytokine response, leucocyte recruitment, oxidative stress and markers of hepatic organ dysfunction in rat models of endotoxaemia or peritoneal contamination and infection (PCI). METHODS: Endotoxemia and polymicrobial sepsis were induced in rats by intraperitoneal injection of lipopolysaccharide (LPS) or stool suspension, respectively. RESULTS: Both insults produced clinical and laboratory signs of multiple organ dysfunction, including hepatic excretory dysfunction. However, TNF alpha, oxidative stress responses and the degree of cell death were significantly higher in endotoxaemia compared to PCI (e.g. serum TNF levels (pg/ml) at 1.5 h post-insult: sham 5 ± 1.4, LPS 1 mg/kg bw 2176.92 ± 373.78, sepsis below detection limit; P P < 0.05). Cholestasis was significantly more pronounced in polymicrobial sepsis whereas serum bilirubin in endotoxaemic animals did not differ from sham-operated controls (plasma levels of bilirubin (µmol/L) at 15 h after the insult: sham 7.1 ± 0.6, LPS 30 mg/kg 9.1 ± 0.6, sepsis 15.2 ± 1.3). CONCLUSIONS: Polymicrobial sepsis produces profound hepatocellular dysfunction in the absence of traditional cytokine-mediated mechanisms of cellular injury. This questions the central role of cytokines and the ensuing oxidative stress as key molecular events in mediating liver dysfunction.