Animal Models Of Septic Shock Research Articles

BPOZ-2 is a negative regulator of the NLPR3 inflammasome contributing to SARS-CoV-2-induced hyperinflammation.

Inflammation play important roles in the initiation and progression of acute lung injury (ALI), acute respiratory distress syndrome (ARDS), septic shock, clotting dysfunction, or even death associated with SARS-CoV-2 infection. However, the pathogenic mechanisms underlying SARS-CoV-2-induced hyperinflammation are still largely unknown. The animal model of septic shock and ALI was established after LPS intraperitoneal injection or intratracheal instillation. Bone marrow-derived macrophages (BMDMs) from WT and BPOZ-2 KO mouse strains were harvested from the femurs and tibias of mice. Immunohistology staining, ELISA assay, coimmunoprecipitation, and immunoblot analysis were used to detect the histopathological changes of lung tissues and the expression of inflammatory factors and protein interaction. We show a distinct mechanism by which the SARS-CoV-2 N (SARS-2-N) protein targets Bood POZ-containing gene type 2 (BPOZ-2), a scaffold protein for the E3 ubiquitin ligase Cullin 3 that we identified as a negative regulator of inflammatory responses, to promote NLRP3 inflammasome activation. We first demonstrated that BPOZ-2 knockout (BPOZ-2 KO) mice were more susceptible to lipopolysaccharide (LPS)-induced septic shock and ALI and showed increased serum IL-1β levels. In addition, BMDMs isolated from BPOZ-2 KO mice showed increased IL-1β production in response to NLRP3 stimuli. Mechanistically, BPOZ-2 interacted with NLRP3 and mediated its degradation by recruiting Cullin 3. In particular, the expression of BPOZ-2 was significantly reduced in lung tissues from mice infected with SARS-CoV-2 and in cells overexpressing SARS-2-N. Importantly, proinflammatory responses triggered by the SARS-2-N were significantly blocked by BPOZ-2 reintroduction. Thus, we concluded that BPOZ-2 is a negative regulator of the NLPR3 inflammasome that likely contributes to SARS-CoV-2-induced hyperinflammation.

CARDIOMYOCYTE REPROGRAMMING IN ANIMAL MODELS OF SEPTIC SHOCK.

Cardiomyocyte reprogramming plays a pivotal role in sepsis-induced cardiomyopathy through the induction or overexpression of several factors and enzymes, ultimately leading to the characteristic decrease in cardiac contractility. The initial trigger is the binding of LPS to TLR-2, -3, -4, and -9 and of proinflammatory cytokines, such as TNF, IL-1, and IL-6, to their respective receptors. This induces the nuclear translocation of nuclear factors, such as NF-κB, via activation of MyD88, TRIF, IRAK, and MAPKs. Among the latter, ROS- and estrogen-dependent p38 and ERK 1/2 are proinflammatory, whereas JNK may play antagonistic, anti-inflammatory roles. Nuclear factors induce the synthesis of cytokines, which can amplify the inflammatory signal in a paracrine fashion, and of several effector enzymes, such as NOS-2, NOX-1, and others, which are ultimately responsible for the degradation of cardiomyocyte contractility. In parallel, the downregulation of enzymes involved in oxidative phosphorylation causes metabolic reprogramming, followed by a decrease in ATP production and the release of fragmented mitochondrial DNA, which may augment the process in a positive feedback loop. Other mediators, such as NO, ROS, the enzymes PI3K and Akt, and adrenergic stimulation may play regulatory roles, but not all signaling pathways that mediate cardiac dysfunction of sepsis do that by regulating reprogramming. Transcription may be globally modulated by miRs, which exert protective or amplifying effects. For all these mechanisms, differentiating between modulation of cardiomyocyte reprogramming versus systemic inflammation has been an ongoing but worthwhile experimental challenge.

