Pain levels and patient comfort after lower limb arthroplasty comparing i.v. patient-controlled analgesia, continuous peripheral nerve block and neuraxial analgesia: a retrospective cohort analysis of clinical routine data.

BACKGROUND: Insufficient pain control after lower limb arthroplasty results in delayed recovery and increased risk for pain chronicization. The ideal kind of analgesia is still discussed controversially. We conducted a retrospective analysis of single-center routine data from a German university hospital, including patients receiving either total hip (THA) or knee arthroplasty (TKA). METHODS: All patients received general anesthesia. Patients undergoing THA received either continuous epidural ropivacaine infusion (0.133%, Epi) or patient-controlled analgesia (PCA) with the Wurzburg Pain Drip (tramadol, metamizole and droperidol, WPD) or with piritramide (Pir). After TKA, patients received either continuous femoral nerve block (ropivacaine 0.2%, PNB) or Pir. RESULTS: The analyzed cohort comprised 769 cases. Use of WPD after THA (n = 333) resulted in significantly reduced Numeric Rating Scale (NRS) values at rest, compared to Epi (n = 48) and Pir (n = 72) (.75 [IQR 1.14] vs. 1.17 [1.5], p = .02 vs. 1.47 [1.33], p < .0001) as well as maximum NRS scores (2.4 [1.7] vs. 3.29 [1.94], p < .001 vs. 3.32 [1.76], p < .0001). Positive feedback during follow-up visits was significantly increased in patients with a WPD PCA (p < .0001), while negative feedback (senso-motoric weakness/technical problems/nausea/dizziness/constipation) was particularly increased in Epi patients and lowest in those with WPD (p < .0001). After TKA, Pir (n = 131) resulted in significantly reduced NRS values at rest, compared to PNB (n = 185) (1.4 [1.4] vs. 1.6 [1.68], p = .02). Positive feedback was increased in patients with a Pir PCA in comparison with PNB (p = .04), while negative feedback was increased in PNB patients (p = .04). Overall, WPD presented with the lowest rate of any complications (8.7%), followed by Pir (20.2%), PNB (27.6%) and Epi (31.3%) (p < .001). CONCLUSIONS: In the assessed population, the use of a WPD PCA after THA offered better pain control and patient comfort in comparison with continuous epidural or piritramide-based analgesia. After TKA, the use of a Pir PCA provided superior analgesia and a lower complication rate compared to continuous PNB.

Comparison of the Newborn Infant Parasympathetic Evaluation (NIPE™) index to changes in heart rate to detect intraoperative nociceptive stimuli in healthy and critically ill children below 2 years: An observational study.

BACKGROUND: The validity of current tools for intraoperative objective assessment of nociception/antinociception balance during anesthesia in young and very young surgery children is unknown. AIM: Primary aim of the study was to test the hypothesis that the Newborn Infant Parasympathetic Evaluation (NIPE) index performs better in indicating nociception in anesthetized children below 2 years than changes in heart rate. Secondary aims were to evaluate associations between intraoperative changes in NIPE index values and postoperative pain and emergence delirium. METHODS: Fifty-one children aged <2 years who underwent surgery were included in this prospective observational study. Patients were assigned to either group 1 (healthy children, n = 31) or group 2 (critically ill, ventilated premature infants and neonates, n = 20). The NIPE index and heart rate in response to three defined nociceptive stimuli were continuously recorded. Two different scales, Kindliche Unbehagens- und Schmerzskala (KUS) and Pediatric Anesthesia Emergence Delirium (PAED) as well as a Pain Questionnaire were used to assess postoperative pain levels and emergence delirium. RESULTS: In total, 110 nociceptive events were evaluated. The analysis revealed a statistically significant association between a decrease in the NIPE index and all nociceptive stimuli, with a sensitivity of 92% and a specificity of 96%. The mean percentage decrease ranged from approx. 15%-30% and was highly statistically significant in both groups and for each of the nociceptive events except for venous puncture (p = .004). In contrast, no consistent change in heart rate was demonstrated. The KUS and PAED scale scores were significantly associated with the duration of anesthesia (p = .04), but not with intraoperative NIPE depression. CONCLUSION: The NIPE index was reliable for assessing intraoperative nociception in children aged <2 years and was more reproducible for detecting specific nociceptive stimuli during general anesthesia than heart rate. An effect on postoperative outcome is still elusive.

From bench to bar side: Evaluating the red wine storage lesion.

