Defective RNA sensing by RIG-I in severe influenza virus infection.

Influenza virus infection causes worldwide seasonal epidemics. Although influenza is usually a mild disease, a minority of patients experience very severe fulminating disease courses. Previous studies have demonstrated a role for type I interferon (IFN) in anti-viral responses during influenza. So far, however, IFN regulatory factor (IRF)7 deficiency is the only genetic cause of severe influenza described in humans. In this study we present a patient with severe influenza A virus (IAV) H1N1 infection during the 2009 swine flu pandemic. By whole exome sequencing we identified two variants, p.R71H and p.P885S, located in the caspase activation and recruitment domain (CARD) and RNA binding domains, respectively, of DExD/H-box helicase 58 (DDX58) encoding the RNA sensor retinoic acid inducible gene 1 (RIG-I). These variants significantly impair the signalling activity of RIG-I. Similarly, patient cells demonstrate decreased antiviral responses to RIG-I ligands as well as increased proinflammatory responses to IAV, suggesting dysregulation of the innate immune response with increased immunopathology. We suggest that these RIG-I variants may have contributed to severe influenza in this patient and advocate that RIG-I variants should be sought in future studies of genetic factors influencing single-stranded RNA virus infections.

Ultrasound-guidance outperforms the palpation technique for peripheral venous catheterisation in anaesthetised toddlers: a randomised study.

BACKGROUND: Traditional palpation technique for peripheral venous catheter (PVC) placement can be challenging. We aimed to compare the Dynamic Needle Tip Positioning (DNTP) ultrasound technique with the palpation technique for PVC placement in anaesthetised toddlers undergoing elective, low-risk procedures. METHODS: The study was a randomised, controlled, crossover study. Five operators applied both techniques for PVC on 50 children < 4 years. The primary endpoint was first attempt success rate. The secondary endpoints were: (1) overall success rate, (2) number of skin perforations, (3) number of needle redirections, (4) number of catheters used, (5) total time, (6) needle manipulation time, (7) distance to the nearest flexion crease, (8) anatomical region of catheterisation, (9) size of the catheters and (10) the effects of visibility, and palpability of the veins. RESULTS: The first attempt success rate and the overall success rate were significantly higher in the DNTP group (42/50 vs. 30/50, P = 0.029), (50/50 vs. 42/50, P = 0.008). Furthermore, a lower number of skin perforations (60 vs. 84, P = 0.013) and needle redirections (14 vs. 131, P < 0.001) and increased distance to the nearest flexion crease (P < 0.001) were seen in the DNTP group. There were no significant differences in number of catheters used (60 vs. 75 P = 0.050) or total time (P = 0.073), however, the needle manipulation time was longer in the DNTP group (P = 0.011). The success rate decreased with less visible and palpable veins in the palpation group (p = 0.006). CONCLUSION: Ultrasound-guidance outperforms the traditional palpation technique for peripheral venous catheterisation in anaesthetised toddlers undergoing elective, low-risk procedures.

Extracorporeal cardiopulmonary resuscitation after out-of-hospital cardiac arrest in a Danish health region.

BACKGROUND: Extracorporeal Cardiopulmonary Resuscitation (ECPR) has emerged as a feasible rescue therapy for refractory, normothermic out-of-hospital cardiac arrest (OHCA). Reported survival rates vary and comparison between studies is hampered by heterogeneous study populations, differences in bystander intervention and in pre-hospital emergency service organisation. We aimed to describe the first experiences, treatment details, complications and outcome with ECPR for OHCA in a Danish health region. METHODS: Retrospective study of adult patients admitted at Aarhus University Hospital, Denmark between 1 January 2011 and 1 July 2015 with witnessed, refractory, normothermic OHCA treated with ECPR. OHCA was managed with pre-hospital advanced airway management and mechanical chest compression during transport. Relevant pre-hospital and in-hospital data were collected with special focus on low-flow time and ECPR duration. Survival to hospital discharge with Cerebral Performance Category (CPC) of 1 and 2 at hospital discharge was the primary endpoint. RESULTS: Twenty-one patients were included. Median pre-hospital low-flow time was 54 min [range 5-100] and median total low-flow time was 121 min [range 55-192]. Seven patients survived (33%). Survivors had a CPC score of 1 or 2 at hospital discharge. Five survivors had a shockable initial rhythm. In all survivors coronary occlusion was the presumed cause of cardiac arrest. CONCLUSION: Extracorporeal cardiopulmonary resuscitation is feasible as a rescue therapy in normothermic refractory OHCA in highly selected patients. Low-flow time was longer than previously reported. Survival with favourable neurological outcome is possible despite prolonged low-flow duration.

