Sepsis-associated Cardiac Dysfunction Research Articles

Targeting of cold-inducible RNA-binding protein alleviates sepsis via alleviating inflammation, apoptosis and oxidative stress in heart.

A systemic inflammatory response is observed in patients undergoing shock and sepsis. This study aimed to explore the effects of cold-inducible RNA-binding protein (CIRP) on sepsis-associated cardiac dysfunction and the underlying mechanism. In vivo and in vitro lipopolysaccharide (LPS)-induced sepsis models were established in mice and neonatal rat cardiomyocytes (NRCMs), respectively. CRIP expressions were increased in the mouse heart and NRCMs treated with LPS. CIRP knockdown alleviated LPS-induced decreases of left ventricular ejection fraction and fractional shortening. CIRP downregulation attenuated the increases of inflammatory factors in the LPS-induced septic mouse heart, and NRCMs. The enhanced oxidative stress in the LPS-induced septic mouse heart and NRCMs was suppressed after CIRP knockdown. By contrast, CIRP overexpression yielded the opposite results. Our current study indicates that the knockdown of CIRP protects against sepsis-induced cardiac dysfunction through alleviating inflammation, apoptosis and oxidative stress of cardiomyocytes.

Honokiol alleviates sepsis-associated cardiac dysfunction via attenuating inflammation, apoptosis and oxidative stress.

Honokiol, a natural active compound extracted from Chinese herbal medicine, can ameliorate acute lung and kidney injury of sepsis. This study was to explore the effects of honokiol on sepsis-associated cardiac dysfunction and the underlying mechanism. Septic mice were induced by cecal ligation and puncture (CLP) or lipopolysaccharide (LPS), and septic HL-1 or AC16 cells were induced by LPS. Honokiol improved the survival and alleviated cardiac dysfunction in mice with CLP-induced sepsis. Honokiol inhibited the increased interleukin (IL) 1-β, IL-6 and tumour necrosis factor (TNF)-α in the serum and heart of CLP- and LSP-induced septic mice. Honokiol treatment reversed the increased levels of IL1-β, IL-6 and TNF-α in LPS-induced HL-1 cells. Honokiol treatment also decreased the elevated levels of IL1-β, IL-6 and TNF-α in LPS-induced AC16 cells. The increased cardiac apoptosis in CLP- and LPS-induced septic mice was alleviated by honokiol. The enhancement of oxidative stress in the heart of CLP- and LPS-induced septic mice was suppressed after honokiol administration. These results showed that honokiol could ameliorate sepsis-associated cardiac dysfunction via attenuating inflammation, apoptosis, and oxidative stress. Honokiol is a prospective drug for sepsis-associated heart damage in the future.

Hyperoside prevents sepsis-associated cardiac dysfunction through regulating cardiomyocyte viability and inflammation via inhibiting miR-21

BackgroundSepsis-associated cardiac dysfunction results in increased mortality. Hyperoside (Hyp) is a flavonoid, showing significant anti-inflammatory effects. However, its pharmacological effects on sepsis-induced cardiac dysfunction remain unknown. In this study, we attempted to explore whether Hyp could prevent cardiac dysfunction and its underlying mechanisms. MethodsWe established a mice mode of sepsis by cecal ligation and puncture (CLP) treatment, and constructed a cell model of myocardial injury by lipopolysaccharide (LPS) stimulation. The cardiac function indicators and the inflammatory cytokine levels were measured. Effect of Hyp on cardiomyocyte viability was evaluated using MTT assay. The expression and functional role of microRNA-21 (miR-21), a documented molecule that regulated by Hyp, was evaluated in the constructed models, and the potential targets of miR-21 were predicted. ResultsHyp alleviated the impaired cardiac function and stimulated inflammation caused by CLP in the in vivo sepsis model, and alleviated the LPS-induced decrease in cell viability and increase in inflammation of cardiomyocytes. Additionally, Hyp significantly inhibited the expression of miR-21 in LPS-induced cardiomyocytes, and the increased cell viability and decreased inflammation caused by Hyp in the in vitro model could be reversed by miR-21 overexpression. In animal model of sepsis, the protective influence of Hyp against sepsis-induced cardiac dysfunction was attenuated by miR-21 upregulation. ConclusionOur findings demonstrated that Hyp may serve as a promising natural drug for the treatment of sepsis-associated cardiac dysfunction, and its protective role may exerted through regulating cardiomyocyte viability and inflammation by suppressing miR-21.

Left Ventricular Impaired Relaxation and Interstitial Myocarditis Identified in Sepsis-Associated Cardiac Dysfunction: Use of a Rodent Model.