Vitally essential red fluids like packed cells and red wine are seriously influenced in quality when stored over prolonged periods. In the case of red cell concentrates, the resulting storage lesion has particular significance in perioperative medicine. We hypothesized that, in contrast, aging rather improves the properties of red wine in several ways. A translational approach, including (I) in vitro experiments, (II) a randomized, blinded crossover trial of acute clinical effects, and (III) a standardized red wine blind tasting was used. Three monovarietal wines (Cabernet Sauvignon, Chianti, Shiraz) in three different vintages (range 2004-2016), each 5 years different, were assessed. Assessments were performed at a German university hospital (I, II) and on a garden terrace during a mild summer evening (III). Young wines induced cell stress and damage while significantly reducing cytoprotective proteins in HepG2 hepatoma cells. Sympathetic activity and multitasking skills were altered depending on wines' ages. Hangovers tended to be aggravated by young red wine. Aged variants performed better in terms of aroma and overall quality but worse in optical appearance. We found no evidence for a red wine storage lesion. However, we plead for consensus-based guidelines for proper storage, as it is common in clinical medicine.

Perioperative Vascular Biomarker Profiling in Elective Surgery Patients Developing Postoperative Delirium: A Prospective Cohort Study.

BACKGROUND: Postoperative delirium (POD) ranks among the most common complications in surgical patients. Blood-based biomarkers might help identify the patient at risk. This study aimed to assess how serum biomarkers with specificity for vascular and endothelial function and for inflammation are altered, prior to or following surgery in patients who subsequently develop POD. METHODS: This was a study on a subcohort of consecutively recruited elective non-cardiac as well as cardiac surgery patients (age > 60 years) of the single-center PROPDESC trial at a German tertiary care hospital. Serum was sampled prior to and following surgery, and the samples were subjected to bead-based multiplex analysis of 17 serum proteins (IL-3, IL-8, IL-10, Cripto, CCL2, RAGE, Resistin, ANGPT2, TIE2, Thrombomodulin, Syndecan-1, E-Selectin, VCAM-1, ICAM-1, CXCL5, NSE, and uPAR). Development of POD was assessed during the first five days after surgery, using the Confusion Assessment Method for ICU (CAM-ICU), the CAM, the 4-'A's test (4AT), and the Delirium Observation Scale (DOS). Patients were considered positive if POD was detected at least once during the visitation period by any of the applied methods. Non-parametric testing, as well as propensity score matching were used for statistical analysis. RESULTS: A total of 118 patients were included in the final analysis; 69% underwent non-cardiac surgery, median overall patient age was 71 years, and 59% of patients were male. In the whole cohort, incidence of POD was 28%. The male gender was significantly associated with the development of POD (p = 0.0004), as well as a higher ASA status III (p = 0.04). Incidence of POD was furthermore significantly increased in cardiac surgery patients (p = 0.002). Surgery induced highly significant changes in serum levels of almost all biomarkers except uPAR. In preoperative serum samples, none of the analyzed parameters was significantly altered in subsequent POD patients. In postoperative samples, CCL2 was significantly increased by a factor of 1.75 in POD patients (p = 0.03), as compared to the no-POD cohort. Following propensity score matching, CCL2 remained the only biomarker that showed significant differences in postoperative values (p = 0.01). In cardiac surgery patients, postoperative CCL2 serum levels were more than 3.5 times higher than those following non-cardiac surgery (p < 0.0001). Moreover, after cardiac surgery, Syndecan-1 serum levels were significantly increased in POD patients, as compared to no-POD cardiac surgery patients (p = 0.04). CONCLUSIONS: In a mixed cohort of elective non-cardiac as well as cardiac surgery patients, preoperative serum biomarker profiling with specificity for vascular dysfunction and for systemic inflammation was not indicative of subsequent POD development. Surgery-induced systemic inflammation-as evidenced by the significant increase in CCL2 release-was associated with POD, particularly following cardiac surgery. In those patients, postoperative glycocalyx injury might furthermore contribute to POD development.

Gene expression in the Angiopoietin/TIE axis is altered in peripheral tissue of ovarian cancer patients: A prospective observational study.

