[Accidental Awareness during General Anaesthesia]. / Akzidentelle Wachheit während Allgemeinanästhesie.

Accidental awareness during general anaesthesia (AAGA) is a rare but severe complication. The reported incidence of AAGA may depend on the assessment of intraoperative awareness with explicit recall and there are substantial variations between subspecialties and groups of patients. The majority of prospective studies using structured interviews reported an incidence of AAGA at 0.1-0.2% during general anaesthesia, however, higher values were observed in paediatric (0.2-1.2%) and obstetric patients (0.47%). Risk factors that predispose to AAGA are patient conditions, ASA status, female gender, patient age, history of AAGA, surgical procedure, anaesthetic drug type, muscle relaxation, dosages of hypnotic or analgesic drugs, monitoring and malfunction of anaesthesia systems. Preventive strategies include careful assessment of risk factors, avoidance of underdosages of hypnotics and analgetics during general anaesthesia and monitoring of depth of anaesthesia in risk patients. The health-related consequences can be serious and psychopharmacological and psychotherapeutic interventions are indicated in patients who have experienced AAGA.

Learning in the Single-Cell Organism Physarum polycephalum: Effect of Propofol.

Propofol belongs to a class of molecules that are known to block learning and memory in mammals, including rodents and humans. Interestingly, learning and memory are not tied to the presence of a nervous system. There are several lines of evidence indicating that single-celled organisms also have the capacity for learning and memory which may be considered as basal intelligence. Here, we introduce a new experimental model for testing the learning ability of Physarum polycephalum, a model organism frequently used to study single-celled "intelligence". In this study, the impact of propofol on Physarum's "intelligence" was tested. The model consists of a labyrinth of subsequent bifurcations in which food (oat flakes soaked with coconut oil-derived medium chain triglycerides [MCT] and soybean oil-derived long chain triglycerides [LCT]) or propofol in MCT/LCT) is placed in one of each Y-branch. In this setting, it was tested whether Physarum memorized the rewarding branch. We saw that Physarum was a quick learner when capturing the first bifurcations of the maze; thereafter, the effect decreased, perhaps due to reaching a state of satiety. In contrast, when oat flakes were soaked with propofol, Physarum's preference for oat flakes declined significantly. Several possible actions, including the blocking of gamma-aminobutyric acid (GABA) receptor signaling, are suggested to account for this behavior, many of which can be tested in our new model.

Cholinesterase alterations in delirium after cardiosurgery: a German monocentric prospective study.

OBJECTIVES: Postoperative delirium (POD) is a common complication after elective cardiac surgery. Recent evidence indicates that a disruption in the normal activity of the cholinergic system may be associated with delirium. DESIGN: Prospective observational study. SETTING: Single-centre at a European academic hospital. PRIMARY AND SECONDARY OUTCOME MEASURES: In our study the enzyme activities of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) were determined preoperatively as well as on the first and second postoperative day. The confusion assessment method for the intensive care unit was used to screen patients for the presence of POD. RESULTS: A total of 114 patients were included in the study. POD was associated with a decrease in BChE activity on postoperative day 1 (p=0.03). In addition, patients who developed POD, had significantly lower preoperative AChE activity than patients without POD (p<0.01). Multivariate analysis identified a preoperatively decreased AChE activity (OR 3.1; 95% CI 1.14 to 8.46), anticholinergic treatment (OR 5.09; 95% CI 1.51 to 17.23), elevated European System for Cardiac Operative Risk Evaluation (OR 3.68; 95% CI 1.04 to 12.99) and age (OR 3.02; 95% CI 1.06 to 8.62) to be independently associated with the development of POD. CONCLUSIONS: We conclude that a reduction in the acetylcholine hydrolysing enzyme activity in patients undergoing cardiac surgery may correlate with the development of POD.

Acute ketamine dysregulates task-related gamma-band oscillations in thalamo-cortical circuits in schizophrenia.