BackgroundSepsis is a serious clinical problem that results from the systemic response of the body to infection. Left ventricular (LV) diastolic dysfunction is increasingly appreciated as a contributor to morbidity and mortality in sepsis. Animal models may offer a method of studying diastolic dysfunction while controlling for many potential clinical confounders, such as sepsis duration, premorbid condition, and therapeutic interventions. This study sought to evaluate an endotoxemia (LPS) rodent model of sepsis, with regard to echocardiographic evidence, including tissue Doppler, of LV diastolic dysfunction and histopathology findings.Material/MethodsFourteen male Sprague-Dawley rats were randomly allocated (1: 1) to LPS or saline (control). Mean arterial blood pressure (MAP) was measured through cannulation of the carotid artery. After a 30-min stabilization, baseline assessment with echocardiography and blood collection was performed. Rats were administered 0.9% saline or LPS (10 mg/mL). Follow-up echocardiography and blood collection were performed after 2 h. Hearts were removed post-mortem and pathology studied using histology and immunohistochemistry.ResultsLPS was associated with hypotension (MAP 81.86±31.67 mmHg; 124.29±20.16; p=0.02) and LV impaired relaxation (myocardial early diastolic velocity [e′] 0.06±0.02 m/s; 0.09±0.02; P=0.008). Histopathology and immunohistochemistry demonstrated evidence of interstitial myocarditis (hydropic changes and inflammation).ConclusionsLPS was associated with both diastolic dysfunction (impaired relaxation) and interstitial myocarditis. These features may offer a link between the structural and functional changes that have previously been described separately in clinical sepsis. This may facilitate further studies focused upon the mechanism and potential benefit treatment of sepsis-associated cardiac dysfunction.

Protective role of matrine in sepsis-associated cardiac dysfunction through regulating the lncRNA PTENP1/miR-106b-5p axis

BackgroundMatrine has attractive cardioprotective effects in some diseases. This study aimed to evaluate the therapeutic potential of matrine against cardiac dysfunction induced by sepsis in vivo and in vitro, and further explore the related mechanisms. MethodsCecal ligation and puncture (CLP) was used to induce a sepsis mice model, and H9C2 cells treated with lipopolysaccharide (LPS) were used as a cardiac myoblast injury model. The evaluation of cardiac function of mice was performed by measuring cardiac function biomarker levels and hemodynamic indicators. An ELISA method was used to examine inflammatory cytokine levels. H9C2 cell viability was measured using MTT assay. The expression of non-coding RNAs that might be involved in matrine function was analyzed using real-time quantitative PCR. ResultsMatrine could significantly improve the cardiac function and attenuate the inflammatory response of the mice model, and could increase H9C2 viability and inhibit inflammation in the cell model. By matrine administration, the expression of PTENP1 was downregulated, but miR-106b-5p expression was upregulated both in vivo and in vitro. The cardioprotective effects of matrine in mice and cell models could be reversed by the overexpression of PTENP1 or the knockdown of miR-106b-5p, and the overexpression of miR-106b-5p could significantly abolish the effects of PTENP1 on cardiac function and inflammation. ConclusionAll the data revealed that matrine can alleviate sepsis-related cardiac dysfunction by enhancing cardiac myoblast viability and attenuating inflammatory responses through the PTENP1/miR-106b-5p axis.

Overexpression of miR-451a in sepsis and septic shock patients is involved in the regulation of sepsis-associated cardiac dysfunction and inflammation

The purpose of this study was to investigate the expression and clinical value of microRNA-451a (miR-451a) in septic patients and analyze its effect on sepsis-associated cardiac dysfunction and inflammation response. A rat model of sepsis was constructed by cecal ligation and puncture. The expression of miR-451a was measured by quantitative real-time PCR. Receiver operating characteristic (ROC) analysis was used to assess the diagnostic value of serum miR-451a. The cardiac function and inflammatory responses in septic rats were measured to explore the functional role of miR-451a. Serum expression of miR-451a was increased in septic patients compared with healthy controls, and had the ability to distinguish septic patients from healthy volunteers with a sensitivity and specificity of 87.8% and 81.5%, respectively. Elevated serum miR-451a was associated with sepsis severity, as evidenced by the increased expression of miR-451a in septic shock patients and its correlation with key clinical indicators. Significantly upregulated expression of miR-451a was found in septic patients with cardiac dysfunction, and the knockdown of miR-451a in sepsis rats improved cardiac function and inhibited inflammatory responses. All the data revealed that serum miR-451a serves as a candidate diagnostic biomarker of sepsis and a potential parameter to indicate disease severity. The reduction of miR-451a may mitigate sepsis-induced cardiac dysfunction and inflammatory responses.

Bruton's Tyrosine Kinase Inhibition Attenuates the Cardiac Dysfunction Caused by Cecal Ligation and Puncture in Mice.

