Thromboprophylaxis with argatroban in critically ill patients with sepsis: a review.

During sepsis, an initial prothrombotic shift takes place, in which coagulatory acute-phase proteins are increased, while anticoagulatory factors and platelet count decrease. Further on, the fibrinolytic system becomes impaired, which contributes to disease severity. At a later stage in sepsis, coagulation factors may become depleted, and sepsis patients may shift into a hypo-coagulable state with an increased bleeding risk. During the pro-coagulatory shift, critically ill patients have an increased thrombosis risk that ranges from developing micro-thromboses that impair organ function to life-threatening thromboembolic events. Here, thrombin plays a key role in coagulation as well as in inflammation. For thromboprophylaxis, low molecular weight heparins (LMWH) and unfractionated heparins (UFHs) are recommended. Nevertheless, there are conditions such as heparin resistance or heparin-induced thrombocytopenia (HIT), wherein heparin becomes ineffective or even puts the patient at an increased prothrombotic risk. In these cases, argatroban, a direct thrombin inhibitor (DTI), might be a potential alternative anticoagulatory strategy. Yet, caution is advised with regard to dosing of argatroban especially in sepsis. Therefore, the starting dose of argatroban is recommended to be low and should be titrated to the targeted anticoagulation level and be closely monitored in the further course of treatment. The authors of this review recommend using DTIs such as argatroban as an alternative anticoagulant in critically ill patients suffering from sepsis or COVID-19 with suspected or confirmed HIT, HIT-like conditions, impaired fibrinolysis, in patients on extracorporeal circuits and patients with heparin resistance, when closely monitored.

A review of European guidelines for patient blood management with a particular emphasis on antifibrinolytic drug administration for cardiac surgery.

The concept of patient blood management (PBM) was introduced by the World Health Organization in 2011 and is defined as a "patient-focused, evidence-based and systematic approach for optimizing the management of patients and transfusion of blood products to ensure high quality and effective patient care". Patient blood management is a multimodal approach based on three pillars: optimization of blood mass, minimization of blood loss and optimization of patient tolerance to anaemia. Antifibrinolytics play a major role in cardiac surgery, where the risk of perioperative bleeding is high and affects a majority of patients, by effectively reducing bleeding, transfusions, re-operations, as well as their associated morbidity and mortality. They represent an essential part of the pharmacological arsenal of patient blood management. However, despite the trend towards high-level PBM practices, currently very few European countries have national PBM guidelines and these guidelines, taken as a whole, are heterogeneous in form and content. In particular, the use of antifibrinolytics in cardiac surgery is often not discussed in detail beyond general prophylactic use and any recommendations lack detail including choice of drug, dosing, and mode of administration. Thus, the implementation of PBM programs in Europe is still challenging. In 2021, the WHO published a new document highlighting the urgent need to close the gap in PBM awareness and implementation and announced their upcoming initiative to develop specific PBM implementation guidelines. This review aims first, to summarize the role played by fibrinolysis in haemostatic disorders; second, to give an overview of the current available guidelines in Europe detailing PBM implementation in cardiac surgery; and third, to analyse the place and use of antifibrinolytics in these guidelines.

A survey of patient blood management for patients undergoing cardiac surgery in nine European countries.