Myocardial effects of angiotensin II compared to norepinephrine in an animal model of septic shock

BackgroundAngiotensin II is one of the vasopressors available for use in septic shock. However, its effects on the septic myocardium remain unclear. The aim of the study was to compare the effects of angiotensin II and norepinephrine on cardiac function and myocardial oxygen consumption, inflammation and injury in experimental septic shock.MethodsThis randomized, open-label, controlled study was performed in 20 anesthetized and mechanically ventilated pigs. Septic shock was induced by fecal peritonitis in 16 animals, and four pigs served as shams. Resuscitation with fluids, antimicrobial therapy and abdominal drainage was initiated one hour after the onset of septic shock. Septic pigs were randomly allocated to receive one of the two drugs to maintain mean arterial pressure between 65 and 75 mmHg for 8 h.ResultsThere were no differences in MAP, cardiac output, heart rate, fluid balance or tissue perfusion indices in the two treatment groups but myocardial oxygen consumption was greater in the norepinephrine-treated animals. Myocardial mRNA expression of interleukin-6, interleukin-6 receptor, interleukin-1 alpha, and interleukin-1 beta was higher in the norepinephrine than in the angiotensin II group.ConclusionsIn septic shock, angiotensin II administration is associated with a similar level of cardiovascular resuscitation and less myocardial oxygen consumption, and inflammation compared to norepinephrine.

A rat model of septic shock using live Escherichia coli and response to Noradrenaline

Objectives: Popular animal models of septic shock involve injections of endotoxin (bacterial lipopolysaccharide). Other methods that induce sepsis are often time-consuming and require long-term monitoring facilities. Further, individual models using different bacterial strains can deepen our understanding of sepsis pathophysiology. Hence, our objective was to develop an acute and functional Wistar rat model of septic shock using live strains of Escherichia coli and then administer Noradrenaline, a known sympathomimetic drug, to study if the response is along expected lines. Materials and Methods: After random allocation to one of three groups (Group 1 – E. coli alone, n=7; Group 2 – E. coli followed by Noradrenaline, n = 7 and Group 3 – control (n = 4), which received saline injections), Wistar rats were anesthetised and intra-arterial pressure was recorded from carotid artery catheter. Live E. coli suspended in normal saline (5 Mcfarland concentration; dose – 650 uL/100 g body weight) was injected through the tail vein to induce sepsis. When mean arterial pressure dropped to 50% of its value before E. coli injection, Noradrenaline was injected in Group 2. Results: The average time (t1, n = 14) for the septic shock to set in was about 1.94 ± 0.97 h. Six out of seven rats (Group 1) died within 60 min without intervention. The addition of Noradrenaline after hypotension in Group 2 prolonged the time to death significantly by about 170 min. Conclusion: The rat septic shock model using E. coli described in the study is an acute, stable, and functional model to study various aspects of septic shock. Administration of Noradrenaline prolonged the animal’s life in septic shock as expected. Future studies using other common sepsis agents encountered in clinics can be undertaken similarly.

In vitro and in vivo Experimental Investigation of Anti-Inflammatory Effects of Peucedanum japonicum Aqueous Extract by Suppressing the LPS-Induced NF-[Formula: see text]B/MAPK JNK Pathways.

Peucedanum japonicum Thunberg has been used to treat cold, cough, and inflammatory diseases in Southern and Eastern Asia. The effects of P. japonicum root aqueous extract (PJ) on lipopolysaccharide (LPS)-induced inflammation were investigated in RAW264.7 macrophages and an animal model of septic shock. Lipopolysaccharides are endotoxins that trigger excessive inflammatory responses, similar to those elicited by gram-negative bacteria. Inflammation is characterized by a primary defense system against pathogens and the onset of sundry diseases or illnesses, and macrophages are important components of the phagocytic system during inflammatory processes. The present study evaluated the effects of PJ on the production of pro-inflammatory mediators, such as nitric oxide and prostaglandin E2, and assessed the expression of enzymes that induce the production of pro-inflammatory mediators using western blotting. We also evaluated the production and mRNA expression of pro-inflammatory cytokines using enzyme-linked immunosorbent assay and reverse transcription polymerase chain reaction, respectively. Using western blotting, we determined whether nuclear factor-kappa B (NF-[Formula: see text]B) and c-Jun N-terminal kinase (JNK) are involved in the molecular mechanisms induced by PJ that suppressed LPS-induced inflammatory responses. We also found that PJ inhibited NF-[Formula: see text]B and JNK pathways in macrophages and reduced LPS-induced mortality in the mouse model of septic shock by inhibiting the activation of NF-[Formula: see text]B and JNK pathways that downregulated the expression of inflammatory mediators. These results indicated that PJ is an effective inflammatory suppressor.