Sepsis is one of the most prevalent diseases in the world. The development of cardiac dysfunction in sepsis results in an increase of mortality. It is known that Bruton's tyrosine kinase (BTK) plays a role in toll-like receptor signaling and NLRP3 inflammasome activation, two key components in the pathophysiology of sepsis and sepsis-associated cardiac dysfunction. In this study we investigated whether pharmacological inhibition of BTK (ibrutinib 30 mg/kg and acalabrutinib 3 mg/kg) attenuates sepsis associated cardiac dysfunction in mice. 10-week old male C57BL/6 mice underwent CLP or sham surgery. One hour after surgery mice received either vehicle (5% DMSO + 30% cyclodextrin i.v.), ibrutinib (30 mg/kg i.v.), or acalabrutinib (3 mg/kg i.v.). Mice also received antibiotics and an analgesic at 6 and 18 h. After 24 h, cardiac function was assessed by echocardiography in vivo. Cardiac tissue underwent western blot analysis to determine the activation of BTK, NLRP3 inflammasome and NF-κB pathway. Serum analysis of 33 cytokines was conducted by a multiplex assay. When compared to sham-operated animals, mice subjected to CLP demonstrated a significant reduction in ejection fraction (EF), fractional shortening (FS), and fractional area change (FAC). The cardiac tissue from CLP mice showed significant increases of BTK, NF-κB, and NLRP3 inflammasome activation. CLP animals resulted in a significant increase of serum cytokines and chemokines (TNF-α, IL-6, IFN-γ, KC, eotaxin-1, eotaxin-2, IL-10, IL-4, CXCL10, and CXCL11). Delayed administration of ibrutinib and acalabrutinib attenuated the decline of EF, FS, and FAC caused by CLP and also reduced the activation of BTK, NF-κB, and NLRP3 inflammasome. Both ibrutinib and acalabrutinib significantly suppressed the release of cytokines and chemokines. Our study revealed that delayed intravenous administration of ibrutinib or acalabrutinib attenuated the cardiac dysfunction associated with sepsis by inhibiting BTK, reducing NF-κB activation and the activation of the inflammasome. Cytokines associated with sepsis were significantly reduced by both BTK inhibitors. Acalabrutinib is found to be more potent than ibrutinib and could potentially prove to be a novel therapeutic in sepsis. Thus, the FDA-approved BTK inhibitors ibrutinib and acalabrutinib may be repurposed for the use in sepsis.

Linagliptin Attenuates the Cardiac Dysfunction Associated With Experimental Sepsis in Mice With Pre-existing Type 2 Diabetes by Inhibiting NF-κB.

The mortality rate of patients who develop sepsis-related cardiac dysfunction is high. Many disease conditions (e.g., diabetes) increase the susceptibility to infections and subsequently sepsis. Activation of the NF-κB pathway plays a crucial role in the pathophysiology of sepsis-associated cardiac dysfunction and diabetic cardiomyopathy. The effect of diabetes on outcomes in patients with sepsis is still highly controversial. We here hypothesized that type 2 diabetes (T2DM) augments the cardiac (organ) dysfunction associated with sepsis, and that inhibition of the NF-κB pathway with linagliptin attenuates the cardiac (organ) dysfunction in mice with T2DM/sepsis. To investigate this, 10-week old male C57BL/6 mice were randomized to receive normal chow or high fat diet (HFD), 60% of calories derived from fat). After 12 weeks, mice were subjected to sham surgery or cecal ligation and puncture (CLP) for 24 h. At 1 hour after surgery, mice were treated with linagliptin (10 mg/kg, i.v.), IKK-16 (1 mg/kg, i.v.), or vehicle (2% DMSO, 3 ml/kg, i.v.). Mice also received analgesia, fluids and antibiotics at 6 and 18 h after surgery. Mice that received HFD showed a significant increase in body weight, impairment in glucose tolerance, reduction in ejection fraction (%EF), and increase in alanine aminotransferase (ALT). Mice on HFD subjected to CLP showed further reduction in %EF, increase in ALT, developed acute kidney dysfunction and lung injury. They also showed significant increase in NF-κB pathway, iNOS expression, and serum inflammatory cytokines compared to sham surgery group. Treatment of HFD-CLP mice with linagliptin or IKK-16 resulted in significant reductions in (i) cardiac, liver, kidney, and lung injury associated with CLP-sepsis, (ii) NF-κB activation and iNOS expression in the heart, and (iii) serum inflammatory cytokine levels compared to HFD-CLP mice treated with vehicle. Our data show that pre-existing type 2 diabetes phenotype worsens the organ dysfunction/injury associated with CLP-sepsis in mice. Most notably, inhibition of NF-κB reduces the organ dysfunction/injury associated with sepsis in mice with pre-existing T2DM.