Hypofunction of the N-methyl-d-aspartate receptor (NMDAR) has been implicated as a possible mechanism underlying cognitive deficits and aberrant neuronal dynamics in schizophrenia. To test this hypothesis, we first administered a sub-anaesthetic dose of S-ketamine (0.006 mg/kg/min) or saline in a single-blind crossover design in 14 participants while magnetoencephalographic data were recorded during a visual task. In addition, magnetoencephalographic data were obtained in a sample of unmedicated first-episode psychosis patients (n = 10) and in patients with chronic schizophrenia (n = 16) to allow for comparisons of neuronal dynamics in clinical populations versus NMDAR hypofunctioning. Magnetoencephalographic data were analysed at source-level in the 1-90 Hz frequency range in occipital and thalamic regions of interest. In addition, directed functional connectivity analysis was performed using Granger causality and feedback and feedforward activity was investigated using a directed asymmetry index. Psychopathology was assessed with the Positive and Negative Syndrome Scale. Acute ketamine administration in healthy volunteers led to similar effects on cognition and psychopathology as observed in first-episode and chronic schizophrenia patients. However, the effects of ketamine on high-frequency oscillations and their connectivity profile were not consistent with these observations. Ketamine increased amplitude and frequency of gamma-power (63-80 Hz) in occipital regions and upregulated low frequency (5-28 Hz) activity. Moreover, ketamine disrupted feedforward and feedback signalling at high and low frequencies leading to hypo- and hyper-connectivity in thalamo-cortical networks. In contrast, first-episode and chronic schizophrenia patients showed a different pattern of magnetoencephalographic activity, characterized by decreased task-induced high-gamma band oscillations and predominantly increased feedforward/feedback-mediated Granger causality connectivity. Accordingly, the current data have implications for theories of cognitive dysfunctions and circuit impairments in the disorder, suggesting that acute NMDAR hypofunction does not recreate alterations in neural oscillations during visual processing observed in schizophrenia.

The Fibrin Cleavage Product Bß15-42 Channels Endothelial and Tubular Regeneration in the Post-acute Course During Murine Renal Ischemia Reperfusion Injury.

Early and adequate restoration of endothelial and tubular renal function is a substantial step during regeneration after ischemia reperfusion (IR) injury, occurring, e.g., in kidney transplantation, renal surgery, and sepsis. While tubular epithelial cell injury has long been of central importance, recent perception includes the renal vascular endothelium. In this regard, the fibrin cleavage product fibrinopeptide Bß15-42 mitigate IR injury by stabilizing interendothelial junctions through its affinity to VE-cadherin. Therefore, this study focused on the effect of Bß15-42 on post-acute physiological renal regeneration. For this, adult male C57BL/6 mice were exposed to a 30 min bilateral renal ischemia and reperfusion for 24 h or 48 h. Animals were randomized in a non-operative control group, two operative groups each treated with i.v. administration of either saline or Bß15-42 (2.4 mg/kg) immediately prior to reperfusion. Endothelial activation and inflammatory response was attenuated in renal tissue homogenates by single application of Bß15-42. Meanwhile, Bß15-42 did not affect acute kidney injury markers. Regarding the angiogenetic players VEGF-A, Angiopoietin-1, Angiopoietin-2, however, we observed significant higher expressions at mRNA and trend to higher protein level in Bß15-42 treated mice, compared to saline treated mice after 48 h of IR, thus pointing toward an increased angiogenetic activity. Similar dynamics were observed for the intermediate filament vimentin, the cytoprotective protein klotho, stathmin and the proliferation cellular nuclear antigen, which were significantly up-regulated at the same points in time. These results suggest a beneficial effect of anatomical contiguously located endothelial cells on tubular regeneration through stabilization of endothelial integrity. Therefore, it seems that Bß15-42 represents a novel pharmacological approach in the targeted therapy of acute renal failure in everyday clinical practice.

Systemic TLR2 Antibody Application in Renal Ischaemia and Reperfusion Injury Decreases AKT Phosphorylation and Increases Apoptosis in the Mouse Kidney.