Severe hypoglycemia during methadone escalation in an 8-year-old child.

This case report describes hypoglycemia during methadone escalation in an 8-year-old girl with acute lymphatic leukemia. After a significant increase in the methadone dosage, the blood glucose decreased from 12 mmol/l to 1 mmol/l. Nutrition and other medications were meticulously checked and no other apparent explanation was found. There are few reports from the adult palliative setting which describe this serious side effect. This is to our knowledge, the first described pediatric case.

Cytokine response in peripheral blood indicates different pathophysiological mechanisms behind anastomotic leakage after low anterior resection: a pilot study.

BACKGROUND: Anastomotic leakage (AL) after rectosigmoid resection is a serious complication associated with high morbidity and mortality. This case-control pilot study investigated the changes in blood concentration of 10 different cytokines and 2 complement factors in relation to symptomatic AL after low anterior resection for rectosigmoid cancer. METHODS: Fifty patients scheduled for resection of rectosigmoid cancer had blood samples taken the day before surgery and on post-operative days 1, 3 and 5. Four patients with symptomatic AL were identified. Twenty-two age- and disease-matched patients constituted the control group. The concentration of 10 cytokines (granulocyte macrophage colony-stimulating factor, interferon-γ, interleukin-1ß, interleukin-2, interleukin-4, interleukin-5, interleukin-6, interleukin-8, interleukin-10 and tumour necrosis factor-α) and 2 complement factors (mannan-binding lectin and membrane attack complex) were measured. RESULTS: The present study demonstrated that plasma concentration of interleukin-1ß, interleukin-6, interleukin-8 and interleukin 10 within the first 5 post-operative days was increased in patients who developed early clinical AL, whereas there were no changes in patients with late-onset AL. CONCLUSIONS: The demonstrated differences in the cytokine response in early and late AL may support the theory of different pathological mechanisms of AL.

Metabolic effects of free fatty acids during endotoxaemia in a porcine model--free fatty acid inhibition of growth hormone secretion as a potential catabolic feedback mechanism.

Critical illness and severe inflammation are catabolic states characterised by breakdown of tissue and protein stores, by increased levels of free fatty acids, and by insulin resistance. These metabolic features contribute to morbidity and mortality. Growth hormone and insulin are the two major anabolic hormones. The present study was designed to test whether increased levels of free fatty acids (i) inhibit growth hormone secretion and (ii) induce insulin resistance during acute endotoxin exposure in a porcine model of critical illness. We studied 20 pigs for 6 h during combined anaesthesia and endotoxin infusion and a hyperinsulinaemic glucose clamp to control glucose, insulin, and free fatty acid concentrations. Pigs were randomised to two different continuous infusion rates of Intralipid resulting in different, sustained, and elevated free fatty acid concentrations (1.63 mmol l(-1) vs. 0.58 mmol l(-1), p=0.0002). Concomitantly, we observed reduced growth hormone concentrations in the group with high free fatty acid concentrations (3.5 ng ml(-1) vs. 6.6 ng ml(-1), p<0.003). No difference in insulin sensitivity, measured as the glucose infusion rate necessary to maintain euglycaemia, was observed. We conclude that high levels of free fatty acids reduce circulating growth hormone concentrations in porcine endotoxaemia; this probably constitutes a negative feedback mechanism whereby growth hormone induced-stimulation of free fatty acids release inhibit growth hormone secretion. This mechanism may further contribute to protein loss in critical illness. We found no evidence that the increment of plasma free fatty acids between groups contribute to insulin resistance in critical illness.

Dose-response effects of free fatty acids on glucose and lipid metabolism during somatostatin blockade of growth hormone and insulin in humans.