STUDY OBJECTIVE: To describe and compare patient blood management (PBM) practices in cardiac surgery in nine European countries and identify the main risk factors for bleeding or transfusion according to the surveyed centres. DESIGN: We set up an online survey to evaluate PBM practices in two clinical scenarios, risk factors for bleeding or transfusion, and previous experience with antifibrinolytics. SETTING: This survey was completed by European anesthesiologists in 2019. PATIENTS: No patients were included in the survey. INTERVENTION: None. MEASUREMENTS: We evaluated the degree of implementation of PBM practices in patients undergoing cardiac surgery. MAIN RESULTS: Ninety-eight of 177 responses (38%) were complete with variable response rates by country. In a non-emergent situation, no respondents would transfuse red cells preoperatively in an anaemic patient, while cell salvage (89%) and antifibrinolytics (82%) would almost always be used. Optimization of Hemoglobin level (36%) and use of off-pump techniques (34%), minimally invasive surgery (25%) and relatively recently-developed CPB technologies such as mini-bypass (32%) and autologous priming (38%), varied greatly across countries. In an emergent clinical situation, topical haemostatic agents would frequently be used (61%). Tranexamic acid (72%) and aprotinin (20%) were the main antifibrinolytics used, with method of administration and dose varying markedly across countries. Five factors were considered to increase risk of bleeding or transfusion by at least 90% of respondents: pre-operative anaemia, prior cardiac surgery, clopidogrel 5 days or less before surgery, use of other P2Y12 inhibitors at any point, and thrombocytopenia <100.109 platelets/mm3. CONCLUSION: PBM guidelines are not universally implemented in European cardiac surgery centres or countries, resulting in discrepancies in techniques and products used for a given clinical situation.

A comparative study on adequate anesthesia depth: clinical judgement and the Narcotrend® measurement.

PURPOSE: To compare the clinical judgement of electroencephalogram (EEG)-naïve anesthesiologists with an EEG-based measurement of anesthetic depth (AD) using the Narcotrend® monitor. METHODS: In this prospective cohort study including 600 patients, AD during stable anesthesia was assessed by clinical judgement of the attending, EEG-blinded anesthesiologist (using a scale staging the AD as mid-adequate, adequate but fairly deep, or adequate but fairly light) and by simultaneously recorded Narcotrend measurements. RESULTS: In 42% of patients (n = 250), the anesthesiologist's clinical judgement was in agreement with anesthetic levels as measured by the Narcotrend monitor. In 46% of patients (n = 274), the anesthesiologist's judgement and the Narcotrend monitor differed by one AD level (minor discordance). Major discordance was observed in 76 (13%) measurements (judged deeper than measured, n = 29 [5%]; judged lighter than measured, n = 47 [8%]). In 7% of patients (n = 44), the Narcotrend index was outside the limits of adequate AD (too deep, n = 28 [5%]; too superficial, n = 16 [3%]). The overall level of agreement between the anesthesiologist's judgement and the Narcotrend monitor was not statistically significant (Cohen's kappa, -0.039; P = 0.17). Using a random forests algorithm, age, mean blood pressure, the American Society of Anesthesiologists classification, body mass index, and frailty were the variables with the highest relative feature importance to predict the level of agreement. CONCLUSION: These results suggest that clinical judgement of AD during stable anesthesia was not in agreement with EEG-based assessment of anesthetic depth in 58% of cases. Nevertheless, this finding could be influenced by the lack of validated scales to clinically judge AD. TRIAL REGISTRATION: www.clinicaltrials.gov (NCT02766894); registered 10 May, 2016.