CD63 and C3AR1: The Potential Molecular Targets in the Progression of Septic Shock.

BackgroundThe molecular mechanism of septic shock is unknown. We studied the pathogenesis of septic shock and provide a novel strategy for treating and improving the prognosis of septic shock.MethodsGluten-Sensitive Enteropathy (GSE) 131761, GSE119217, GSE26378 datasets were downloaded from the Gene Expression Omnibus (GEO) database. The three datasets included 204 septic shock samples and 48 normal samples. The R packages “affy” and “limma” were employed to identify the differently expressed genes (DEGs) between septic shock and normal samples. Weighted gene co-expression network analysis (WGCNA) was performed to search for modules that play an important role in septic shock. Functional annotation of DEGs and construction and analysis of hub genes were used to explore the pathomechanism of septic shock. The receiver operating characteristic (ROC) curves were obtained using MedCalc software. The drug molecules that could regulate hub genes associated with septic shock were searched for in the CMap database. An animal model of septic shock was constructed to analyze the role of these hub genes.ResultsThe merged series contained 321 up-regulated and 255 down-regulated genes. WGCNA showed the brown module had the highest correlation with the status of septic shock. GO and KEGG enrichment analysis results of the brown module genes showed they were mainly enriched in “leukocyte differentiation”, “Ras-proximate-1 (Rap1) signaling pathway”, and “cytokine–cytokine receptor interaction”. Through construction and analysis of a protein–protein interaction (PPI) network, cluster of differentiation 63 (CD63) and complement component 3a receptor 1 (C3AR1) were identified as hub genes of septic shock. The area under curve (AUC) of C3AR1 for the septic shock is 0.772 (P<0.001), and the AUC of CD63 for the septic shock is 0.871 (P<0.001). Small molecule drugs were filtered by the number of instances (n>3) and P-values <0.05, including “monensin”, “verteporfin”, “ikarugamycin”, “tetrahydroalstonine”, “cefamandole”, “etoposide”. In the animal model, the relative expression levels of interleukin-6 (IL-6), Tumor Necrosis Factor-α (TNF-α), and lactic acid were significantly higher in the septic shock group compared with the control group. Results of Real Time Quantitative PCR (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA) analysis for CD63 and C3AR1 showed that their relative expression levels were significantly lower in the septic shock group compared with the control group (P<0.05).ConclusionCD63 and C3AR1 are significant hub genes of septic shock and may represent potential molecular targets for future studies of septic shock.

A double-blind, placebo-controlled, randomised, multicentre, proof-of-concept and dose-finding phase II clinical trial to investigate the safety, tolerability and efficacy of adrecizumab in patients with septic shock and elevated adrenomedullin concentration (AdrenOSS-2)

IntroductionSepsis remains a major health problem with an increasing incidence, high morbidity and high mortality. Apart from treatment with antibiotics and organ support, no approved specific adjunct therapies currently exist....

A bio-artificial renal epithelial cell system conveys survival advantage in a porcine model of septic shock.

Renal cell therapy using the hollow fiber based renal assist device (RAD) improved survival time in an animal model of septic shock (SS) through the amelioration of cardiac and vascular dysfunction. Safety and ability of the RAD to improve clinical outcomes was demonstrated in a Phase II clinical trial, in which patients had high prevalence of sepsis. Even with these promising results, clinical delivery of cell therapy is hampered by manufacturing hurdles, including cell sourcing, large-scale device manufacture, storage and delivery. To address these limitations, the bioartificial renal epithelial cell system (BRECS) was developed. The BRECS contains human renal tubule epithelial cells derived from adult progenitor cells using enhanced propagation techniques. Cells were seeded onto trabeculated disks of niobium-coated carbon, held within cryopreservable, perfusable, injection-moulded polycarbonate housing. The study objective was to evaluate the BRECS in a porcine model of SS to establish conservation of efficacy after necessary cell sourcing and design modifications; a pre-clinical requirement to move back into clinical trials. SS was incited by peritoneal injection of E. coli simultaneous to insertion of BRECS (n=10) or control (n=15), into the ultrafiltrate biofeedback component of an extracorporeal circuit. Comparable to RAD, prolonged survival of the BRECS cohort was conveyed through stabilization of cardiac output and vascular leak. In conclusion, the demonstration of conserved efficacy with BRECS therapy in a porcine SS model represents a crucial step toward returning renal cell therapy to the clinical setting, initially targeting ICU patients with acute kidney injury requiring continuous renal replacement therapy. Copyright © 2014 John Wiley & Sons, Ltd.