Acute kidney injury remains an important cause of renal dysfunction. In this context, Toll-like receptors have been demonstrated to play a critical role in the induction of innate and inflammatory responses. Among these, Toll-like receptor 2 (TLR2) is constitutively expressed in tubular epithelial cells (TECs) of the kidney and is also known to mediate ischaemia reperfusion (IR) injury. Adult male C57BL/6JRj mice were randomized into seven groups (n = 8): a non-operative control group (CTRL) and six interventional groups in which mice were subjected to a 30 min. bilateral renal ischaemia. Immediately before reperfusion, mice were treated either with saline or with TLR2 antibody (clone T2.5) and harvested after ischaemia and reperfusion for 3, 24 and 48 hr. Analysed kidney homogenates of TLR2 antibody-treated mice displayed significantly decreased levels of TLR2 protein after 3 hr of IR compared to saline-treated mice. Accordingly, the degree of AKT phosphorylation was significantly decreased after 3 hr of IR compared to saline-treated animals. TUNEL staining revealed significantly higher apoptosis rates in TLR2 antibody-treated animals compared to saline-treated mice after 3 and 24 hr of IR. Further, a positive correlation between TLR2 protein expression and phosphorylation of AKT as well as a negative correlation with the number of TUNEL-positive cells could be observed. Inhibition of TLR2 and its signalling pathway by a single application of TLR2 antibody results in reduced phosphorylation of AKT and consecutively increased apoptosis.

Mortality of Septic Mice Strongly Correlates With Adrenal Gland Inflammation.

OBJECTIVES: Sepsis and septic shock are commonly present in the ICU and accompanied by significant morbidity, mortality, and cost. The frequency of secondary adrenal insufficiency in sepsis remains open to debate and a challenge to identify and treat appropriately. Animal models of sepsis using genetic or surgical initiation of adrenal insufficiency resulted in increased mortality, but the mechanisms are still unclear. The present study investigates the impact of adrenal inflammation in septic mice challenged with cecal ligation and puncture. DESIGN: Prospective experimental study. SETTING: University laboratory. SUBJECTS: C57BL/6N wild-type mice. INTERVENTIONS: Sepsis, induced by cecal ligation and puncture for 24 and 48 hours. MEASUREMENTS AND MAIN RESULTS: Both septic and control mice were carefully monitored (every 30 min) for up to 48 hours and divided into survivors and nonsurvivors. We observed a significant and massive increase of interleukin-6, interleukin-1ß, and tumor necrosis factor-α in adrenal protein extracts of nonsurvivors compared with sham animals and survivors. This pattern was partly reflected in liver and lung but not in plasma samples. Notably, a significant increase in nonsurvivors compared with survivors was only found for lung interleukin-6. In line with these findings, we detected a higher degree of leukocyte infiltration and hemorrhage in the adrenal glands of deceased mice. Evaluation of the hypothalamic-pituitary-adrenal axis response in these animals revealed an increase of adrenocorticotropic hormone, which was only partly reflected in the corticosterone level. Notably, using the adrenocorticotropic hormone stimulation test, we found an impaired adrenocorticotropic hormone response, particularly in nonsurvivors, which significantly correlated with the number of infiltrated leukocytes. CONCLUSIONS: Cecal ligation and puncture-induced murine sepsis induces a strong inflammatory response in the adrenal glands, which is accompanied by cell death and hemorrhage. Our data suggest that mortality and adrenal incapacitation are associated with the degree of adrenal inflammation, thereby underscoring the importance of adrenal function on survival.

Toll-like receptor 3 signalling mediates angiogenic response upon shock wave treatment of ischaemic muscle.

AIMS: Shock wave therapy (SWT) represents a clinically widely used angiogenic and thus regenerative approach for the treatment of ischaemic heart or limb disease. Despite promising results in preclinical and clinical trials, the exact mechanism of action remains unknown. Toll-like receptor 3, which is part of the innate immunity, is activated by binding double-stranded (ds) RNA. It plays a key role in inflammation, a process that is needed also for angiogenesis. We hypothesize that SWT causes cellular cavitation without damaging the target cells, thus liberating cytoplasmic RNA that in turn activates TLR3. METHODS AND RESULTS: SWT induces TLR3 and IFN-ß1 gene expression as well as RNA liberation from endothelial cells in a time-dependant manner. Conditioned medium from SWT-treated HUVECs induced TLR3 signalling in reporter cells. The response was lost when the medium was treated with RNase III to abolish dsRNAs or when TLR3 was silenced using siRNAs. In a mouse hind limb ischaemia model using wt and TLR3(-/-) mice (n = 6), SWT induced angiogenesis and arteriogenesis only in wt animals. These effects were accompanied by improved blood perfusion of treated limbs. Analysis of main molecules of the TLR3 pathways confirmed TLR3 signalling in vivo following SWT. CONCLUSION: Our data reveal a central role of the innate immune system, namely Toll-like receptor 3, to mediate angiogenesis upon release of cytoplasmic RNAs by mechanotransduction of SWT.