RéSUMé: OBJECTIF: Notre objectif était de comparer le jugement clinique d'anesthésiologistes n'ayant pas accès à un électroencéphalogramme (EEG) à une mesure de la profondeur anesthésique (PA) fondée sur l'EEG utilisant le moniteur Narcotrend®. MéTHODE: Dans cette étude de cohorte prospective de 600 patients, la PA a été évaluée pendant la phase de maintien stable de l'anesthésie selon le jugement clinique de l'anesthésiologiste traitant, qui n'avait pas accès à l'EEG (sur une échelle évaluant la PA comme étant adéquate, adéquate mais relativement profonde ou adéquate mais relativement légère) et par des mesures simultanément enregistrées par le Narcotrend. RéSULTATS: Chez 42 % des patients (n = 250), le jugement clinique de l'anesthésiologiste concordait aux niveaux anesthésiques tels que mesurés par le moniteur Narcotrend. Chez 46 % des patients (n = 274), le jugement de l'anesthésiologiste et le moniteur Narcotrend différaient d'un niveau de PA (discordance mineure). Une discordance majeure a été observée dans 76 (13 %) mesures (jugées plus profondes que mesurées, n = 29 [5 %], jugées plus légères que mesurées, n = 47 [8 %]). Chez 7 % des patients (n = 44), l'indice Narcotrend était situé au-delà des limites d'une PA adéquate (trop profond, n = 28 [5 %]; trop superficiel, n = 16 [3 %]). Le niveau global de concordance entre le jugement de l'anesthésiologiste et le moniteur Narcotrend n'était pas significatif d'un point de vue statistique (kappa de Cohen, -0,039; P = 0,17). En se fondant sur un algorithme de forêt d'arbres décisionnels (random forests algorithm), l'âge, la tension artérielle moyenne, la classification selon l'American Society of Anesthesiologists, l'indice de masse corporelle et l'index de fragilité ont été identifiés comme les variables ayant la plus grande importance relative pour prédire le niveau de concordance. CONCLUSION: Ces résultats suggèrent que, dans 58 % des cas, le jugement clinique de la PA ne concordait pas à l'évaluation par EEG de la profondeur anesthésique pendant une phase de maintien stable de l'anesthésie. Toutefois, ces résultats pourraient être influencés par l'absence d'échelles validées pour juger la PA d'un point de vue clinique. ENREGISTREMENT DE L'éTUDE: www.clinicaltrials.gov (NCT02766894); enregistrée le 10 mai 2016.

miR-19a-3p containing exosomes improve function of ischaemic myocardium upon shock wave therapy.

AIMS: As many current approaches for heart regeneration exert unfavourable side effects, the induction of endogenous repair mechanisms in ischaemic heart disease is of particular interest. Recently, exosomes carrying angiogenic miRNAs have been described to improve heart function. However, it remains challenging to stimulate specific release of reparative exosomes in ischaemic myocardium. In the present study, we sought to test the hypothesis that the physical stimulus of shock wave therapy (SWT) causes the release of exosomes. We aimed to substantiate the pro-angiogenic impact of the released factors, to identify the nature of their cargo, and to test their efficacy in vivo supporting regeneration and recovery after myocardial ischaemia. METHODS AND RESULTS: Mechanical stimulation of ischaemic muscle via SWT caused extracellular vesicle (EV) release from endothelial cells both in vitro and in vivo. Characterization of EVs via electron microscopy, nanoparticle tracking analysis and flow cytometry revealed specific exosome morphology and size with the presence of exosome markers CD9, CD81, and CD63. Exosomes exhibited angiogenic properties activating protein kinase b (Akt) and extracellular-signal regulated kinase (ERK) resulting in enhanced endothelial tube formation and proliferation. A miRNA array and transcriptome analysis via next-generation sequencing were performed to specify exosome content. miR-19a-3p was identified as responsible cargo, antimir-19a-3p antagonized angiogenic exosome effects. Exosomes and target miRNA were injected intramyocardially in mice after left anterior descending artery ligation. Exosomes resulted in improved vascularization, decreased myocardial fibrosis, and increased left ventricular ejection fraction as shown by transthoracic echocardiography. CONCLUSION: The mechanical stimulus of SWT causes release of angiogenic exosomes. miR-19a-3p is the vesicular cargo responsible for the observed effects. Released exosomes induce angiogenesis, decrease myocardial fibrosis, and improve left ventricular function after myocardial ischaemia. Exosome release via SWT could develop an innovative approach for the regeneration of ischaemic myocardium.

Shock Wave Therapy Improves Cardiac Function in a Model of Chronic Ischemic Heart Failure: Evidence for a Mechanism Involving VEGF Signaling and the Extracellular Matrix.