AIMS: Clinical studies suggest altered systemic vascular biology in cancer patients. We assessed expression patterns of endothelial activation- and vascular leakage-related genes in tumor as well as in tumor-free peripheral tissues from patients with and without ovarian cancer (OC). MAIN METHODS: Patients being scheduled for laparotomy for either gynecologic benign diagnosis (n = 10) or for advanced-stage OC (n = 22) were prospectively recruited to this observational study. Serum samples were taken preoperatively, and tissue samples were taken from peripheral abdominal wall musculature, tumor-free peritoneum and the tumor itself. KEY FINDINGS: Patients in OC group received significantly more fluid per time intraoperatively (p = 0.01). IL-8 and MCP-1/CCL2, VCAM-1 (CD 106) and ICAM-1 (CD 54) as well as Thrombomodulin were significantly increased in cancer patients' serum at baseline (p = 0.03). Expression of distinct vascular leakage-related genes (Angiopoietin-1 (ANG-1), ANG-2, TIE2, VEGFR1, VEGFR2) was significantly altered in tumor tissue of OC patients (p = 0.003), while in tumor-free peritoneal tissue, ANG-2/1 expression ratio was more than doubled in OC group (p = 0.03). In peripheral musculature, particularly genes from the ANG/TIE axis were significantly changed in OC patients (p = 0.005), suggesting a distinct vascular leakage-related genotype. Gene expression changes in OC patients were significantly associated with the postoperative fluid balance (p = 0.03). SIGNIFICANCE: Altered expression of barrier dysfunction- and angiogenesis-associated genes from the ANG/TIE axis was detected not only in tumor but also in peripheral tissues of cancer patients. This may contribute to a systemic vascular leakage-related genotype.

Risk Factors for Severe Complications in Ovarian Cancer Surgery.

Backround: Due to extensive surgical intervention for macroscopic complete cytoreduction in epithelial ovarian cancer (EOC) patients, severe complications in the postoperative course are possible. PATIENTS AND METHODS: A total of 345 EOC patients who underwent cytoreductive surgery were retrospectively evaluated regarding risk factors for an unfavorable postoperative course. Possible pre-, intra- and postoperative risk factors were statistically analyzed performing multivariate ordinal logistic regression. RESULTS: A total of 345 EOC patients underwent cytoreductive surgery. There were no complications in 114 patients, mild complications in 114 patients and severe complications in 117 patients. The risk factor evaluation identified age (p=0.049), smoking (p=0.032) and duration of surgery (p<0.0001) as significant factors for severe postoperative morbidity. CONCLUSION: In EOC patients age, smoking and the duration of surgery have significant impact on the postoperative course. Only the duration of surgery can be positively influenced by a well-trained EOC team.

Differential modulation of endothelial cell function by fresh frozen plasma.

AIMS: In vivo studies suggest a positive influence of fresh frozen plasma (FFP) on endothelial properties and vascular barrier function, leading to improved outcomes in animal sepsis models as well as in major abdominal surgery. However, those effects are incompletely described. It was our aim to evaluate in vitro effects of FFP on endothelial key functions and to identify underlying mechanisms. MATERIALS AND METHODS: Human pulmonary microvascular endothelial cells (HPMECs) were prestimulated with LPS, followed by incubation with FFP. Permeability for FITC-dextran was assessed, and intercellular gap formation was visualized. NF-κB nuclear translocation and expression of pro-inflammatory, pro-adhesion, and leakage-related genes were evaluated, and monocyte adhesion to ECs was assessed. Intracellular cAMP levels as well as phosphorylation of functional proteins were analyzed. In patients undergoing major abdominal surgery, Syndecan-1 serum levels were assessed prior to and following FFP transfusion. KEY FINDINGS: Post-incubation of HPMVECs with FFP increased intracellular cAMP levels that had been decreased by preceding LPS stimulation. On one hand, this reduced endotoxin-mediated upregulation of IL-8, ICAM-1, VCAM-1, VEGF, and ANG-2. Impaired phosphorylation of functional proteins was restored, and intercellular cohesion and barrier function were rescued. On the other hand, NF-κB nuclear translocation as well as monocyte adhesion was markedly increased by the combination of LPS and FFP. Syndecan-1 serum levels were lower in surgery patients that were transfused with FFP compared to those that were not. SIGNIFICANCE: Our data provide evidence for a differential modulation of crucial endothelial properties by FFP, potentially mediated by elevation of intracellular cAMP levels.

Dynamic changes of angiopoietins and endothelial nitric oxide supply during fluid resuscitation for major gyn-oncological surgery: a prospective observation.