Individual Organ Failure and Concomitant Risk of Mortality Differs According to the Type of Admission to ICU - A Retrospective Study of SOFA Score of 23,795 Patients.

INTRODUCTION: Organ dysfunction or failure after the first days of ICU treatment and subsequent mortality with respect to the type of intensive care unit (ICU) admission is poorly elucidated. Therefore we analyzed the association of ICU mortality and admission for medical (M), scheduled surgery (ScS) or unscheduled surgery (US) patients mirrored by the occurrence of organ dysfunction/failure (OD/OF) after the first 72h of ICU stay. METHODS: For this retrospective cohort study (23,795 patients; DIVI registry; German Interdisciplinary Association for Intensive Care Medicine (DIVI)) organ dysfunction or failure were derived from the Sequential Organ Failure Assessment (SOFA) score (excluding the Glasgow Coma Scale). SOFA scores were collected on admission to ICU and 72h later. For patients with a length of stay of at least five days, a multivariate analysis was performed for individual OD/OF on day three. RESULTS: M patients had the lowest prevalence of cardiovascular failure (M 31%; ScS 35%; US 38%), and the highest prevalence of respiratory (M 24%; ScS 13%; US 17%) and renal failure (M 10%; ScS 6%; US 7%). Risk of death was highest for M- and ScS-patients in those with respiratory failure (OR; M 2.4; ScS 2.4; US 1.4) and for surgical patients with renal failure (OR; M 1.7; ScS 2.7; US 2.4). CONCLUSION: The dynamic evolution of OD/OF within 72h after ICU admission and mortality differed between patients depending on their types of admission. This has to be considered to exclude a systematic bias during multi-center trials.

Combined peri-ischemic administration of Bß15-42 in treating ischemia reperfusion injury of the mouse kidney.

The disruption of endothelial integrity is a crucial step for the development of vascular leakage and consequently ischemia-reperfusion injury (IRI). Regarding the molecular cell-cell interaction, the fibrinopeptide Bß15-42 prevents vascular leakage by stabilizing the inter-endothelial junctions via association with the vascular endothelial-cadherin. In a previous study we showed that a renoprotective effect in early IRI may be achieved by intravenous administration of Bß15-42 at the time of reperfusion. We now aimed to investigate whether additional pre-ischemic application of Bß15-42 could enhance this effect. Therefore C57BL/6 mice were subjected to 0.5h bilateral renal ischemia followed by reperfusion. The animals were randomized into 6 groups (n=6): two control groups treated with i.v. administration of NaCl at reperfusion for 0.5h (NaCl 1h) and 2.5h (NaCl 3h), two groups with Bß15-42 at reperfusion for 0.5h (Bß(rep) 1h) and 2.5h (Bß(rep) 3h), and two groups with administration of Bß15-42 immediately pre-ischemic as well as at reperfusion for 0.5h (Bß(peri) 1h) and 2.5h (Bß(peri) 3h). We found that both Bß(rep) and Bß(peri) mice displayed reduced early renal damage compared with NaCl treated mice. However, there was no further reduction of the IR damage through added pre-ischemic application of Bß15-42. Overall, we detected significantly reduced endothelial activation, lower tissue infiltration of neutrophils as well as lower tissue levels of neutrophil gelatinase-associated lipocalin (NGAL) in all mice treated with Bß15-42 compared to mice treated with NaCl. Our data confirm the renoprotective effect of Bß15-42 in the early therapeutic treatment of acute kidney injury due to ischemia and reperfusion. However, a combined pre-and post-ischemic administration of Bß15-42 appears to provide no additional benefit compared with a sole administration at reperfusion.

Ketamine Dysregulates the Amplitude and Connectivity of High-Frequency Oscillations in Cortical-Subcortical Networks in Humans: Evidence From Resting-State Magnetoencephalography-Recordings.