Background Mechanical stimulation of acute ischemic myocardium by shock wave therapy ( SWT ) is known to improve cardiac function by induction of angiogenesis. However, SWT in chronic heart failure is poorly understood. We aimed to study whether mechanical stimulation upon SWT improves heart function in chronic ischemic heart failure by induction of angiogenesis and postnatal vasculogenesis and to dissect underlying mechanisms. Methods and Results SWT was applied in a mouse model of chronic myocardial ischemia. To study effects of SWT on postnatal vasculogenesis, wild-type mice received bone marrow transplantation from green fluorescence protein donor mice. Underlying mechanisms were elucidated in vitro in endothelial cells and murine aortic rings. Echocardiography and pressure/volume measurements revealed improved left ventricular ejection fraction, myocardial contractility, and diastolic function and decreased myocardial fibrosis after treatment. Concomitantly, numbers of capillaries and arterioles were increased. SWT resulted in enhanced expression of the chemoattractant stromal cell-derived factor 1 in ischemic myocardium and serum. Treatment induced recruitment of bone marrow-derived endothelial cells to the site of injury. In vitro, SWT resulted in endothelial cell proliferation, enhanced survival, and capillary sprouting. The effects were vascular endothelial growth factor receptor 2 and heparan sulfate proteoglycan dependent. Conclusions SWT positively affects heart function in chronic ischemic heart failure by induction of angiogenesis and postnatal vasculogenesis. SWT upregulated pivotal angiogenic and vasculogenic factors in the myocardium in vivo and induced proliferative and anti-apoptotic effects on endothelial cells in vitro. Mechanistically, these effects depend on vascular endothelial growth factor signaling and heparan sulfate proteoglycans. SWT is a promising treatment option for regeneration of ischemic myocardium.

Valve-sparing root replacement for freestanding pulmonary autograft aneurysm after the Ross procedure.

OBJECTIVE: Autograft dilatation is the main long-term complication following the Ross procedure using the freestanding root replacement technique. We reviewed our 25-year experience with the Ross procedure with a special emphasis on valve-sparing reoperations. METHODS: From 1991 to 2016, 153 patients (29.6 ± 16.6 years; 29.4% pediatric) underwent a Ross operation at our institution with implantation of the autograft as freestanding root replacement. The follow-up is 98.7% complete with a mean of 12.2 ± 5.5 years. RESULTS: Mortality at 30-days was 2.0%. Echocardiography documented no or trivial aortic regurgitation in 99.3% of the patients at discharge. Survival probability at 20 years was 85.4%. No case of autograft endocarditis occurred. Autograft deterioration rate was 2.01% per patient-year, and freedom from autograft reoperation was 75.3% at 15 years. A reoperation for autograft aneurysm was required in 35 patients (22.9%) at a mean interval of 11.1 ± 4.6 years after the Ross procedure. A valve-sparing root replacement was performed in 77% of patients, including 10 David and 17 Yacoub procedures with no early mortality. Three patients required prosthetic valve replacement within 2 years after a Yacoub operation. At latest follow-up, 92% of all surviving patients still carry the pulmonary autograft valve. Freedom from autograft valve replacement was 92.1% at 15 years. CONCLUSIONS: Using the David or Yacoub techniques, the autograft valve can be preserved in the majority of patients with root aneurysms after the Ross procedure. Reoperations can be performed with no early mortality, a good functional midterm result, and an acceptable reintervention rate.

Shock wave treatment after hindlimb ischaemia results in increased perfusion and M2 macrophage presence.