BACKGROUND: Despite goal-directed hemodynamic therapy, vascular function may deteriorate during surgery for advanced abdominal tumor masses. Fluid administration has been shown to be associated with distinct changes in serum levels of functional proteins. We sought to determine how serum total protein and angiopoietin (ANG) levels change during major abdominal tumor surgery. In addition, ex vivo endothelial nitric oxide synthase (eNOS) activation as well as NO bioavailability in vivo were assessed. METHODS: 30 patients scheduled for laparotomy for late-stage ovarian or uterine cancer were prospectively included. Advanced hemodynamic monitoring as well as protocol-driven goal-directed fluid optimization were performed. Total serum protein, ANG-1, -2, and soluble TIE2 were determined pre-, intra-, and postoperatively. Phosphorylation of eNOS was assessed in microvascular endothelial cells after incubation with patient serum, and microvascular reactivity was determined in vivo by near-infrared spectroscopy and arterial vascular occlusion. RESULTS: Cardiac output as well as preload gradually decreased during surgery and were associated with a median total fluid intake of 12.8 (9.7-15.4) mL/kg*h and a postoperative fluid balance of 6710 (4113-9271) mL. Total serum protein decreased significantly from baseline (66.5 (56.4-73.3) mg/mL) by almost half intraoperatively (42.7 (36.8-51.5) mg/mL, p < 0.0001) and remained at low level. While ANG-1 showed no significant dilutional change (baseline: 12.7 (11.9-13.9) ng/mL, postop.: 11.6 (10.8 -13.5) ng/mL, p = 0.06), serum levels of ANG-2 were even increased postoperatively (baseline: 2.2 (1.6-2.6) ng/mL vs. postop.: 3.4 (2.3-3.8) ng/mL, p < 0.0001), resulting in a significant shift in ANG-2 to ANG-1 ratio. Ex vivo phosphorylation of eNOS was decreased depending on increased ANG-2 levels and ANG-2/1 ratio (Spearman r = - 0.37, p = 0.007). In vivo, increased ANG-2 levels were associated with impaired capillary recruitment and NO bioavailability (Spearman r = - 0.83, p = 0.01). CONCLUSIONS: Fluid resuscitation-associated changes in serum vascular mediator profile during abdominal tumor surgery were accompanied by impaired eNOS activity ex vivo as well as reduced NO bioavailability in vivo. Our results may explain disturbed microvascular function in major surgery despite goal-directed hemodynamic optimization.

Supraglottic devices for airway management in awake craniotomy.

Awake craniotomy is a unique technique utilized for mapping neuro and motor function during neurosurgical procedures close to eloquent brain tissue. Since active communication is required only during surgical manipulation of eloquent brain tissue and the patient is "sedated" during other parts of the procedure, different methods for anesthesia management have been explored. Furthermore, airway management ranges from spontaneous breathing to oro or nasotracheal intubation. Case reports have described the use of laryngeal masks (LMs) previously; however, its safety compared to tracheal intubation has not been assessed.We conducted a retrospective analysis of 30 patients that underwent awake craniotomy for tumor surgery to compare the feasibility and safety of different airway management strategies. Nasal fiberoptic intubation (FOI) was performed in 21 patients while 9 patients received LM for airway management. Ventilation, critical events, and perioperative complications were evaluated.Cannot intubate situation occurred in 4 cases reinserting the tube after awake phase, while no difficulties were described reinserting the LM (P < .0001). Furthermore, duration of mechanical ventilation after tumor removal was significantly lower in the LM group compared to FOI group (62 ±â€Š24 vs. 339 ±â€Š82 [min] mean ±â€Šsem, P < .0001). Postoperatively, 2 patients in each group were diagnosed with and treated for respiratory complications including pneumonia, without statistical significance between groups.In summary, LM is a feasible airway management method for patients undergoing awake craniotomy, resulting in reduced ventilation duration compared to FOI procedure.

Network of Mediators for Vascular Inflammation and Leakage Is Dysbalanced during Cytoreductive Surgery for Late-Stage Ovarian Cancer.