Hypofunctioning of the N-methyl-D-aspartate receptor (NMDA-R) has been prominently implicated in the pathophysiology of schizophrenia (ScZ). The current study tested the effects of ketamine, a dissociative anesthetic and NMDA-R antagonist, on resting-state activity recorded with magnetoencephalography (MEG) in healthy volunteers. In a single-blind cross-over design, each participant (n = 12) received, on 2 different sessions, a subanesthetic dose of S-ketamine (0.006 mg/Kg) and saline injection. MEG-data were analyzed at sensor- and source-level in the beta (13-30 Hz) and gamma (30-90 Hz) frequency ranges. In addition, connectivity analysis at source-level was performed using transfer entropy (TE). Ketamine increased gamma-power while beta-band activity was decreased. Specifically, elevated 30-90 Hz activity was pronounced in subcortical (thalamus and hippocampus) and cortical (frontal and temporal cortex) regions, whilst reductions in beta-band power were localized to the precuneus, cerebellum, anterior cingulate, temporal and visual cortex. TE analysis demonstrated increased information transfer in a thalamo-cortical network after ketamine administration. The findings are consistent with the pronounced dysregulation of high-frequency oscillations following the inhibition of NMDA-R in animal models of ScZ as well as with evidence from electroencephalogram-data in ScZ-patients and increased functional connectivity during early illness stages. Moreover, our data highlight the potential contribution of thalamo-cortical connectivity patterns towards ketamine-induced neuronal dysregulation, which may be relevant for the understanding of ScZ as a disorder of disinhibition of neural circuits.

Contribution of Ninjurin1 to Toll-like receptor 4 signaling and systemic inflammation.

Nerve injury-induced protein (Ninjurin [Ninj]) 1 is an adhesion molecule originally identified in Schwann cells after nerve injury, whereas it is also expressed in leukocytes, epithelium, endothelium, and various organs, and is induced under inflammatory conditions. Its contribution to inflammation was so far restricted to the nervous system and exclusively attributed to its role during leukocyte migration. We hypothesized a proinflammatory role for Ninj1 also outside the nervous system. To elucidate its impact during inflammation, we analyzed expression levels and its contribution to inflammation in septic mice and studied its effect on inflammatory signaling in vitro. The effect on inflammation was analyzed by genetic (only in vitro) and pharmacologic repression in septic mice (cecal ligation and puncture) and cell culture, respectively. Repression of Ninj1 by an inhibitory peptide or small interfering RNA attenuated LPS-triggered inflammation in macrophages and endothelial cells by modulating p38 phosphorylation and activator protein-1 activation. Inhibition of Ninj1 in septic mice reduced systemic and pulmonary inflammation as well as organ damage, and ameliorated survival after 24 hours. Ninj1 is elevated under inflammatory conditions and contributes to inflammation not only by mediating leukocyte migration, but also by modulating Toll-like receptor 4-dependent expression of inflammatory mediators. We assume that, owing to both mechanisms, inhibition reduces systemic inflammation and organ damage in septic mice. Our data contribute to a better understanding of the complex inflammatory mechanisms and add a novel therapeutic target for inflammatory conditions such as sepsis.

Intestinal Fatty Acid Binding Protein: A Sensitive Marker in Abdominal Surgery and Abdominal Infection.

BACKGROUND: Intestinal fatty acid binding protein (iFABP) is elevated in plasma by intestinal injury. We investigated the influence of surgical trauma and severe sepsis caused by abdominal and pulmonary infection on plasma iFABP concentrations. METHODS: Seventy-nine patients were included in this prospective observational study: 31 patients before elective major abdominal surgery (EMS), 33 patients with severe sepsis on admission to the intensive care unit (ICU), and 15 healthy volunteers who served as controls. Blood samples were taken before and after surgery for a period up to 5 d. RESULTS: Prior to surgery, EMS patients had increased iFABP concentrations in those patients with intestinal cancer compared with patients without intestinal cancer (217 pg/mL, interquartile range [IQR] I-III 100-369 pg/mL versus 79 pg/mL, IQR I-III: 0-182 pg/mL; p<0.01) and with controls (114 pg/mL, IQR I-III: 103-124 pg/mL; p<0.01). Surgical trauma increased iFABP levels in patients without intestinal cancer (240 pg/mL, IQR I-III 111-305 pg/mL; p<0.01). Within 24 h after surgery, iFABP levels decreased to normal values. Patients with severe sepsis of abdominal origin had elevated concentrations compared with controls (324 pg/mL [IQR I-III 0-649 pg/mL]; p=0.05); in patients with pneumonia, iFABP levels were not significantly increased. Discrimination between intestinal- and pulmonary-induced sepsis was low (area under the curve [AUC] 0.693; 95% confidence interval 0.512-0.874). CONCLUSIONS: Surgical trauma and severe sepsis lead to elevated iFABP concentrations. However, intestinal malignant disease and in some patients severe sepsis caused by pneumonia also resulted in elevated iFABP concentrations. The results support the idea that epithelial injury of many causes leads to elevated concentrations of iFABP. The value of iFABP for differentiating pulmonary from intestinal sepsis is limited.