Shock wave therapy (SWT) has been shown to induce angiogenesis in ischaemic muscle. However, the mechanism of action remains unknown. Macrophages are crucial for angiogenic responses after ischaemic injury. The M2 macrophage subset enables tissue repair and induces angiogenesis. It was hypothesized that the angiogenic effects of SWT are at least partly caused by enhanced macrophage recruitment. C57BL/6 mice were subjected to hind limb ischaemia with subsequent SWT or sham treatment. Muscles were analysed via immunofluorescence staining, reverse-transcription polymerase chain reaction and western blot. Gene expression and proteins involved in macrophage recruitment were analysed and tissue sections were stained for macrophages, including subsets, capillaries and arterioles. Laser Doppler perfusion imaging was performed to assess functional outcome. Treated muscles showed increased expression of the pivotal macrophage recruiting factor monocyte chemotactic protein 1 (MCP-1). Higher levels of macrophage marker CD14 were found. Increased numbers of macrophages after SWT could be confirmed by immunofluorescence staining. The expression of the M2 polarization promoting chemokine interleukin 13 was significantly elevated in the treatment group. Elevated mRNA expression of the M2 scavenger receptor CD163 was found after SWT. Immunofluorescence staining confirmed increased numbers of M2 macrophages after treatment. It was found that SWT resulted in higher number of capillaries and arterioles. Assessment of functional outcome revealed significantly improved limb perfusion in treated animals. Shock wave therapy causes increased macrophage recruitment and enhanced polarization towards reparative M2 macrophages in ischaemic muscle resulting in angiogenesis and improved limb perfusion and therefore represents a promising new treatment option for the treatment of ischaemic heart disease. Copyright © 2016 John Wiley & Sons, Ltd.

Long-term Evaluation of the Ross Procedure in Acute Infective Endocarditis.

Optimal valve substitute for young patients with aortic valve endocarditis remains controversial. Given its better resistance to infection, the Ross procedure is an attractive alternative to prosthetic valve replacement or homograft implantation. The objective of this study was to assess long-term outcomes of the Ross procedure in this indication. From January 1991 to April 2017, 190 patients underwent a Ross procedure at our institution. Acute endocarditis was the indication for operation in 19 patients, including 6 patients with a bicuspid aortic valve. The pulmonary autograft was implanted as freestanding root replacement in all patients. The clinical follow-up is 100% complete, with a mean of 12.0 ± 5.7 years. The mean age of the study population was 35.9 ± 11.5 years. Moderate or severe aortic regurgitation was present in 84.2% of the patients. Systemic embolization had occurred in 36.8% of the patients. The mean aortic cross-clamp time was 126 ± 24 minutes. The median length of stay on the intensive care unit was 1 day. Mortality at 30 days was 5.3% (1 patient with gastrointestinal bleeding). Echocardiography at hospital discharge documented no or trivial aortic regurgitation in all patients. No case of recurrent endocarditis affecting the autograft occurred. One patient (0.4% per patient-year) was reoperated 1.8 years after the Ross procedure for homograft endocarditis. Three patients (15.8%) were reoperated for autograft aneurysm. The Ross procedure is a safe and effective alternative to prosthetic valve replacement or homograft implantation in selected young patients with acute endocarditis with a low rate of recurrent infection.

Risk of long-term anticoagulation under sustained severe arterial hypertension: A translational study comparing warfarin and the new oral anticoagulant apixaban.

New oral anticoagulants for the prevention of stroke and systemic embolism in patients with atrial fibrillation have recently been introduced. In this translational study, we explored the risk of long-term anticoagulation on intracerebral hemorrhage under sustained severe arterial hypertension. We initiated anticoagulation with warfarin or apixaban in spontaneously hypertensive rats prone to develop severe hypertension and subsequent intracerebral bleeding complications. A non-anticoagulated group served as control. During an 11-week-study period, blood pressure, anticoagulation parameters, and clinical status were determined regularly. The incidence of histopathologically proven intracerebral hemorrhage was defined as the primary endpoint. Both warfarin and apixaban anticoagulation was fairly stable during the study period, and all rats developed severe hypertension. Intracerebral hemorrhage was determined in 29% (4/14) of warfarin rats and in 10% (1/10) of apixaban rats. Controls did not show cerebral bleeding complications (chi-square not significant). Mortality rate at study termination was 33% (2/6) in controls, 43% (6/14) in the warfarin group, and 60% (6/10) in the apixaban group. Animals died from extracerebral complications in most cases. Our study describes an experimental intracerebral hemorrhage model in the context of sustained hypertension and long-term anticoagulation. Extracerebral bleeding complications occurred more often in warfarin-treated animals compared with apixaban and control rats.

