The G protein-coupled receptor kinase 6 is associated with inflammation and pathological pain. Impairment of GRK6 expression was described in chronic inflammatory diseases such as rheumatoid ...arthritis and this was shown to be accompanied by an imbalance of downstream signaling pathways. Here, we discuss novel aspects of GRK6 interaction and its impact upon hyperalgesia and inflammatory processes. In this review, we compile important findings concerning GRK6 regulation for a better pathophysiological understanding of the intracellular interaction in the context of inflammation and show clinical implications-for example, the identification of possible therapy goals in the treatment of chronic inflammatory hyperalgesia.
Sepsis is now operationally defined as life-threatening organ dysfunction caused by an infection, identified by an acute change in SOFA-Score of at least two points, including clinical chemistry such ...as creatinine or bilirubin concentrations. However, little knowledge exists about organ-specific microRNAs as potentially new biomarkers. Accordingly, we tested the hypotheses that micro-RNA-122, the foremost liver-related micro-RNA (miR), 1) discriminates between sepsis and infection, 2) is an early predictor for mortality, and 3) improves the prognostic value of the SOFA-score.
We analyzed 108 patients with sepsis (infection + increase SOFA-Score ≥2) within the first 24h of ICU admission and as controls 20 patients with infections without sepsis (infection + SOFA-Score ≤1). Total circulating miR was isolated from serum and relative miR-122 expression was measured (using spiked-in cel-miR-54) and associated with 30-day survival.
30-day survival of the sepsis patients was 63%. miR-122 expression was 40-fold higher in non-survivors (p = 0.001) and increased almost 6-fold in survivors (p = 0.013) compared to controls. miR-122 serum-expression discriminated both between sepsis vs. infection (AUC 0.760, sensitivity 58.3%, specificity 95%) and survivors vs. non-survivors (AUC 0.728, sensitivity 42.5%, specificity 94%). Multivariate Cox-regression analysis revealed miR-122 (HR 4.3; 95%-CI 2.0-8.9, p<0.001) as independent prognostic factor for 30-day mortality. Furthermore, the predictive value for 30-day mortality of the SOFA-Score (AUC 0.668) was improved by adding miR-122 (AUC 0.743; net reclassification improvement 0.37, p<0.001; integrated discrimination improvement 0.07, p = 0.007).
Increased miR-122 serum concentration supports the discrimination between infection and sepsis, is an early and independent risk factor for 30-day mortality, and improves the prognostic value of the SOFA-Score, suggesting a potential role for miR-122 in sepsis-related prediction models.
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The choice of the anesthetic regime is suggested to affect clinical outcomes following major surgery. Propofol was shown to exert beneficial effects on different cancer outcomes, ...while volatile anesthetics may be favorable in cardiac surgery. Recently, extracellular vesicles (EVs) were discovered as essential signal mediators in physiological and pathophysiological processes including carcinogenesis and metastasis. Furthermore, depending on their cell source, EVs fulfill therapeutic functions. In addition to extracorporally produced EVs, appropriate systemic intervention such as remote ischemic preconditioning (RIPC) is considered to promote endogenous release of therapeutically active EVs to mediate cardioprotective effects. EVs are assembled in cell-type specific manners and the composition of EVs is not only affected by the disease, but also by the applied anesthetic of anesthetized patients. Here, we compare known impacts of anesthetic agents on outcomes in cancer surgery and cardioprotection and link these effects to the composition and therapeutic potential of EVs.