BACKGROUND: Cytoreductive surgery (CS) in late-stage ovarian cancer patients is often challenging due to extensive volume shifts, and high fluid intake may provoke postoperative complications. Expression of vasoactive mediators is altered in cancer patients, which may affect systemic vascular function. We sought to assess how serum levels of vasoactive markers and mediators change during CS in ovarian cancer. METHODS: Following IRB approval and informed consent, pre- and postoperative serum samples were analyzed in 26 late-stage ovarian cancer patients using multiplex protein arrays and ELISA. RESULTS: The proinflammatory cytokines and chemokines IL-6, IL-8, and CCL2 were significantly elevated after 24 hrs compared to the baseline values, with IL-6 and IL-8 being most prominently increased. While ANGPT1 remained unchanged after surgery, its competitive antagonist ANGPT2 was significantly increased. In contrast, serum levels of the ANGPT receptor TIE2 were decreased to 0.6 of the baseline values. While VEGF-D, E-selectin, P-selectin, ICAM-1, and PECAM-1 remained unchanged, serum activity of both thrombomodulin and syndecan-1 was significantly increased following surgery. CONCLUSION: We identified a regulatory network of acute-phase reaction during CS in late-stage ovarian cancer. This suggests that IL-6 exerts positive regulation of other proinflammatory mediators and, by upregulating ANGPT2 and suppressing ANGPT1, induces a serum profile that promotes vascular leakage. This may contribute to the observed hemodynamic alterations during CS procedures.

Sub-cytotoxic doses of pharmaceutical silica nanoparticles show significant impact on the proteome of HepG2 cells.

The ever-growing application of nanoparticles (NPs) in medical and pharmaceutical domains has brought issues related to their toxicity into focus. However, a profound analysis of non-acute, sub-lethal effects of engineered pharmaceutical NPs is often disregarded during such toxicological investigations. Here, two selected NPs were investigated in cultured HepG2 cells in terms of their intracellular localization and the associated impact on pharmacokinetically relevant CYP3A4 isoform, as well as the induced changes observed in the proteome of such cells. Using SILAC (Stable Isotope Labeling by Amino acids in Cell culture)-based mass spectrometry facilitated quantitative proteomics, significant proteomic changes in NP-treated hepatocytes were detected, which were subsequently analyzed via bioinformatic tools. Both, silica NPs (SiO2 NP) and cargo-free PEGylated stealth liposomes resulted in the induction of CYP3A4-activity up to 150% in a dose-dependent manner, with different time-dependent response-patterns as a function of NP-type after a single treatment. Proteomic analysis revealed that the observed metabolic alterations are only one aspect of the cellular response to NP-exposure. SiO2NPs (free in cytoplasm) caused extensive changes in the proteome, whereas liposomes (compartmentalized) seemed unproblematic as they accounted for minimal changes in the protein profile. Based on the obtained results, proteomic analyses were revealed to be highly important for the toxicological assessment of NPs. Although sub-toxic concentrations of many NPs are considered as uncritical based on standard toxicological assays, proteomic analysis indicated that drug-free NPs could cause fundamental cellular modifications, which were attributed to different causal networks and regulatory pathways.

Common carotid artery diameter responds to intravenous volume expansion: an ultrasound observation.

BACKGROUND: In case of intravascular fluid depletion, large veins react to volume expansion with dilation. Little is known about the reaction of arterial vessels. We herein report on the effect of a standardized fluid bolus on the diameter of the common carotid artery (CCA) and its association with hemodynamic parameters, assessed in 20 mechanically ventilated patients after cardiac surgery. CCA was visualized using ultrasound, and the percentage increase in diastolic diameter was calculated by measuring before and after administration of crystalloid infusion solution. Invasive arterial blood pressure and pulse pressure variation (PPV) were assessed in parallel. RESULTS: Median diastolic CCA diameter was 6.2 (Q1-Q3: 5.4-7.1) mm, and it significantly increased to 6.7 (5.8-7.3) mm upon fluid administration [5.0 (1.9-10.5) % increase]. Mean arterial blood (MAP) pressure likewise increased from 68 (70-73) to 85 (71-100) mmHg, whereas PPV was significantly reduced from 17.6 (16.8-23.9) to 13.2 (6.7-18.1) %. There was a significant association between the change in CCA diameter and the hemodynamic response (delta-MAP: r = 0.53, delta-PPV: r = 0.56; p < 0.05). Furthermore, carotid diameter measured before volume expansion significantly correlated with the delta-PPV upon fluid administration (r = -0.5; p = 0.02). CONCLUSIONS: Diameter of the CCA increases in response to intravascular volume expansion. Additional studies on the interplay between carotid geometry and intravascular fluid status are necessary.

The Use of Internal Jugular Vein Ultrasonography to Anticipate Low or High Central Venous Pressure During Mechanical Ventilation.