Frizzled2 signaling regulates growth of high-risk neuroblastomas by interfering with ß-catenin-dependent and ß-catenin-independent signaling pathways.

Frizzled2 (FZD2) is a receptor for Wnts and may activate both canonical and non-canonical Wnt signaling pathways in cancer. However, no studies have reported an association between FZD2 signaling and high-risk NB so far. Here we report that FZD2 signaling pathways are critical to NB growth in MYCN-single copy SK-N-AS and MYCN-amplified SK-N-DZ high-risk NB cells. We demonstrate that stimulation of FZD2 by Wnt3a and Wnt5a regulates ß-catenin-dependent and -independent Wnt signaling factors. FZD2 blockade suppressed ß-catenin-dependent signaling activity and increased phosphorylation of PKC, AKT and ERK in vitro, consistent with upregulation of ß-catenin-independent signaling activity. Finally, FZD2 small interfering RNA knockdown suppressed tumor growth in murine NB xenograft models associated with suppressed ß-catenin-dependent signaling and a less vascularized phenotype in both NB xenografts. Together, our study suggests a role for FZD2 in high-risk NB cell growth and provides a potential candidate for therapeutic inhibition in FZD2-expressing NB patients.

Expression of the anti-inflammatory suppressor of cytokine signaling 3 (SOCS3) in human clear cell renal cell carcinoma.

The oncogenic transcription factor signal transducer and activator of transcription 3 (STAT3) is a cytokine-activated transcription factor controlling inflammation, cell proliferation, survival, and differentiation in normal tissue as well as in tumor growth. One of its most important negative regulators is the suppressor of cytokine signaling 3 (SOCS3). Here, we analyzed SOCS3 and other tumor-associated local immune regulators in human clear cell renal cell carcinoma (ccRCC). Analyses were performed in tumor and adjacent tumor-free healthy renal tissue from 35 patients with ccRCC. For functional analysis, ccRCC Caki-1 cell lines were stimulated with IL-6 and IFNγ in cell culture assays. We observed significantly lower SOCS3 messenger RNA (mRNA) levels in tumor tissue compared to healthy tissue. SOCS3 mRNA strongly correlated within tumor and healthy tissue. Interestingly vice versa, SOCS3 protein levels were significantly higher in tumor tissue than in healthy tissue. IL-22 and IL-22R1 mRNA displayed no differences in tumor and healthy tissue. Stimulation of Caki-1 cells with IFNγ resulted in markedly increased SOCS3 mRNA levels. We conclude that SOCS3 along with STAT3 participates in regulatory mechanisms in ccRCC, which certainly features only one of multiple factors involved but nevertheless merits further attention.

Epicardial shock-wave therapy improves ventricular function in a porcine model of ischaemic heart disease.