The relationship between cardiac filling pressures and fluid status in a controlled animal model of septic shock

The relationship between cardiac filling pressures and fluid status in a controlled animal model of septic shock

Adrenomedullin: its double-edged sword during sepsis slices yet again.

In this first issue of Intensive Care Medicine Experimental, two related studies describe the effects of antibodies directed against adrenomedullin (ADM), a vasoactive peptide that is released from various tissues during systemic inflammation, in murine sepsis. ADM, derived from a larger precursor peptide (pro-ADM), is released into the circulation during systemic inflammation, and the highest plasma concentrations are measured in patients with septic shock [1]. With a plasma half-life of approximately 20 min, it exerts multiple effects of which the most relevant appears to be vasodilation [2]. The consequent clinical sequelae of increased ADM concentrations are far from clear. While elevated ADM levels are associated with impaired outcome in sepsis patients [1] and the measurement of the circulating pro-peptide of ADM might serve as a marker to assess disease severity and predict mortality [3], administration of exogenous ADM actually improved outcome in animal models of septic shock [4]. Intriguingly, in this issue of Intensive Care Medicine Experimental, two studies demonstrate beneficial effects of anti-ADM antibodies administered in a murine cecal ligation and puncture (CLP) model [5,6]. These studies represent the first attempts at antibody targeting of the ADM cascade in vivo. Struck et al. [5] describe the development of monoclonal antibodies directed against various epitopes of ADM and their ability to attenuate sepsis-induced mortality in severe murine sepsis (100% control group mortality within 2 to 3 days). The severity of this model means that any longterm deleterious or beneficial effects of the antibody remain unknown. Notably, all antibodies directed against ADM improved survival. Of interest, the antibody against the N-terminal moiety of ADM prevented mortality most effectively, while it inhibited ADM agonistic activity by only 25%. Wagner et al. [6] performed a more mechanistic study into the effects of HAM1101, the antibody targeted against the N-terminal part of ADM, in a resuscitated murine CLP model. They show that HAM1101 improves vasoactive responsiveness to catecholamines, attenuates systemic inflammation, and improves kidney function. The beneficial effects on the kidneys are likely attributable to reduced iNOS expression and nitrotyrosine formation in HAM1101-treated animals. The effects on inflammatory parameters are surprising as ADM itself has also been reported to inhibit inflammation [7]. In summary, previous work has established that high endogenous levels of ADM correlate with poor outcomes and that exogenous administration improved outcomes, while now it is reported that antagonism of ADM using antibodies improves outcome in an animal sepsis model.

Effects of high volume hemofiltration on hemodynamics and oxygen metabolism at early stage of septic shock in piglet models