CONTEXT: GH and other stress hormones stimulate lipolysis, which may result in free fatty acid (FFA)-mediated insulin resistance. However, there are also indications that FFAs in the very low physiological range have the same effect. OBJECTIVE: The objective of the study was to address systematically the dose-response relations between FFAs and insulin sensitivity. DESIGN: We therefore examined eight healthy men for 8 h (6 h basal and 2 h glucose clamp) on four occasions. INTERVENTION: Intralipid was infused at varying rates (0, 3, 6, 12 microl.kg(-1).min(-1)); lipolysis was blocked by acipimox; and endogenous GH, insulin, and glucagon secretion was blocked by somatostatin and subsequently replaced at fixed rates. RESULTS: This resulted in four different FFA levels between 50 and 2000 micromol/liter, with comparable levels of insulin and counterregulatory hormones. Both in the basal state and during insulin stimulation, we saw progressively decreased glucose disposal, nonoxidative glucose disposal, and forearm muscle glucose uptake at FFA levels above 500 micromol/liter. Apart from forearm glucose uptake, the very same parameters were decreased at low FFA levels (approximately 50 micromol/liter). FFA rate of disposal was linearly related to the level of FFAs, whereas lipid oxidation reached a maximum at FFA levels approximately 1000 micromol/liter. CONCLUSION: In the presence of comparable levels of all major metabolic hormones, insulin sensitivity peaks at physiological levels of FFAs with a gradual decrease at elevated as well as suppressed FFA concentrations. These data constitute comprehensive dose-response curves for FFAs in the full physiological range from close to zero to above 2000 micromol/liter.

Coagulation, inflammatory, and stress responses in a randomized comparison of open and laparoscopic repair of recurrent inguinal hernia.

BACKGROUND: In previous comparisons of inflammatory and stress responses to open (OR) and laparoscopic (LR) hernia repair, all operations were performed under general anesthesia. Since local anesthesia is widely used for OR, a comparison of this approach with LR seemed relevant. METHODS: Patients with recurrent inguinal hernia were randomized to OR under local anesthesia (n = 30) or LR under general anesthesia (n = 31). The magnitude of the surgical trauma was assessed by measuring markers of coagulation (prothrombin fragment 1 + 2), endothelial activation (von Willebrand factor), inflammation [leukocytes, interleukin-6, -8 and -10, granulocyte macrophage colony-stimulating factor, and C-reactive protein (CRP)], and endocrine stress (cortisol) in blood collected before operation, 4 h postincision, and on postoperative day 2. RESULTS: Leukocyte counts and interleukin-6 and CRP levels increased in both groups, with the CRP increase being significantly greater in the OR group. The other markers did not increase significantly. CONCLUSION: The acute phase response was more pronounced after OR, even when this was done under local anesthesia. Both techniques seemed rather atraumatic.

Effects of adrenaline on lactate, glucose, lipid and protein metabolism in the placebo controlled bilaterally perfused human leg.

AIM: Adrenaline has widespread metabolic actions, including stimulation of lipolysis and induction of insulin resistance and hyperlactatemia. Systemic adrenaline administration, however, generates a very complex hormonal and metabolic scenario. No studies employing regional, placebo controlled and adrenaline infusion exist. Our study was designed to test the hypothesis that local placebo controlled leg perfusion with adrenaline directly increases local lactate release, stimulates lipolysis, induces insulin resistance and leaves protein metabolism unaffected. METHODS: We studied seven healthy volunteers with bilateral femoral vein and artery catheters during 3-h basal and 3-h hyperinsulinemic (0.6 mU kg(-1) min(-1) ) euglycemic clamp conditions. One femoral artery was perfused with saline and the other with adrenaline (0.4 µg min m(-2) ). Lipid metabolism was quantified with [9,10-(3) H] palmitate and amino acid metabolism with (15) N-phenylalanine and lactate and glucose by raw arterio-venous differences. RESULTS: Femoral vein plasma adrenaline increased ≈eightfold in the perfused leg with unaltered blood flows. Adrenaline perfusion significantly increased local leg lactate release from 0.01 to 0.25 mmol min(-1) per leg, palmitate release in the basal state 11.5-16.9 µmol min(-1) per leg and during the clamp 2.62-8.44 µmol min(-1) per leg. Glucose uptake decreased during the clamp from ≈180 to 30 µmol min(-1) per leg. Phenylalanine kinetics was not affected by adrenaline. CONCLUSION: Adrenaline directly increases lactate release and lipolysis and inhibits insulin-stimulated glucose uptake in the perfused human leg. Adrenaline has no direct effects on peripheral amino acid metabolism. Adrenaline-induced lactate release from striated muscle may be an important mechanism underlying hyperlactatemia in the critically ill.