Septic cardiomyopathy: pathophysiology and potential new therapeutic approaches

Sepsis is the leading cause of death in critically ill patients, and its incidence continues to rise. Sepsis was defined as a systemic inflammatory response syndrome with an identifiable focus of infection, but therapeutic strategies aimed at eliminating the inflammatory response have only modest clinical benefit. The development of a failure of one or more organs poses a major threat to the survival of patients with sepsis, and mortality in sepsis is most often attributed to multiple organ dysfunction. Accordingly, sepsis has been recently redefined as life-threatening organ dysfunction due to a dysregulated host response to infection. Cardiac dysfunction is a well-recognized important component of septic multiple organ failure and can compromise the balance between oxygen supply and demand, ultimately leading to the development of multiple organ failure. The existence of cardiac dysfunction in sepsis is associated with much higher mortality when compared with septic patients without heart problems. Dobutamine, a β1-selective adrenoceptor agonist, has been used in septic shock for many years as an only inotrope, but limited clinical outcome measures have been provided as to advisability of the usefulness of dobutamine in septic shock management. Here we provide an overview on the possible mechanisms underlying intrinsic myocardial depression during sepsis and discuss the perspective of several inotropes for sepsis-associated cardiac dysfunction.

Cardiovascular disease and resuscitated septic shock lead to the downregulation of the H2S-producing enzyme cystathionine-\u03b3-lyase in the porcine coronary artery

BackgroundDownregulation of the hydrogen sulfide (H2S)-producing enzymes cystathionine-γ-lyase (CSE), cystathionine-β-synthase (CBS), and/or 3-mercaptopyruvate sulfurtransferase (3-MST) is associated with chronic cardiovascular pathologies. Nevertheless, equivocal data are available on both the expression and function of these enzymes in coronary arteries (CA). We recently reported that atherosclerotic pigs subjected to sepsis developed impaired cardiac function, which coincided with decreased myocardial CSE expression and increased nitrotyrosine formation. To define the endogenous source(s) of H2S in the CA, we studied the expression of CBS, CSE, or 3-MST in the CA of pigs subjected to septic shock with/without pre-existing cardiovascular co-morbidity.MethodsAnesthetized and instrumented FBM "familial hypercholesterolemia Bretoncelles Meishan" pigs with high-fat diet-induced hypercholesterolemia and atherosclerosis were subjected to polymicrobial septic shock, or sham procedure, and subsequent intensive care therapy for 24 h. Young German domestic pigs were used as naïve controls. CSE, CBS, 3-MST, HO-1, eNOS, and nitrotyrosine expression was quantified by immunohistochemistry of formalin-fixed paraffin sections.ResultsFBM pigs, in the absence of septic shock, showed decreased CSE expression in the media. This decrease became more pronounced after sepsis. The expression pattern of HO-1 resembled the pattern of CSE expression. CBS protein was not detected in the media of any of the CA examined but was localized to the adventitia and only in the atheromatous plaques containing foam cells of the CA, in regions that also displayed abundant nitrotyrosine formation. The CBS expression in the adventitia was not associated with nitrotyrosine formation. 3-MST expression was not found in any of the CA samples.ConclusionsWe hypothesize that (i) the reduced CSE expression in FBM pigs may contribute to their cardiovascular disease phenotype and moreover (ii) the further decrease in CA CSE expression in sepsis may contribute to the sepsis-associated cardiac dysfunction.

MiR-21-3p controls sepsis-associated cardiac dysfunction via regulating SORBS2

Cardiac dysfunction with sepsis is a major cause of death in intensive care units. Several lines of evidence have revealed the potential of microRNAs (miRNAs, miRs) as biomarkers for detecting sepsis, though direct evidence of their functional roles in septic cardiac dysfunction is still lacking. In this study, C57BL/6 mice were exposed to lipopolysaccharide (LPS) to induce sepsis-associated cardiac dysfunction, as evidenced by reduced fractional shortening (FS) and ejection fraction (EF) and detrimental changes in cardiac contractility, inflammation, and energy metabolism. Microarray analysis and qRT-PCRs revealed that miR-21-3p was significantly induced in heart samples challenged with LPS. Impressively, pharmacological inhibition of miR-21-3p using antagomiR was able to preserve FS and EF and prevent mitochondria ultrastructural damage and autophagy in LPS-treated mice, while forced expression of miR-21-3p using agomiR aggravated that. Besides that, miR-21-3p antagomiR improved the survival of mice treated with LPS. Meanwhile, our data showed that SH3 domain-containing protein 2 (SORBS2) was inversely correlated with miR-21-3p expression level in mice hearts, and was repressed in hearts challenged with LPS, suggesting SORBS2 as a target gene of miR-21-3p. Additionally, plasma miR-21-3p was markedly elevated in septic patients with cardiac dysfunction as compared to septic patients without cardiac dysfunction. The ROC curve showed that plasma miR-21-3p could be a specific predictor of septic patients developing cardiac dysfunction with an area under the curve of 0.939. Collectively, the present study provides strong evidence that miR-21-3p controls sepsis-associated cardiac dysfunction via regulating SORBS2. Inhibition of miR-21-3p might be a protective strategy to treat sepsis-induced cardiac dysfunction.