Preoperative interleukin-22 values add valuable information for outcome prediction following orthotopic liver transplantation: a preliminary study.

BACKGROUND: Recent findings support the idea that interleukin (IL)-22 serum levels are related to disease severity in end-stage liver disease. Existing scoring systems--Model for End-Stage Liver Disease (MELD), Survival Outcomes Following Liver Transplantation (SOFT) and Pre-allocation-SOFT (P-SOFT)--are well-established in appraising survival rates with or without liver transplantation. We tested the hypothesis that IL-22 serum levels at transplantation date correlate with survival and potentially have value as a predictive factor for survival. MATERIAL AND METHODS: MELD, SOFT, and P-SOFT scores were calculated to estimate post-transplantation survival. Serum levels of IL-22, IL-6, IL-10, C-reactive protein (CRP), and procalcitonin (PCT) were collected prior to transplantation in 41 patients. Outcomes were assessed at 3 months, 1 year, and 3 years after transplantation. RESULTS: IL-22 significantly correlated with MELD, P-SOFT, and SOFT scores (Rs 0.35, 0.63, 0.56 respectively, p<0.05) and with the discrimination in post-transplantation survival. IL-6 showed a heterogeneous pattern (Rs 0.40, 0.63, 0.57, respectively, p<0.05); CRP and PCT did not correlate. We therefore added IL-22 serum values to existing scoring systems in a generalized linear model (GLM), resulting in a significantly improved outcome prediction in 58% of the cases for both the P-SOFT (p<0.01) and SOFT scores (p<0.001). CONCLUSIONS: Further studies are needed to address the concept that IL-22 serum values at the time of transplantation provide valuable information about survival rates following orthotopic liver transplantation.

The early activation of toll-like receptor (TLR)-3 initiates kidney injury after ischemia and reperfusion.

Acute kidney injury (AKI) is one of the most important complications in hospitalized patients and its pathomechanisms are not completely elucidated. We hypothesize that signaling via toll-like receptor (TLR)-3, a receptor that is activated upon binding of double-stranded nucleotides, might play a crucial role in the pathogenesis of AKI following ischemia and reperfusion (IR). Male adult C57Bl6 wild-type (wt) mice and TLR-3 knock-out (-/-) mice were subjected to 30 minutes bilateral selective clamping of the renal artery followed by reperfusion for 30 min 2.5h and 23.5 hours or subjected to sham procedures. TLR-3 down-stream signaling was activated already within 3 h of ischemia and reperfusion in post-ischemic kidneys of wt mice lead to impaired blood perfusion followed by a strong pro-inflammatory response with significant neutrophil invasion. In contrast, this effect was absent in TLR-3-/- mice. Moreover, the quick TLR-3 activation resulted in kidney damage that was histomorphologically associated with significantly increased apoptosis and necrosis rates in renal tubules of wt mice. This finding was confirmed by increased kidney injury marker NGAL in wt mice and a better preserved renal perfusion after IR in TLR-3-/- mice than wt mice. Overall, the absence of TLR-3 is associated with lower cumulative kidney damage and maintained renal blood perfusion within the first 24 hours of reperfusion. Thus, we conclude that TLR-3 seems to participate in the pathogenesis of early acute kidney injury.

VEGF-A blockade reduces reperfusion edema but favors arterial thromboembolism in a rat model of orthotopic lung transplantation.