Forearm and leg amino acid metabolism in the basal state and during combined insulin and amino acid stimulation after a 3-day fast.

AIM: Fasting is characterized by a progressive loss of protein, but data on protein kinetics are unclear and few have studied the effects of re-feeding. The present study was designed to test the hypothesis that a combined infusion of insulin and amino acids after fasting would induce compensatory increases in protein synthesis and reductions in protein breakdown at the whole body level and in muscle. METHODS: We included 10 healthy male volunteers and studied them twice: (1) in the post-absorptive state and (2) after 72 h of fasting. Amino acid kinetics was measured using labelled phenylalanine and tyrosine, whole body energy expenditure was assessed and urea nitrogen synthesis rates were calculated. RESULTS: After fasting we observed an increase in arterial blood concentration of branched chain amino acids and a decrease in gluconeogenic amino acids (P < 0.05). Isotopically determined whole body, forearm and leg phenylalanine fluxes were unaltered apart from a 30% decrease in phenylalanine-to-tyrosine conversion (2.0 vs. 1.4 mumol kg(-1) h(-1), P < 0.01). During infusion of insulin and amino acids, amino acid concentrations increased. CONCLUSION: Our data indicate that after a 72-h fast basal and insulin/amino acid-stimulated regional phenylalanine fluxes in leg and forearm muscle are unaltered. During fasting concentrations of gluconeogenic amino acids decrease and hepatic and/or renal phenylalanine-to-tyrosine conversion decreases. Thus, as opposed to glucose and lipid metabolism, fasting does not induce insulin resistance as regards amino acid metabolism.

Dose-response effects of free fatty acids on amino acid metabolism and ureagenesis.

AIM: Free fatty acids (FFAs) are important fuels and have vital protein-sparing effects, particularly during conditions of metabolic stress and fasting. However, it is uncertain whether these beneficial effects are evident throughout the physiological range or only occur at very high FFA concentrations. It is also unclear whether secondary alterations in hormone levels and ketogenesis play a role. We therefore aimed at describing dose-response relationships between amino acid metabolism and circulating FFA concentrations at clamped hormone levels. METHODS: Eight healthy men were studied on four occasions (6 h basal, 2 h glucose clamp). Endogenous lipolysis was blocked with acipimox and Intralipid was infused at varying rates (0, 3, 6 or 12 microL kg(-1) min(-1)) to obtain four different levels of circulating FFAs. Endogenous growth hormone, insulin and glucagon secretion was blocked by somatostatin (300 microg h(-1)) and replaced exogenously. 15N-phenylalanine, 2H4-tyrosine and 13C-urea were infused continuously to assess protein turnover and ureagenesis. RESULTS: We obtained four distinct levels of FFA concentrations ranging from 0.03 to 2.1 mmol L(-1) and 3-hydroxybutyrate concentrations from 10 to 360 micromol L(-1). Whole-body phenylalanine turnover and phenylalanine-to-tyrosine degradation decreased with increasing FFA levels as did insulin-stimulated forearm fluxes of phenylalanine. Phenylalanine, tyrosine and urea concentrations also decreased progressively, whereas urea turnover was unperturbed. CONCLUSION: Circulating FFAs decrease amino acid concentrations and inhibit whole-body phenylalanine fluxes and phenylalanine-to-tyrosine conversion. Our data cover FFA concentrations from 0 to 2 mmol L(-1) and indicate that FFAs exert their protein conserving effects in the upper physiological range (>1.5 mmol L(-1)).

Effects of a 3-day fast on regional lipid and glucose metabolism in human skeletal muscle and adipose tissue.