Abstract 20517: MicroRNA-155: A New Player in Sepsis-Induced Cardiac Dysfunction

Introduction: Cardiac dysfunction is an important outcome predictor in patients with septic shock and lacks effective and targeted therapies. MicroRNA-155 (miR-155) is a multifunctional microRNA with important target genes in inflammation, immunity and cardiovascular system. Hypothesis: The present study assessed the role of miR-155 in experimental and human sepsis-induced cardiac dysfunction. Methods: Experimental sepsis was induced using endotoxin injection (LPS) or cecal ligation and puncture (CLP) models. Genetic loss (miR-155 KO) and pharmacological inhibition (LNA-anti-miR; AM) of miR-155 were evaluated. Cardiac function was assessed through echocardiography; mortality analysis was also performed. Quantification of myocardial miR-155, pro-inflammatory cytokine mRNA profile, miR-155 targets, apoptosis, cellular adhesion proteins and activation of pro-inflammatory intracellular signaling pathways were assessed. In another experiment, myocardial miR-155 expression, cellular localization and associations between its levels and other clinical variables were evaluated using post-mortem samples from septic shock (SS) patients. Results: Myocardial miR-155 levels were increased in experimental and human septic myocardium. This was associated with a decreased ejection fraction (EF) and cardiac output (CO) and increased diastolic LV diameter (LVDd) in septic WT animals. Knockout and anti-miR-155 treated animals and presented with preserved EF, CO and LVDd, compared with WT; this was accompanied by ~50% mortality reduction. Experimental sepsis induced downregulation of SOCS1 and CD47, myocardial cytokine upregulation, increased apoptosis and adhesion protein expression as well as pro-inflammatory kinase and transcription factor activation, all of which were attenuated in KO and AM. In human myocardium, miR-155 expression was positively associated with 24h antemortem period plasma troponin I levels. Conclusions: miR-155 is upregulated in experimental and human septic myocardium. Loss or inhibition of miR-155 attenuated sepsis-induced cardiac dysfunction, blunted proinflammatory activation and reduced mortality. This suggests miR-155 as a potential target in human sepsis-associated cardiac dysfunction.

Cardiac fibroblasts: contributory role in septic cardiac dysfunction

BackgroundCardiac dysfunction is a frequent and severe complication of septic shock and contributes to the high mortality of sepsis. Although several mechanisms have been suspected to be responsible for sepsis-associated cardiac dysfunction, the precise cause(s) remains unclear to date. Materials and methodsWe tested the hypothesis that cardiac fibroblasts may play a critical role as a disease modifier involved in sepsis-associated cardiac dysfunction. Human cardiac fibroblasts (HCFs) cultured in vitro were exposed to lipopolysaccharide (LPS). Changes in cardiac morphology and function were assessed in mice with cecal ligation and puncture-induced sepsis. ResultsIn LPS-stimulated HCFs, messenger RNA and protein levels of proinflammatory molecules, including tumor necrosis factor-α, interleukin-1β, interleukin-6, and monocyte chemoattractant protein-1, were strikingly upregulated. LPS also increased expression and activity of matrix metalloproteinase (MMP)-9, but not MMP-2. LPS-induced expression of α-smooth muscle actin, a classical marker for myoblast differentiation, which was abrogated when MMP-9 small interfering RNA was transfected into HCFs. High gene expression levels of proinflammatory cytokines and MMP-9 were observed in the heart tissues of cecal ligation and puncture-induced septic mice. Histology sections of the hearts from septic mice showed perivascular and interstitial cardiac fibrosis, and echocardiography demonstrated that septic mice had profound cardiac dysfunction. The broad-spectrum MMP inhibitor ONO-4817 significantly alleviated these histologic and functional changes during the acute phase. ConclusionsWe suggest that cardiac fibroblasts are of pathogenetic importance in inflammation and fibrosis in the heart during sepsis, leading to cardiac dysfunction that would affect the outcome of sepsis syndrome.