To observe the effects of hemofiltration at early stage of septic shock with different ultrafiltration doses, including hemodynamics, oxygen metabolism, inflammatory mediator in piglet models, and to evaluate the therapeutic effects of HVHF. The 18 healthy young piglets (Shanghai species) were divided randomly into three groups:control group (n = 6), conventional volume hemofiltration (CVVH) group [n = 6, ultrafiltration volume = 30 ml/(kg·h)] and high volume hemofiltration (HVHF) group [n = 6, ultrafiltration volume = 50 ml/(kg·h)], the animal model of septic shock was established by injection of lipopolysaccharide (LPS) (150 µg/kg) O111: B4. During the experiment, the following observations were carried out for all groups:1) Changes of hemodynamics [heart rate (HR), mean arterial pressure (MABP), cardiac output (CO), systemic vascular resistance index (SVRI), intrathoracic blood volume (ITBV)] and oxygen metabolism [oxygen delivery (DO2), oxygen consumption (VO2), oxygen extraction rate (O2ER) ] at the time of B0h, 0 h, 2 h, 4 h and 6 h.2) changes of TNF-α, IL-6, IL-10 in plasma at different time points (B0h, 0 h, 2 h, 4 h, 6 h). Significant difference in circulatory parameters, inflammatory mediators in plasma were found at B0h and 0 h among three groups; the CO in two treatment groups were higher than that in control group at 4 h, 6 h after model establishment (P < 0.05), and SVRI in HVHF groups were higher than that in other two groups at 4 h, 6 h after model was established (P < 0.05). The MABP in HVHF group [4 h (82 ± 17) mm Hg, 6 h (80 ± 12) mm Hg](1 mm Hg = 0.133 kPa) were higher than that in CVVH group at 4 h [(67 ± 12) mm Hg], 6 h [(69 ± 14) mm Hg] after model was established (P < 0.05). The levels of IL-6, IL-10, TNF-α in two treatment groups were lower than that in control group at 4 h and 6 h after model was established (P < 0.05), and the IL-6 [(281 ± 51) pg/ml], TNF-α [(67 ± 13) pg/ml] level in HVHF group was lower than that in CVVH group [IL-6(281 ± 51) pg/ml, TNF-α (67 ± 13) pg/ml] at 6 h (P < 0.05). The DO2 and VO2 in two treatment groups were higher than that in control group at 4 h, 6 h (P < 0.05), the O2ER in HVHF group were higher than that in CVVH group at 4 h (44% ± 3% vs. 33% ± 4%), 6 h (43% ± 5% vs. 31% ± 3%, P < 0.05). High volume hemofiltration (HVHF) at early stage of septic shock piglet models was more effective in improving hemodynamics, oxygen metabolism than conventional CVVH. And HVHF eliminated blood inflammatory mediators more effectively than conventional CVVH.

Role of Peroxynitrite in the Cardiovascular Dysfunction of Septic Shock

The intense systemic inflammatory response characterizing septic shock is associated with an increased generation of free radicals by multiple cell types in cardiovascular and non cardiovascular tissues. The oxygen-centered radical superoxide anion (O2 .-) rapidly reacts with the nitrogen-centered radical nitric oxide (NO.) to form the potent oxidant species peroxynitrite. Peroxynitrite oxidizes multiple targets molecules, either directly or via the secondary generation of highly reactive radicals, resulting in significant alterations in lipids, proteins and nucleic acids, with significant cytotoxic consequences. The formation of peroxynitrite is a key pathophysiological mechanism contributing to the cardiovascular collapse of septic shock, promoting vascular contractile failure, endothelial and myocardial dysfunction, and is also implicated in the occurrence of multiple organ dysfunction in this setting. The recent development of various porphyrin-based pharmacological compounds accelerating the degradation of peroxynitrite has allowed to specifically address these pathophysiological roles of peroxynitrite in experimental septic shock. Such agents, including 5,10,15,20-tetrakis(4- sulfonatophenyl)porphyrinato iron III chloride (FeTTPs), manganese tetrakis(4-N-methylpyridyl)porphyrin (MnTMPyP), Fe(III) tetrakis-2-(N-triethylene glycol monomethyl ether)pyridyl porphyrin) (FP-15) and WW-85, have been shown to improve the cardiovascular and multiple organ failure in small and large animal models of septic shock. Therefore, these findings support the development of peroxynitrite decomposition catalysts as potentially useful novel therapeutic agents to restore cardiovascular function in sepsis.