Remote ischemic preconditioning (RIPC) can evoke cardioprotection following ischemia/reperfusion and this may depend on the anesthetic used. We tested whether 1) extracellular vesicles (EVs) isolated ...from humans undergoing RIPC protect cardiomyoblasts against hypoxia-induced apoptosis and 2) this effect is altered by cardiomyoblast exposure to isoflurane or propofol. EVs were isolated before and 60 min after RIPC or Sham from ten patients undergoing coronary artery bypass graft surgery with isoflurane anesthesia and quantified by Nanoparticle Tracking Analysis. Following EV-treatment for 6 hours under exposure of isoflurane or propofol, rat H9c2 cardiomyoblasts were cultured for 18 hours in normoxic or hypoxic atmospheres. Apoptosis was detected by flow cytometry. Serum nanoparticle concentrations in patients had increased sixty minutes after RIPC compared to Sham (2.5x1011±4.9x1010 nanoparticles/ml; Sham: 1.2x1011±2.0x1010; p = 0.04). Hypoxia increased apoptosis of H9c2 cells (hypoxia: 8.4%±0.6; normoxia: 2.5%±0.1; p<0.0001). RIPC-EVs decreased H9c2 cell apoptosis compared to control (apoptotic ratio: 0.83; p = 0.0429) while Sham-EVs showed no protection (apoptotic ratio: 0.97). Prior isoflurane exposure in vitro even increased protection (RIPC-EVs/control, apoptotic ratio: 0.79; p = 0.0035; Sham-EVs/control, apoptotic ratio:1.04) while propofol (50μM) abrogated protection by RIPC-EVs (RIPC-EVs/control, Apoptotic ratio: 1.01; Sham-EVs/control, apoptotic ratio: 0.94; p = 0.602). Thus, EVs isolated from patients undergoing RIPC under isoflurane anesthesia protect H9c2 cardiomyoblasts against hypoxia-evoked apoptosis and this effect is abrogated by propofol. This supports a role of human RIPC-generated EVs in cardioprotection and underlines propofol as a possible confounder in RIPC-signaling mediated by EVs.
Extracorporeal membrane oxygenation (ECMO) is a life-saving therapy in acute respiratory distress syndrome (ARDS) patients but is associated with complications and costs. Here, we validate various ...scores supposed to predict mortality and develop an optimized categorical model.
In a derivation cohort, 108 ARDS patients (2010-2015) on veno-venous ECMO were retrospectively analysed to assess four established risk scores (ECMOnet-Score, RESP-Score, PRESERVE-Score, Roch-Score) for mortality prediction (receiver operating characteristic analysis) and to identify by multivariable logistic regression analysis independent variables for mortality to yield the new PRESET-Score (PREdiction of Survival on ECMO Therapy-Score). This new score was then validated both in independent internal (n = 82) and external (n = 59) cohorts.
The median (25%; 75% quartile) Sequential Organ Failure Assessment score was 14 (12; 16), Simplified Acute Physiology Score II was 62.5 (57; 72.8), median intensive care unit stay was 17 days (range 1-124), and mortality was 62%. Only the ECMOnet-Score (area under curve (AUC) 0.69) and the RESP-Score (AUC 0.64) discriminated survivors and non-survivors. Admission pH
, mean arterial pressure, lactate, platelet concentrations, and pre-ECMO hospital stay were independent predictors of death and were used to build the PRESET-Score. The score's internal (AUC 0.845; 95% CI 0.76-0.93; p < 0.001) and external (AUC 0.70; 95% CI 0.56-0.84; p = 0.008) validation revealed excellent discrimination.
While our data confirm that both the ECMOnet-Score and the RESP-Score predict mortality in ECMO-treated ARDS patients, we propose a novel model also incorporating extrapulmonary variables, the PRESET-Score. This score predicts mortality much better than previous scores and therefore is a more precise choice for decision support in ARDS patients to be placed on ECMO.
G protein-coupled receptor kinase 6 (GRK6) is part of the G protein-coupled receptor kinase family, whose members act as key regulators of seven-transmembrane receptor signalling. GRK6 seems to play ...a role in regulation of inflammatory processes, but mechanisms of transcriptional regulation of GRK6 expression in inflammatory cell lines have not been characterized. Protein kinase C (PKC) signalling is also involved in inflammatory regulation and an impact of PKC activation on GRK6 protein expression was described previously. Thus, the aim of this study was to 1) characterize the GRK6 promoter, and 2) investigate a potential influence of PKC on GRK6 expression.
Five deletion constructs of the GRK6 promoter were cloned. After transient transfection into a human T cell line, promoter activity was assessed using luciferase reporter gene assays. Putative transcription factor binding sites were identified, mutated, and binding was investigated using electrophoretic mobility shift assays (EMSA). Following stimulation with a PKC activator, GRK6 expression on mRNA and protein levels was assessed by reverse transcriptase qPCR and Western blots.