The impact of cardiac dysfunction on acute respiratory distress syndrome and mortality in mechanically ventilated patients with severe sepsis and septic shock: An observational study

PurposeAcute respiratory distress syndrome (ARDS) is associated with significant mortality and morbidity in survivors. Treatment is only supportive, therefore elucidating modifiable factors that could prevent ARDS could have a profound impact on outcome. The impact that sepsis-associated cardiac dysfunction has on ARDS is not known. Materials and MethodsIn this retrospective observational cohort study of mechanically ventilated patients with severe sepsis and septic shock, 122 patients were assessed for the impact of sepsis-associated cardiac dysfunction on incidence of ARDS (primary outcome) and mortality. ResultsSepsis-associated cardiac dysfunction occurred in 44 patients (36.1%). There was no association of sepsis-associated cardiac dysfunction with ARDS incidence (p= 0.59) or mortality, and no association with outcomes in patients that did progress to ARDS after admission. Multivariable logistic regression demonstrated that higher BMI was associated with progression to ARDS (adjusted OR 11.84, 95% CI 1.24 to 113.0, p= 0.02). ConclusionsCardiac dysfunction in mechanically ventilated patients with sepsis did not impact ARDS incidence, clinical outcome in ARDS patients, or mortality. This contrasts against previous investigations demonstrating an influence of nonpulmonary organ dysfunction on outcome in ARDS. Given the frequency of ARDS as a sequela of sepsis, the impact of cardiac dysfunction on outcome should be further studied.

233 ERYTHROPOIETIN ATTENUATES CARDIAC DYSFUNCTION IN EXPERIMENTAL SEPSIS VIA ACTIVATION OF THE Β-COMMON RECEPTOR

Rationale There is limited evidence that the tissue-protective effects of erythropoietin are mediated by a heterocomplex of the erythropoietin-receptor and the β-common receptor (‘tissue-protective receptor9), which is pharmacologically distinct from the ‘classical9 erythropoietin-receptor homodimer responsible for erythropoiesis. However, the role of the β-common receptor and/or erythropoietin in sepsis-induced cardiac dysfunction (a well known, serious complication of sepsis) is unknown. Here we report for the first time that β-common receptor is essential for the improvements in the impaired systolic contractility afforded by erythropoietin in experimental sepsis. Methods and Results Cardiac function was assessed in vivo (echocardiography) and ex vivo (Langendorff-perfused heart) in wild-type and β-common receptor knock-out mice, that were subjected to lipopolysaccharide (9 mg/kg; young mice) for 16–18 h or cecal ligation and puncture (aged mice) for 24 h. Mice received erythropoietin (1000 IU/kg) 1 h after lipopolysaccahride or cecal ligation and puncture. Erythropoietin reduced the impaired systolic contractility ( in vivo and ex vivo ) caused by endotoxemia/sepsis in young as well as old wild-type mice in a β-common receptor-dependent fashion. Activation by erythropoietin of the β-common receptor also resulted in the activation of well-known survival pathways (Akt and endothelial nitric oxide synthase) and inhibition of pro-inflammatory pathways (glycogen synthase kinase-3β, nuclear factor-κB and interleukin-1β). All the above pleitropic effects of erythropoietin were lost inβ-common receptor knock-out mice. Conclusions Erythropoietin attenuates the impaired systolic contractility associated with sepsis by activation of the β-common receptor, which in turn, results in activation of survival pathways and inhibition of inflammation. Thus, targeting the 9tissue protective receptor9 with specific agonists devoid of the erythropoietic effects of erythropoietin may represent a novel approach for the treatment of sepsis-associated cardiac dysfunction. *AIK and SMC contributed equally to this study.

Beneficial Effects of Levosimendan in Infants With Sepsis-Associated Cardiac Dysfunction

Although international guidelines for hemodynamic support of pediatric septic shock suggest considering the potent calcium sensitizer inodilator levosimendan as a second-line inotropic agent to treat cold hypodynamic shock resistant to catecholamines, clinical experience in septic infants is lacking. We report the beneficial effects of levosimendan infusion in 2 infants (39 and 64 days old) with low cardiac output septic shock (ejection fraction, 29% and 33%; fractional shortening, 10% and 16% respectively) refractory to volume replacement and to the catecholamines dopamine and dobutamine. Levosimendan infusion (0.15 µg/kg/min) promptly increased both patients' myocardial contractility and improved tissue perfusion, thereby reducing lactate levels and increasing urine output. The only adverse effect was moderate hypotension in one infant, who reversed by adding norepinephrine to levosimendan. This clinical experience confirms the potential beneficial effects of levosimendan infusion to restore hemodynamics in infants with low cardiac output septic shock resistant to catecholamines.