Concomitant use of beta-1 adrenoreceptor blocker and norepinephrine in patients with septic shock

Betablockade has been shown to have cardioprotective effects in patients under perioperative stress. Besides animal model of septic shock and a small cohort of septic patients, these benefits have not been studied in septic shock patients who require norepinephrine administration. After correction of preload, an esmolol bolus (0.2-0.5mg/kg) followed by continuous 24h infusion was administered in septic patients with sinus or supraventricular tachycardia (HR > 120/min). Exclusion criteria were severe LV systolic dysfunction, atrioventricular blockade and norepinephrine infusion at rates over 0.5mg/kg/min. Monitoring with echocardiography and pulmonary artery catheter before, at 2, 6, 12, 24h following the start and 6h after ceasing of the esmolol drip. Patients were maintained normovolemic throughout the study and adjustments of concomitant norepinephrine infusion rates were made as required. Ten septic patients (mean age 54.4 ± 18.7), APACHE II 21.5 ± 6.2, CRP 275 ± 78mg/l, procalcitonin 14.5 ± 10.1mg/l, were given esmolol drip of 212.5 ± 63.5mg/h at start to 272.5 ± 89.5mg/h at 24h. Heart rate decreased from mean 142 ± 11/min to 112 ± 9/min (p < 0.001) with parallel insignificant reduction of cardiac index (4.94 ± 0.76 to 4.35 ± 0.72l/min/m(2)). Stroke volume insignificantly increased from 67.1 ± 16.3ml to 72.9 ± 15.3ml. No parallel change of pulmonary artery wedge pressure was observed (15.9 ± 3.2 to 15.0 ± 2.4mmHg) as well as no significant changes of norepinephrine infusion (0.13 ± 0.17 to 0.17 ± 0.19mg/kg/min), DO(2), VO(2), OER or arterial lactate. Saving the heart 30beats/min did not demonstrate adverse impact on global haemodynamics in rates above 110/min. Using well titratable betablocker seems to be safe and cardioprotective in septic shock patients with high cardiac output.

24 Confocal Laser Endomicroscopy to Assess Mucosal Microcirculation: A Quantitative Analysis in a Porcine Model of Septic Shock and in Patients With Severe Sepsis

Aim Microcirculatory alterations play a central role in the pathophysiology of sepsis. Even if systemic hemodynamic parameters appear to be adequate, septic patients may suffer from significant intestinal microcirculatory alterations. It has been suggested that these changes play a central role in regard to morbidity and mortality. The present study first assessed the feasibility of In Vivo detection of mucosal microcirculation in different segments of the gastrointestinal tract in an animal model of septic shock using probe-based confocal laser endomicroscopy (pCLE) (Mauna Kea Technologies, Paris, France). Subsequently, we assessed duodenal microcirculation in patients suffering from severe sepsis in the early phase of the disease. Materials and Methods First, anesthetized and mechanically ventilated pigs were observed over 8 hrs. Septic shock was triggered by inducing fecal peritonitis (0.75 g autologous feces per kg body weight). Mucosal microcirculation was assessed simultaneously using pCLE in stomach, duodenum, terminal ileum and rectum at baseline, 4 hours after induction of septic shock as well as 2 hours after treatment of the condition. Second, six patients who presented with a severe sepsis on the intensive care unit were examined regarding their duodenal microcirculation after obtaining informed consent from their nearest relatives. These patients were compared to ten healthy controls. Four to six areas were examined for each site and images were analysed in a blinded fashion offline thereafter in both parts of the study. Mean capillary diameter, capillary length and functional capillary density (FCD) were measured quantitatively. Results Two hours after induction of sepsis in the animal model, FCD was markedly decreased in the duodenal (-20.8, p < 0.001), the ileal (-13.4, p < 0.001), the gastric (-11.9%, p < 0.001), and in the rectal mucosal beds (-5.5, p<0,01). After administration of 30 mg x kg-1 of gelatine (30 kDa) FCD increased in all mucosal compartments to 90.0% (duodenum), 94.4% (ileum), 95.4% (gastric) and 97% (rectum) of baseline values. Interestingly, mean vessel diameter was unchanged in all compartments investigated. In patients suffering from severe sepsis we found a significant decrease in mean vessel diameter (-5.0%, p<0.001) as well as in FCD (-9,4%, p<0.001) of the duodenal mucosa, as well. Conclusions During the early phase of septic shock, pCLE may be able to quantify microcirculatory alterations in the gastrointestinal mucosa. Fluid resuscitation improves but does not completely restore intestinal microcirculation in the septic shock model. pCLE may represent a useful tool in order to assess the efficacy of therapeutic interventions on mucosal microcirculation, possibly even in clinical practice.