Previously we have shown that epicardial shock-wave therapy improves left ventricular ejection fraction (LVEF) in a rat model of myocardial infarction. In the present experiments we aimed to address the safety and efficacy of epicardial shock-wave therapy in a preclinical large animal model and to further evaluate mechanisms of action of this novel therapy. Four weeks after left anterior descending (LAD) artery ligation in pigs, the animals underwent re-thoracotomy with (shock-wave group, n = 6) or without (control group, n = 5) epicardial shock waves (300 impulses at 0.38 mJ/mm2 ) applied to the infarcted anterior wall. Efficacy endpoints were improvement of LVEF and induction of angiogenesis 6 weeks after shock-wave therapy. Safety endpoints were haemodynamic stability during treatment and myocardial damage. Four weeks after LAD ligation, LVEF decreased in both the shock-wave (43 ± 3%, p < 0.001) and control (41 ± 4%, p = 0.012) groups. LVEF markedly improved in shock-wave animals 6 weeks after treatment (62 ± 9%, p = 0.006); no improvement was observed in controls (41 ± 4%, p = 0.36), yielding a significant difference. Quantitative histology revealed significant angiogenesis 6 weeks after treatment (controls 2 ± 0.4 arterioles/high-power field vs treatment group 9 ± 3; p = 0.004). No acute or chronic adverse effects were observed. As a potential mechanism of action in vitro experiments showed stimulation of VEGF receptors after shock-wave treatment in human coronary artery endothelial cells. Epicardial shock-wave treatment in a large animal model of ischaemic heart failure exerted a positive effect on LVEF improvement and did not show any adverse effects. Angiogenesis was induced by stimulation of VEGF receptors. Copyright © 2014 John Wiley & Sons, Ltd.

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.

Regulation of tumorigenic Wnt signaling by cyclooxygenase-2, 5-lipoxygenase and their pharmacological inhibitors: A basis for novel drugs targeting cancer cells?

Canonical Wnt signaling is a highly conserved pathway with a prominent role in embryogenic development, adult tissue homeostasis, cell polarization, stem cell biology, cell differentiation, and proliferation. Furthermore, canonical Wnt signaling is of pivotal importance in the pathogenesis of a number of cancer types and crucially affects tumor initiation, cancer cell proliferation, cancer cell apoptosis, and metastasis. Reports over the last decade have provided strong evidence for a pathophysiological role of Wnt signaling in non-malignant classical inflammatory and neurodegenerative diseases. Although, several agents suppressing the Wnt pathway at different levels have been identified, the development of clinically relevant Wnt-inhibiting agents remains challenging due to selectivity and toxicity issues. Several studies have shown that long-term administration of non-steroidal anti-inflammatory drugs protects against colon cancer and potentially other tumor types by interfering both with the COX and the Wnt pathway. Our own studies have shown that non-steroidal anti-inflammatory drugs suppress Wnt signaling by targeting the pro-inflammatory enzyme 5-lipoxygenase which is the key enzyme pathophysiologically involved in the synthesis of leukotrienes. Furthermore, we found a direct link between the 5-lipoxygenase and Wnt signaling pathways, which is essential for the maintenance of leukemic stem cells. Accordingly, genetic and pharmacological inhibition of 5-lipoxygenase led to an impairment of Wnt-dependent acute and chronic myeloid leukemic stem cells. We believe that 5-lipoxygenase inhibitors might represent a novel type of Wnt inhibitor activating a potentially naturally occurring novel mechanism of suppression of Wnt signaling that is non-toxic, at least in mice, and is potentially well tolerated in patients.

Shock Wave Treatment Protects From Neuronal Degeneration via a Toll-Like Receptor 3 Dependent Mechanism: Implications of a First-Ever Causal Treatment for Ischemic Spinal Cord Injury.

BACKGROUND: Paraplegia following spinal cord ischemia represents a devastating complication of both aortic surgery and endovascular aortic repair. Shock wave treatment was shown to induce angiogenesis and regeneration in ischemic tissue by modulation of early inflammatory response via Toll-like receptor (TLR) 3 signaling. In preclinical and clinical studies, shock wave treatment had a favorable effect on ischemic myocardium. We hypothesized that shock wave treatment also may have a beneficial effect on spinal cord ischemia. METHODS AND RESULTS: A spinal cord ischemia model in mice and spinal slice cultures ex vivo were performed. Treatment groups received immediate shock wave therapy, which resulted in decreased neuronal degeneration and improved motor function. In spinal slice cultures, the activation of TLR3 could be observed. Shock wave effects were abolished in spinal slice cultures from TLR3(-/-) mice, whereas the effect was still present in TLR4(-/-) mice. TLR4 protein was found to be downregulated parallel to TLR3 signaling. Shock wave-treated animals showed significantly better functional outcome and survival. The protective effect on neurons could be reproduced in human spinal slices. CONCLUSIONS: Shock wave treatment protects from neuronal degeneration via TLR3 signaling and subsequent TLR4 downregulation. Consequently, it represents a promising treatment option for the devastating complication of spinal cord ischemia after aortic repair.