Investigation of the GRK6 promoter revealed a putative cAMP responsive element (CRE), whose mutation led to decreased promoter activity (p = 0.0006). Functionality of the CRE binding protein (CREB) binding site was verified in EMSA blots. Stimulation with a PKC activator resulted in decreased GRK6 promoter activity (p = 0.0027), mRNA (p = 0.04) and protein expression.
We characterized the human GRK6 promoter and identified promoter activity to be influenced by a CREB binding site. PKC might be one determinant contributing to altered GRK6 expression.
Postoperative nausea and vomiting (PONV) is the most frequent side effect following anaesthesia. Predisposition to developing PONV is multifactorial with patient risk factors and anaesthetic ...techniques both being contributory. However, there is also a genetic susceptibility to PONV, and several studies have aimed to identify polymorphisms contributing to a genetic PONV risk.
We summarised previous published studies investigating genetic contribution to PONV risk.
Systematic review without meta-analysis.
We searched MEDLINE until June 2019.
Articles were chosen for review when PONV and polymorphisms were included. Exclusion criteria were reviews/meta-analysis/comments, articles not in the English language, nonappropriate content (e.g. PONV not as primary aim of the study, study investigated opioid-induced nausea) or if articles were pharmacogenetic studies addressing treatment of PONV.
A total of 59 studies were screened and 14 articles were reviewed including one genome-wide association study (GWAS). Seven studies were performed in East Asians, and seven in Caucasians. Seventeen polymorphisms have been positively associated with PONV in at least one study. Allele frequency of the investigated polymorphisms differs widely between the ethnicities. Furthermore, the anaesthesia regimen and the postoperative time point at which the association with PONV was reported were quite different. Only two polymorphisms, the CHRM3 rs2165870 and the KCNB2 rs349358 (both first associated with PONV in a GWAS), have been significantly associated with PONV incidence in Caucasians in independent studies.
There is a genetic susceptibility to the development of PONV. Two single nucleotide polymorphisms (SNPs), the CHRM3 rs2165870 and the KCNB2 rs349358 SNP, seem to have a major influence on PONV incidence, at least in Caucasians. Both SNPs were primarily identified in a GWAS and this association may lead to a better understanding of the disease aetiology. Further high-quality studies are needed to reveal more insights in genetic PONV susceptibility, particularly so in non-Caucasian ethnicities.
Echocardiographic quantification of ejection fraction (EF) by manual endocardial tracing requires training, is time-consuming and potentially user-dependent, whereas determination of cardiac output ...by pulmonary artery catheterization (PAC) is invasive and carries a risk of complications. Recently, a novel software for semi-automated EF and CO assessment (AutoEF) using transthoracic echocardiography (TTE) has been introduced. We hypothesized that AutoEF would provide EF values different from those obtained by the modified Simpson's method in transoesophageal echocardiography (TOE) and that AutoEF CO measurements would not agree with those obtained via VTI
in TOE and by thermodilution using PAC.
In 167 patients undergoing coronary artery bypass graft surgery (CABG), TTE cine loops of apical 4- and 2-chamber views were recorded after anaesthesia induction under steady-state conditions. Subsequently, TOE was performed following a standardized protocol, and CO was determined by thermodilution. EF and CO were assessed by TTE AutoEF as well as TOE, using the modified Simpson's method, and Doppler measurements via velocity time integral in the LV outflow tract (VTI
). We determined Pearson's correlation coefficients r and carried out Bland-Altman analyses. The primary endpoints were differences in EF and CO. The secondary endpoints were differences in left ventricular volumes at end diastole (LVEDV) and end systole (LVESV).
AutoEF and the modified Simpson's method in TOE showed moderate EF correlation (r = 0.38, p < 0.01) with a bias of -12.6% (95% limits of agreement (95%LOA): -36.6 - 11.3%). AutoEF CO correlated poorly both with VTI
in TOE (r = 0.19, p < 0.01) and thermodilution (r = 0.28, p < 0.01). The CO bias between AutoEF and VTI
was 1.33 l min
(95%LOA: -1.72 - 4.38 l min
) and 1.39 l min
(95%LOA -1.34 - 4.12 l min
) between AutoEF and thermodilution, respectively. AutoEF yielded both significantly lower EF (EF
: 42.0% (IQR 29.0 - 55.0%) vs. EF
: 55.2% (IQR 40.1 - 70.3%), p < 0.01) and CO values than the reference methods (CO
: 2.30 l min
(IQR 1.30 - 3.30 l min
) vs. CO
: 3.64 l min
(IQR 2.05 - 5.23 l min
) and CO
: 3.90 l min
(IQR 2.30 - 5.50 l min
), p < 0.01)).