DNA‐DEPENDENT PROTEIN KINASE IS CRITICAL FOR VCAM‐1 EXPRESSION UPON TNF TREATMENT THROUGH PHOSPHORYLATION OF p50 NF‐kappa B: A CRITICAL INVOLVEMENT IN VASCULAR AND LUNG INFLAMMATION

NF‐κB is critical for vascular cell adhesion molecule 1 (VCAM‐1) expression during vascular inflammation. We show that DNA‐dependent protein kinase (DNA‐PK) is required for VCAM‐1 expression in response to TNF‐α in an established cell line and in primary endothelial cells. DNA‐PK deficiency, achieved by DNA‐PKcs heterozygosity, reduced VCAM‐1 expression in the aorta in an animal model of septic shock and blocked airway neutrophilia in an animal model of acute lung injury. Interestingly, such effect unravels a novel function for DNA‐PK independent of its role in immune competence. Phosphorylation and nuclear translocation of p65 NF‐κB (p65) was insufficient for VCAM‐1 expression in response to TNF‐α. The requirement for p50 NF‐κB (p50) was associated with its interaction with and phosphorylation by DNA‐PK, which appears to be dominant over the requirement for p65 activation. Additionally, DNA‐PK activity appeared to increase the κB‐site binding ability of p50/p50 and p50/p65 dimers to VCAM‐1 promoter. Serine 20 in p50 was found to be phosphorylated by DNA‐PK. Re‐establishing wild‐type p50, but not its S20/A mutant, in p50‐knockout cells reversed VCAM‐1 expression. Together, these results elucidate a novel mechanism for the regulation of VCAM‐1 expression by DNA‐PK through phosphorylation of p50 and its critical involvement in vascular and lung inflammation.

Melatonin in Bacterial and Viral Infections with Focus on Sepsis: A Review

Melatonin is a versatile molecule, synthesized not only by the pineal gland, but also in small amounts by many other organs like retina, gastrointestinal tract, thymus, bone marrow, lymphocytes etc. It plays an important role in various functions of the body like sleep and circadian rhythm regulation, immunoregulatory mechanism, free radical scavenger, antioxidant functions, oncostatic actions, control of reproductive functions, regulation of mood etc. Melatonin has also been found to be effective in combating various bacterial and viral infections. Its administration has been shown to be effective in controlling chlamydial infections, infections induced by Mycobacterium tuberculosis, and also in many viral infections. Molecular mechanisms of anti microbial actions of melatonin have suggested to be due to effects on free radical formation, direct regulation of duplication of bacteria, depletion of intracellular substrates like iron etc. Besides, it is effective in sepsis as demonstrated in various animal models of septic shock. Melatonin's protective action against sepsis is suggested to be due to its antioxidant, immunomodulating and inhibitory actions against the production and activation of pro-inflammatory mediators. Use of melatonin has been beneficial in treating premature infants suffering from severe respiratory distress syndrome and septic shock. It has a potential therapeutic value in treating septic shock and associated multi organ failure in critically ill patients in addition to its antimicrobial and antiviral actions. The patents related to melatonin's use for treatment of bacterial infections and its use in clinical disorders are included.

Effects of a Novel Anticoagulant Compound (TV7130) in an Ovine Model of Septic Shock

We compared the effects of a new compound (TV7130) with those of activated protein C (APC) in a large animal model of septic shock. Thirty-two fasted, anesthetized, invasively monitored, mechanically ventilated female sheep received 1.5 g/kg body weight of feces into the abdomen to induce sepsis. Immediately after feces injection, all animals received a bolus followed by a continuous infusion of saline (n = 8, bolus 1.5 mL for 15 min, infusion 1.5 mL/[kg·h]), low-dose TV7130 (n = 8; 0.4 mg/kg bolus, 0.4 mg/[kg·h] infusion), high-dose TV7130 (n = 8; 0.8 mg/kg bolus, 0.8 mg/[kg·h] infusion), or APC (n = 8; saline bolus, APC infusion of 0.024 mg/[kg·h]). Experiments were pursued until each sheep's spontaneous death. There were no significant differences among groups in heart rate or cardiac index, but mean arterial pressure, systemic vascular resistance index, and left ventricular stroke work index decreased less in the high-dose TV7130 and APC groups than in the other groups. Gas exchange was preserved better in the high-dose TV7130 and APC groups. Interleukin 6 and lactate concentrations were lower in the high-dose TV7130 and APC groups than in the other groups. Functional capillary density and proportion of perfused vessels, evaluated in the sublingual region using sidestream dark-field videomicroscopy, were significantly higher in the TV7130 and APC groups than in the vehicle group at 16 h. Survival time was significantly longer in the high-dose TV7130 and APC groups than in the other groups (log-rank test, P = 0.0002). TV7130 has similar effects to APC and may be a promising agent for the management of severe sepsis.