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.

Effects of azithromycin and tanomastat on experimental bronchiolitis obliterans.

OBJECTIVE: Azithromycin has become a standard of care in therapy of bronchiolitis obliterans following lung transplantation. Matrix metalloprotease-9 broncho-alveolar lavage levels increase in airway neutrophilia and bronchiolitis obliterans. Interleukin-17 may play a role in lung allograft rejection, and interleukin-12 is downregulated in bronchiolitis obliterans. Whether these mechanisms can be targeted by azithromycin remains unclear. METHODS: Bronchiolitis obliterans was induced by transplantation of Fischer F344 rat left lungs to Wistar Kyoto rats. Allografts with azithromycin therapy from day 1 to 28 or 56 and mono- or combination therapy with the broad-spectrum matrix metalloprotease inhibitor tanomastat from day 1 to 56 were compared to control allografts and isografts. Graft histology was assessed, and tissue cytokine expression studied using Western blotting and immunofluorescence. RESULTS: The chronic airway rejection score in the azithromycin group did not change between 4 and 8 weeks after transplantation, whereas it significantly worsened in control allografts (P = .041). Azithromycin+tanomastat prevented complete allograft fibrosis, which occurred in 40% of control allografts. Azithromycin reduced interleukin-17 expression (P = .049) and the number of IL-17(+)/CD8(+) lymphocytes at 4 weeks, and active matrix metalloprotease-9 at 8 weeks (P = .017), and increased interleukin-12 expression (P = .025) at 8 weeks following transplantation versus control allografts. CONCLUSIONS: The expression of interleukin-17 and matrix metalloprotease-9 in bronchiolitis obliterans may be attenuated by azithromycin, and the decrease in interleukin-12 expression was prevented by azithromycin. Combination of azithromycin with a matrix metalloprotease inhibitor is worth studying further because it prevented complete allograft fibrosis in this study.

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.

Upper and lower urinary tract infections can be detected early but not be discriminated by urinary NGAL in adults.

PURPOSE: We investigated whether the recently established biomarkers of acute kidney injury, neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule-1 (KIM-1), may help to diagnose acute urinary tract infections (UTI) in adults and are able to distinguish between upper or lower localization. METHODS: NGAL levels were measured in blood and urine, and KIM-1 concentrations in urine of 97 subjects. We recruited age- and gender-matched groups of 30 patients with acute upper UTI and 29 patients with acute lower UTI as well as 38 healthy controls. NGAL and KIM-1 were determined by ELISA, serum and urine creatinine applying the Jaffé's method. RESULTS: Urinary NGAL (uNGAL) was significantly increased in patients with upper as well as with lower UTI compared with the healthy controls (P < 0.01; P < 0.05). Accordingly, uNGAL normalized on urinary creatinine (uNGAL/uCrea) was markedly higher in patients with lower UTI compared with the control group (P < 0.05), and uNGAL/uCrea levels were still raised in patients with upper UTI, though not reaching statistical significance (P = 0.07). However, neither uNGAL nor uNGAL/uCrea levels displayed significant differences between patients with upper or lower UTI (P = 0.75; P = 0.97). uKIM-1 levels were close to the detection limit and too low to reliably reveal differences between the three groups. CONCLUSIONS: While uKIM-1 seems not to serve as a helpful biomarker in this setting, increased levels of uNGAL indicate inflammatory processes in the urinary tract in adults. However, the determination of uNGAL levels does not allow to differentiate between upper and lower UTI.