BACKGROUND: Ischemia-reperfusion edema is a common early complication after lung transplantation where the hypoxia-induced vascular endothelial growth factor (VEGF)-A plays a pivotal role. It remains unclear whether a VEGF blockade is beneficial in lung transplantation. METHODS: VEGF-A blockade was investigated in an orthotopic rat model of lung transplantation. VEGF-A antibody was added into the preservation solution alone (α-VEGF D/-), in the preservation solution and systemically to the recipient before reperfusion (α-VEGF D/R), or applied to the recipient alone before reperfusion (α-VEGF -/R). Forty-eight hours after lung transplantation, left lungs were collected and wet-to-dry ratio, Western blotting, RT-PCR, and immunohistology were performed. RESULTS: VEGF-A blockade in α-VEGF D/-, α-VEGF D/R, and α-VEGF -/R resulted in neutralization of tissue VEGF-A. Reperfusion edema was only reduced in α-VEGF D/R and α-VEGF D/- groups versus Perfadex controls. Some α-VEGF -/R rats showed a hyperinflammation leading to increased pro-inflammatory cytokine expressions as well as increased edema. Whereas generally the α-VEGF D/- group showed decreased inflammation, the combination with anti-VEGF treatment to the recipient resulted in a pro-inflammatory and a pro-apoptotic phenotype. Short-term survival, however, was not significantly different in all groups as compared to the controls. In the α-VEGF (D/R) or (D/-) groups, animals mainly died from arterial thromboembolisms and in the α-VEGF (-/R) group, hyperinflammation was the main cause of death. CONCLUSION: VEGF-A directly contributes to the formation of a reperfusion edema, which might be reduced by its blockade. However, the α-VEGF effect on the endothelial integrity might also favor arterial thrombosis formation.

The small fibrinopeptide Bß15-42 as renoprotective agent preserving the endothelial and vascular integrity in early ischemia reperfusion injury in the mouse kidney.

Disruption of the renal endothelial integrity is pivotal for the development of a vascular leak, tissue edema and consequently acute kidney injury. Kidney ischemia amplifies endothelial activation and up-regulation of pro-inflammatory mechanisms. After restoring a sufficient blood flow, the kidney is damaged through complex pathomechanisms that are classically referred to as ischemia and reperfusion injury, where the disruption of the inter-endothelial connections seems to be a crucial step in this pathomechanism. Focusing on the molecular cell-cell interaction, the fibrinopeptide Bß15-42 prevents vascular leakage by stabilizing these inter-endothelial junctions. The peptide associates with vascular endothelial-cadherin, thus preventing early kidney dysfunction by preserving blood perfusion efficacy, edema formation and thus organ dysfunction. We intended to demonstrate the early therapeutic benefit of intravenously administered Bß15-42 in a mouse model of renal ischemia and reperfusion. After 30 minutes of ischemia, the fibrinopeptide Bß15-42 was administered intravenously before reperfusion was commenced for 1 and 3 hours. We show that Bß15-42 alleviates early functional and morphological kidney damage as soon as 1 h and 3 h after ischemia and reperfusion. Mice treated with Bß15-42 displayed a significantly reduced loss of VE-cadherin, indicating a conserved endothelial barrier leading to less neutrophil infiltration which in turn resulted in significantly reduced structural renal damage. The significant reduction in tissue and serum neutrophil gelatinase-associated lipocalin levels reinforced our findings. Moreover, renal perfusion analysis by color duplex sonography revealed that Bß15-42 treatment preserved resistive indices and even improved blood velocity. Our data demonstrate the efficacy of early therapeutic intervention using the fibrinopeptide Bß15-42 in the treatment of acute kidney injury resulting from ischemia and reperfusion. In this context Bß15-42 may act as a potent renoprotective agent by preserving the endothelial and vascular integrity.

Prednisolone as preservation additive prevents from ischemia reperfusion injury in a rat model of orthotopic lung transplantation.

The lung is, more than other solid organs, susceptible for ischemia reperfusion injury after orthotopic transplantation. Corticosteroids are known to potently suppress pro-inflammatory processes when given in the post-operative setting or during rejection episodes. Whereas their use has been approved for these clinical indications, there is no study investigating its potential as a preservation additive in preventing vascular damage already in the phase of ischemia. To investigate these effects we performed orthotopic lung transplantations (LTX) in the rat. Prednisolone was either added to the perfusion solution for lung preservation or omitted and rats were followed for 48 hours after LTX. Prednisolone preconditioning significantly increased survival and diminished reperfusion edema. Hypoxia induced vasoactive cytokines such as VEGF were reduced. Markers of leukocyte invasiveness like matrix metalloprotease (MMP)-2, or common pro-inflammatory molecules like the CXCR4 receptor or the chemokine (C-C motif) ligand (CCL)-2 were downregulated by prednisolone. Neutrophil recruitment to the grafts was only increased in Perfadex treated lungs. Together with this, prednisolone treated animals displayed significantly reduced lung protein levels of neutrophil chemoattractants like CINC-1, CINC-2α/ß and LIX and upregulated tissue inhibitor of matrix metalloproteinase (TIMP)-1. Interestingly, lung macrophage invasion was increased in both, Perfadex and prednisolone treated grafts, as measured by MMP-12 or RM4. Markers of anti-inflammatory macrophage transdifferentiation like MRC-1, IL-13, IL-4 and CD163, significantly correlated with prednisolone treatment. These observations lead to the conclusion that prednisolone as an additive to the perfusion solution protects from hypoxia triggered danger signals already in the phase of ischemia and thus reduces graft edema in the phase of reperfusion. Additionally, prednisolone preconditioning might also lead to macrophage polarization as a beneficial long-term effect.