Salvaging the septic heart through targeting the interleukin-6/p38 mitogen-activated protein kinase signaling network*

Depression of myocardial function during severe sepsis, which currently accounts for approx. 200,000 deaths/year in the United States (1), is characterized by a decrease in contractility and a poor response to fluid therapy (2). Since the md-1980s it has been recognized that the decreased cardiac function, which undoubtedly contributes to the overall pathophysiology of the septic state, does not arise from factors that are intrinsic to the myocardium, but instead results from the presence of circulating myocardial depressant factors (3, 4). Since much of the massive inflammation and multi-organ dysfunction in sepsis result from the secretion of various cytokines, it was long suspected that these proteins were also responsible, at least in part, for the observed myocardial dysfunction, although their identification, and the molecular basis for their effects on myocyte function were poorly understood. Pathan and colleagues recently showed that the cytokine IL-6 is a major factor responsible for modulating myocardial depression in children with meningococcal septicemia (5). In this issue of Critical Care Medicine, Pathan et al., use an in-vitro cardiac myocyte cell culture system to investigate the signaling pathways (and potential drug targets) that modulate and/or are modulated by IL-6 (new article). They report that the stress activated p38 mitogen activated protein kinase (p38 MAPK) pathway controls IL-6-induced myocardial depression and that this can be reversed using a specific chemical inhibitor of this kinase (SB203580). The authors go on to look at gene expression changes in a small cohort of pediatric patients with meningococcal sepsis and find a number of differences between septic patients and healthy controls. Amongst the genes that show differential expression are a number of upstream and downstream regulators of p38 MAPK signaling indicating that this pathway is deregulated in this cohort of patients. However, the mechanism by which such a large group of genes involved in p38 MAPK signaling could be deregulated remains to be determined. In addition to the work of Pathan et al., are a number of observations that link p38 MAPK and one of it’s downstream kinases, MK2 (MAPKAPK2) to the control of IL-6 biosynthesis (6). For example, during LPS-induced inflammation p38 MAPK activates MK2, which in turn controls IL-6 production by stabilizing the IL-6 messenger RNA (7). Therefore the p38 MAPK/MK2 pathway appears to lie both upstream and downstream of IL-6 in inflammation and sepsis. These observations are of obvious clinical relevance since p38 MAPK and its downstream effectors are under intense scrutiny as viable drug targets for the treatment of a number of diseases from colitis to cancer (6, 8, 9). The current study further highlights the potential utility of p38 MAPK inhibitors in the specific management of sepsis-associated cardiac dysfunction. p38 MAPK, however, activates a variety of downstream effectors that affect a wide variety of other cellular responses during sepsis in addition to cytokine secretion and myocardial contractility, including effects on apoptosis and neutrophil function (10, 11). In this regard, drugs that target the specific components of the p38 MAPK pathway that are central to its IL-6 effects, such as MK2 or its substrates, may also show promise in the clinical treatment of sepsis by preventing IL-6 production thus inhibiting myocardial depression, while having fewer effects on other p38 MAPK-dependent responses. In this regard emerging new drugs targeting IL-6 itself, such as IL-6 blocking monoclonal antibodies (12), may also show utility in this setting. Importantly the utility of these drugs will not be limited to myocardial dysfunction in sepsis since the p38 MAPK/MK2/IL6 pathway appears to be a major determinant of response to chemotherapy in certain cancers (13, 14). For example, p38 MAPK-dependent IL-6 secretion from thymic endothelial cells mediates drug resistance and relapse in a mouse model of Burkitt’s lymphoma (15). Therefore new drugs targeting the p38 MAPK-MK2-IL-6 signaling pathway may have a broader utility in cancer treatment as well as for the treatment of conditions where chronic inflammation is a major driving force of the diseased state, such as sepsis. If such drugs prove useful, it will be yet another example of how agents that were developed to limit the pathology of inflammation, such as cyclooxygenase inhibitors, can have a double life in clinical oncology. That indeed would be quite a lot of heart to salvage.

The effect of diazepam on myocardial function and coronary vascular tone after endotoxemia in the isolated rat heart model