AutoEF correlated moderately with TOE EF determined by the modified Simpson's method but poorly both with VTI
and thermodilution CO. A systematic bias was detected overestimating LV volumes and underestimating both EF and CO compared to the reference methods.
German Register for Clinical Trials (DRKS-ID DRKS00010666, date of registration: 08/07/2016).
An increasing amount of longitudinal health data is available on critically ill septic patients in the age of digital medicine, including daily sequential organ failure assessment (SOFA) score ...measurements. Thus, the assessment in sepsis focuses increasingly on the evaluation of the individual disease's trajectory. Machine learning (ML) algorithms may provide a promising approach here to improve the evaluation of daily SOFA score dynamics. We tested whether ML algorithms can outperform the conventional ΔSOFA score regarding the accuracy of 30-day mortality prediction.
We used the multicentric SepsisDataNet.NRW study cohort that prospectively enrolled 252 sepsis patients between 03/2018 and 09/2019 for training ML algorithms, i.e. support vector machine (SVM) with polynomial kernel and artificial neural network (aNN). We used the Amsterdam UMC database covering 1,790 sepsis patients for external and independent validation.
Both SVM (AUC 0.84; 95% CI: 0.71-0.96) and aNN (AUC 0.82; 95% CI: 0.69-0.95) assessing the SOFA scores of the first seven days led to a more accurate prognosis of 30-day mortality compared to the ΔSOFA score between day 1 and 7 (AUC 0.73; 95% CI: 0.65-0.80; p = 0.02 and p = 0.05, respectively). These differences were even more prominent the shorter the time interval considered. Using the SOFA scores of day 1 to 3 SVM (AUC 0.82; 95% CI: 0.68 0.95) and aNN (AUC 0.80; 95% CI: 0.660.93) led to a more accurate prognosis of 30-day mortality compared to the ΔSOFA score (AUC 0.66; 95% CI: 0.58-0.74; p < 0.01 and p < 0.01, respectively). Strikingly, all these findings could be confirmed in the independent external validation cohort.
The ML-based algorithms using daily SOFA scores markedly improved the accuracy of mortality compared to the conventional ΔSOFA score. Therefore, this approach could provide a promising and automated approach to assess the individual disease trajectory in sepsis. These findings reflect the potential of incorporating ML algorithms as robust and generalizable support tools on intensive care units.
Remote ischemic preconditioning (RIPC) by repeated brief cycles of limb ischemia/reperfusion may reduce myocardial ischemia/reperfusion injury and improve patients' prognosis after elective coronary ...artery bypass graft (CABG) surgery. The signal transducer and activator of transcription (STAT)5 activation in left ventricular myocardium is associated with RIPC´s cardioprotection. Cytokines and growth hormones typically activate STATs and could therefore act as humoral transfer factors of RIPC´s cardioprotection. We here determined arterial plasma concentrations of 25 different cytokines, growth hormones, and other factors which have previously been associated with cardioprotection, before (baseline)/after RIPC or placebo (n = 23/23), respectively, and before/after ischemic cardioplegic arrest in CABG patients. RIPC-induced protection was reflected by a 35% reduction of serum troponin I release. With the exception of interleukin-1α, none of the humoral factors changed in their concentrations after RIPC or placebo, respectively. Interleukin-1α, when normalized to baseline, increased after RIPC (280 ± 56%) but not with placebo (97 ± 15%). The interleukin-1α concentration remained increased until after ischemic cardioplegic arrest and was also higher than with placebo in absolute concentrations (25 ± 6 versus 16 ± 3 pg/mL). Only interleukin-1α possibly fulfills the criteria which would be expected from a substance to be released in response to RIPC and to protect the myocardium during ischemic cardioplegic arrest.
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