Systemic endotoxin activity correlates with clot formation: an observational study in patients with early systemic inflammation and sepsis.

INTRODUCTION: Inflammation and coagulation are closely linked, and both can be triggered by endotoxin. Thrombelastometry and impedance aggregometry are of diagnostic and predictive value in critically ill patients. In this observational study we investigated the correlation of endotoxin activity with thrombelasometric and aggregometric variables in patients with systemic inflammation. METHODS: Based on a daily screening on a tertiary academic surgical ICU, patients, as soon as they fulfilled two or more criteria for systemic inflammatory response syndrome (SIRS), were included. In whole blood we performed endotoxin activity (EA) assay, thrombelastometry (ROTEM®) and impendance aggregometry (Multiplate®). RESULTS: In total, 49 patients were included with a broad spread of EA levels of (median (minimum to maximum)) 0.27 (0.01 to 0.72), allowing expedient correlative analysis. Clot formation time (CFT) (263 s (60 to 1,438 s)) and clotting time (CT) (1,008 s (53 to 1,481 s)) showed a significant negative correlation with EA level (r = -0.38 (P < 0.005) and r = -0.29 (P < 0.05)). Positive correlations were found for alpha-angle (50° (17 to 78°), r = 0.40 (P < 0.005)) and maximum clot firmness (MCF) (55 mm (5/76), r = 0.27 (P < 0.05)). No significant correlations were found between Lysis Index at 60 minutes (LI60) and EA levels. There was no correlation between EA level and aggregometric values, or classical coagulation parameters. CONCLUSIONS: In patients with systemic inflammation, increasing endotoxin concentrations correlate with increased clot formation.

Cecum ligation and dissection: a novel modified mouse sepsis model.

BACKGROUND: The fact that many sepsis therapeutics failed to be translated into the human indicates that there is still a serious need to reassess our models of sepsis research. We aimed to develop a novel modified model of sepsis in the mouse, which simulates the clinical situation more accurately. MATERIALS AND METHODS: Sepsis was induced in C57Bl/6 mice by dissecting the cecum and placing the discontinued organ back into the abdomen (cecum ligation and dissection [CLD]). Septic animals were relaparotomized after 6 h, followed by peritoneal lavage, and antibiotic treatment. Results were compared with shams or the classic colon ligation and puncture (CLP) model. The postoperative lung impairment was assessed using neutrophil invasion as a surrogate. Proinflammatory cytokines were measured by either real-time polymerase chain reaction or Luminex technology, and liver damage was evaluated by aspartate transaminase and alanine transaminase measurements. RESULTS: In CLD animals with relaparotomy after 6 h, lung interleukin (IL) 6, monocyte chemoattractant protein (MCP)-1 messenger RNA levels, and neutrophil invasion were significantly increased. Liver messenger RNA expression in CLD animals was significantly upregulated for IL-6, tumor necrosis factor alpha, IL-10, and MCP-1 compared with sham and CLP animals. Significantly higher levels of alanine transaminase were observed in CLD animals. Finally, systemic inflammation as measured by plasma IL-6, tumor necrosis factor alpha, IL-1ß, IL-10, and MCP-1 was significantly increased in all CLD animals compared with shams, whereas CLP animals only showed an insignificant increase in the latter molecules. CONCLUSIONS: Our modifications to the classic CLP model significantly produced organ inflammation, liver damage, and a similar mortality compared with a clinical setting, with a reliable onset of sepsis.