AIM: Fasting is characterized by increased whole body lipolysis and lipid oxidation, decreased glucose oxidation and insulin resistance. To identify the regional sources and underlying mechanisms, we studied 10 healthy male volunteers post-absorptively and after 72 h of fasting. METHODS: Each study comprised a 3-h basal period and a 3-h hyperinsulinaemic euglycaemic clamp and we used a combination of leg and forearm arteriovenous techniques, upper and lower body microdialysis and glucose and palmitate tracers. RESULTS: In the basal state, plasma levels, fluxes and oxidation rates of free fatty acids all roughly doubled after fasting. Palmitate fluxes across the forearm and leg also increased by two to threefold and interstitial leg muscle glycerol concentrations doubled. Subcutaneous femoral glycerol concentrations and blood flows were unaltered, but abdominal subcutaneous blood flow increased by 50% in the presence of unchanged glycerol concentrations, indicating stimulated abdominal lipolysis. During the clamp, we observed whole body insulin resistance and glucose uptake across the leg and forearm decreased by 60%. CONCLUSION: Our data show that fasting induces insulin resistance in upper and lower body muscles and suggest that increased lipolysis, is primarily due to the activation of lipolysis in muscle-associated fat (in the leg) and in upper body subcutaneous fat, whereas peripheral subcutaneous fat is spared.

Oxygenation and release of inflammatory mediators after off-pump compared with after on-pump coronary artery bypass surgery.

BACKGROUND: In a previous study, we showed that oxygenation was impaired for up to 5 day after conventional coronary artery bypass grafting (CABG). As cardiopulmonary bypass (CPB) may have a detrimental effect on pulmonary function, we hypothesized that coronary revascularization grafting without the use of CPB (OPCAB) would affect post-operative oxygenation and release of inflammatory mediators less compared with CABG. METHODS: Low-risk patients scheduled for elective coronary revascularization were randomly assigned to one of two groups (CABG, n = 17 or OPCAB, n = 18). Two parameters of oxygenation, shunt (%) and ventilation-perfusions mismatch, described as DeltaPO(2) (kPa), were estimated for up to 5 days post-operatively. Systemic release of interleukin (IL)-6, -8 and -10, C-reactive protein (CRP) and neutrophils were measured in peripheral blood samples for up to 3 days post-operatively. The lungs participation in the cytokine response was evaluated from mixed venous blood samples taken within the first 16 h post-operatively. RESULTS: OPCAB was followed by a higher shunt (P = 0.047), with no difference (P = 0.47) in the deterioration of DeltaPO(2) between the groups. OPCAB was followed by an attenuated systemic release of IL-8 (P = 0.041) and IL-10 (P = 0.006), while the release of IL-6 (P = 0.94), CRP (P = 0.121) and neutrophils (P = 0.078) did not differ between the groups. Indications of an uptake of cytokines in the lungs were found after OPCAB. CONCLUSIONS: When comparing OPCAB with CABG, oxygenation was more affected and only part of the systemic inflammatory response was attenuated.

Inflammatory response during hyperglycemia and hyperinsulinemia in a porcine endotoxemic model: the contribution of essential organs.

BACKGROUND: During euglycemia acute hyperinsulinemia diminishes the cytokine response to endotoxin [Lipopolysaccharide (LPS)] exposure. In this study we elucidated whether acute hyperglycemia and hyperinsulinemia modify the cytokine content in several organs during LPS challenge in a porcine model. METHODS: Pigs (35-40 kg) were randomized to either normoglycemia (group 1, n = 8) or hyperglycemia and hyperinsulinemia (group 2, n = 8), anesthetized and mechanically ventilated. Both groups received a 180-min intravenous infusion of LPS (total 10 microg kg(-1)). Groups 1 and 2 were clamped at plasma glucose concentrations of 5 mM and 15 mM, respectively. Group 1 maintained a baseline insulin level while the hyperglycemic group exhibited increased insulin levels. RESULTS: Circulating cytokines, cytokine mRNA and cytokine protein content were examined in the heart, liver, kidneys, lungs, spleen, adipose and muscle tissue. After LPS exposure, in both groups vast and equal plasma cytokines were elicited by approximately 70-5000-fold. A 10-fold higher level of IL-10, IL-6 and TNF-alpha protein was found in kidney tissue compared to the other organs together with a 3-10-fold increase of TNF-alpha in adipose tissue. However, cytokine mRNAs as well as organ function were without statistical difference between the groups. CONCLUSION: Endotoxemia elicited a pronounced cytokine response in both plasma and at organ level. The kidneys and adipose tissue showed the highest cytokine protein content. Acute hyperglycemia apparently counteracts the well-established anti-inflammatory effects of insulin on the inflammatory response in a LPS challenged porcine model. Whether the observation can be extrapolated to more long-term stress-exposure remains to be clarified.