An ovine intensive care model of septic shock

Translation of previous animal studies into human ICU clinical trials has frequently produced negative results. Most of these animal studies have had high baseline mortality and have not employed standardised management of sepsis as usually provided in an ICU. The aim of this study was to develop a large animal model of septic shock receiving standardised intensive care management, thus replicating the management of septic shock in humans.

Septic shock induces distinct changes in sympathetic nerve activity to the heart and kidney in conscious sheep

Sepsis and septic shock are the chief cause of death in intensive care units, with mortality rates between 30 and 70%. In a large animal model of septic shock, we have demonstrated hypotension, increased cardiac output, and tachycardia, together with renal vasodilatation and renal failure. The changes in cardiac sympathetic nerve activity (CSNA) that may contribute to the tachycardia have not been investigated, and the changes in renal SNA (RSNA) that may mediate the changes in renal blood flow and function are unclear. We therefore recorded CSNA and RSNA during septic shock in conscious sheep. Septic shock was induced by administration of Escherichia coli, which caused a delayed hypotension and an immediate, biphasic increase in heart rate (HR) associated with similar changes in CSNA. After E. coli, RSNA decreased for over 3 h, followed by a sustained increase (180%), whereas renal blood flow progressively increased and remained elevated. There was an initial diuresis, followed by oliguria and decreased creatinine clearance. There were differential changes in the range of the arterial baroreflex curves; it was depressed for HR, increased for CSNA, and unchanged for RSNA. Our findings, recording CSNA for the first time in septic shock, suggest that the increase in SNA to the heart is not driven solely by unloading of baroreceptors and that the increase has an important role to increase HR and cardiac output. There was little correlation between the changes in RSNA and renal blood flow, suggesting that the renal vasodilatation was mediated mainly by other mechanisms.

Activation of the PGI 2 /IP System Contributes to the Development of Circulatory Failure in a Rat Model of Endotoxic Shock

Prostacyclin levels are increased in septic patients and several animal models of septic shock, and selective inhibition of cyclooxygenase-2 improved cardiovascular dysfunction in rats treated with lipopolysaccharide (LPS). Here, we examine the specific role of prostacyclin and of the receptor for prostacyclin (IP) in the development of LPS-induced circulatory failure. Intravenous injection of LPS (10 mg/kg) into male Sprague-Dawley rats caused a strong increase in plasma prostacyclin levels, which was paralleled by a decrease in blood pressure and an increase in heart rate. Moreover, LPS injection increased the mRNA expression of the IP receptor in the heart, aorta, lung, liver, adrenal glands, and kidneys. Cotreatment with the IP antagonist CAY-10441 (1, 10, 30, and 100 mg/kg) dose-dependently moderated the LPS-induced changes in mean arterial blood pressure, heart rate, cardiac output, and systemic vascular resistance. The development of cardiovascular failure was ameliorated by CAY-10441 in spite of the typical LPS-induced increases in plasma levels of cytokines and NO. In vitro, cytokines dose- and time-dependently induced IP expression in rat vascular smooth muscle cells. Incubation of cells with the stable IP agonist iloprost in the presence of the phosphodiesterase inhibitor 3-isobutyl-1-mehylxanthine resulted in higher cAMP levels in cytokine-treated cells compared with untreated cells. Taken together, our data demonstrate a prominent role of the prostacyclin/IP system in the development of LPS-induced cardiovascular failure.