Tumor necrosis factor alpha (TNF-α) has been implicated in the pathogenesis of cardiovascular disease and sepsis-associated cardiac dysfunction. Although initially described solely as a lipopolysaccharide (LPS)-induced macrophage product, evidence exists that cardiac myocytes themselves produce substantial amounts of TNF-α in response to ischemia as well as LPS. The use of phosphodiesterase inhibitors has been shown to decrease LPS-induced TNF-α elaboration. The aim of the present study was to determine the effect of diazepam (Type IV phosphodiesterase inhibitor) on (1) myocardial function and (2) coronary vascular flow after LPS-induced endotoxic shock in an isolated rat heart model. Endotoxemia was induced by intraperitoneal LPS administration in adult male Wistar rats. Hearts were isolated after 6 h and perfused in a working mode with oxygenated Krebs-Henseleit buffer at 37°C. Diazepam was mixed with Krebs-Henseleit buffer and administered (3.0 μg/ml) for 20 min. LPS-treated hearts showed depressed cardiac function and reduced coronary flow. Myocardial functional parameters (LVDP, +dP/dt, -dP/dt, RPP) and coronary flow (ml/min) were significantly (p < 0.01) improved by diazepam administration. These findings suggest that diazepam can salvage myocardial function and undo coronary vascular constriction in the endotoxemic rat heart. These findings are clinically relevant to the treatment of cardiovascular depression caused by endotoxic shock.

Protein phosphatase 2A contributes to the cardiac dysfunction induced by endotoxemia

AimsSepsis-associated cardiac dysfunction represents an intrinsic impairment of cardiomyocyte function due in part to a decrease in myofilament Ca2+ sensitivity associated with a sustained increase in cardiac troponin I (cTnI) phosphorylation at Ser23/24. Dephosphorylation of cTnI is under regulatory control. Thus, muscarinic and adenosine A1-receptor agonists antagonize β-adrenergic stimulation via activation of protein phosphatase 2A (PP2A). The aim of this study was to determine whether modulation of PP2A and thus cTnI phosphorylation could improve sepsis-induced contractile dysfunction.Methods and resultsCardiomyocytes were isolated from control or septic mice 16–18 h after an injection of vehicle or lipopolysaccharide (LPS; 9 mg/kg ip) respectively. Protein expression and phosphatase activity were determined in homogenates of control and septic hearts. Our data showed that LPS significantly increased cTnI phosphorylation at Ser23/24 in cardiomyocytes and reduced contraction amplitude without affecting Ca2+-transients. Treatment of cardiomyocytes with the A1 agonist cyclopentyladenosine (CPA) or the protein kinase A inhibitor H89 significantly attenuated the LPS-induced contractile dysfunction without effect on Ca2+-transients. Co-treatment with CPA and H89 completely reversed the contractile dysfunction. Increased cTnI phosphorylation in septic hearts was associated with a significant reduction in the protein expression of both the catalytic and regulatory subunits (B56α) of PP2A and a decrease in PP2A activity. CPA treatment of septic hearts increased PP2A activity. An increase in the protein expression of demethylated PP2A and a decrease in the PP2A-methyltransferase (PPMT; the methyltransferase that catalyses this reaction) were also observed.ConclusionThese data support the hypothesis that sustained cTnI phosphorylation underlies the contractile dysfunction seen in sepsis.

THE RESPONSE OF NEONATAL RAT VENTRICULAR MYOCYTES TO LIPOPOLYSACCHARIDE-INDUCED STRESS

Sepsis induced by exposure to lipopolysaccharide (LPS) can be life-threatening and lead to multiple-organ dysfunction. Sepsis-associated cardiac dysfunction is a primary cause of mortality. The response of isolated cardiac myocytes to LPS exposure is poorly understood. Cultured neonatal rat ventricular cardiomyocytes were used to evaluate the response to LPS exposure. Other authors have reported that LPS exposure at doses sufficient to induce tumor necrosis factor alpha (TNF-alpha) production and apoptosis in adult cardiomyocytes do not induce apoptosis in neonatal cardiomyocytes. We therefore hypothesized that neonatal cardiomyocytes have innate protective mechanisms that protect from septic damage. Cultured neonatal rat ventricular cardiomyocytes were stimulated by exposure to LPS for varying lengths of time. NFkappaB signaling pathways, TNF-alpha production, and Akt activation were monitored. We also assessed the induction of apoptosis in these cells by monitoring caspase-3 activity. LPS rapidly stimulates nuclear translocation of NFkappaB and Akt activation. TNF-alpha production is also stimulated. However, high doses of LPS are unable to induce apoptosis in these cells, and protection is not a function of Akt activation. LPS treatment also stimulated the levels of cyclooxygenase-2 and the production of downstream metabolites, specifically PGE2 and 15deoxyDelta12-14PGJ2 (15dPGJ2). Specific inhibition of cyclooxygenase-2 activity induced apoptosis in the presence of LPS, whereas direct exposure to 15dPGJ2 at pharmacological levels induced apoptosis. Neonatal rat ventricular cardiomyocytes have innate protective mechanisms that prevent apoptotic cell death after LPS exposure. Metabolic products of arachidonic acid metabolized by the cyclooxygenase pathway can be potentially apoptotic or antiapoptotic. The balance of these products within these cells may define the cellular response to LPS exposure.