BACKGROUND: Critically low or high central venous pressure (CVP) values, together with systemic hypotension, can indicate hypovolemia or acute heart failure. However, measuring CVP requires the insertion of a central venous catheter, a time-consuming procedure that can be associated with severe complications. OBJECTIVE: We sought to evaluate the use of ultrasonography of the internal jugular vein (IJV) to estimate low or high CVP values in patients who were on ventilation. METHODS: Ultrasonography of IJV dimensions and the collection of hemodynamic data was performed in 47 patients, and the ratio between IJV diameter in the 30° and 0° position was calculated (ratio(30/0)). The predictive value of ratio(30/0) for estimating low and high CVP levels was analyzed using receiver operating characteristic curves. RESULTS: The median IJV diameter ratio(30/0) was 0.49. CVP ranged from 1 to 13 mm Hg (median 7 mm Hg). Seventeen patients had a CVP ≤ 5 mm Hg or lower (defined as "low"), and in 11 patients, values of ≥ 10 mm Hg were measured (defined as "high"). The corresponding IJV diameter ratios increased significantly from 0.34 (in the low CVP group) to 0.9 (in the high CVP group). Receiver operating characteristic analysis revealed a good predictive value of the ratio(30/0) for the prediction of low or high CVP values, respectively. A ratio(30/0) of < 0.45 optimally indicated a low CVP, while > 0.65 was the cutoff value to detect a CVP ≥ 10 mm Hg. CONCLUSION: The estimation of low or high CVP values by IJV ultrasonography in different patient positions can be a helpful instrument for the rapid hemodynamic assessment of the critically ill patient.

Synergistic Stimulation with Different TLR7 Ligands Modulates Gene Expression Patterns in the Human Plasmacytoid Dendritic Cell Line CAL-1.

OBJECTIVE: TLR7 ligation in plasmacytoid dendritic cells is promising for the treatment of cancer, allergy, and infectious diseases; however, high doses of ligands are required. We hypothesized that the combination of structurally different TLR7 ligands exponentiates the resulting immune response. METHODS: CAL-1 (human pDC line) cells were incubated with the TLR7-specific adenine analog CL264 and single-stranded 9.2s RNA. Protein secretion was measured by ELISA. Microarray technique was used to detect modified gene expression patterns upon synergistic stimulation, revealing underlying functional groups and networks. Cell surface binding properties were studied using FACS analysis. RESULTS: CL264 in combination with 9.2s RNA significantly enhanced cytokine and interferon secretion to supra-additive levels. This effect was due to a stronger stimulation of already regulated genes (by monostimulation) as well as to recruitment of thus far unregulated genes. Top scoring canonical pathways referred to immune-related processes. Network analysis revealed IL-1ß, IL-6, TNF, and IFN-ß as major regulatory nodes, while several minor regulatory nodes were also identified. Binding of CL264 to the cell surface was enhanced by 9.2s RNA. CONCLUSION: Structurally different TLR7 ligands act synergistically on gene expression patterns and on the resulting inflammatory response. These data could impact future strategies optimizing TLR7-targeted drug design.

Characterizing the genetic basis of innate immune response in TLR4-activated human monocytes.

Toll-like receptors (TLRs) play a key role in innate immunity. Apart from their function in host defense, dysregulation in TLR signalling can confer risk to autoimmune diseases, septic shock or cancer. Here we report genetic variants and transcripts that are active only during TLR signalling and contribute to interindividual differences in immune response. Comparing unstimulated versus TLR4-stimulated monocytes reveals 1,471 expression quantitative trait loci (eQTLs) that are unique to TLR4 stimulation. Among these we find functional SNPs for the expression of NEU4, CCL14, CBX3 and IRF5 on TLR4 activation. Furthermore, we show that SNPs conferring risk to primary biliary cirrhosis (PBC), inflammatory bowel disease (IBD) and celiac disease are immune response eQTLs for PDGFB and IL18R1. Thus, PDGFB and IL18R1 represent plausible candidates for studying the pathophysiology of these disorders in the context of TLR4 activation. In summary, this study presents novel insights into the genetic basis of the innate immune response and exemplifies the value of eQTL studies in the context of exogenous cell stimulation.

Priming with synthetic oligonucleotides attenuates pressure overload-induced inflammation and cardiac hypertrophy in mice.

AIMS: Inflammation and Toll-like receptor (TLR) signalling have been linked to the development of cardiac hypertrophy following transverse aortic constriction (TAC). In the present study, we investigated whether pre-treatment with the synthetic TLR9 ligands 1668-thioate or 1612-thioate modulates the progression of TAC-induced cardiac inflammation and hypertrophy. METHODS AND RESULTS: C57BL/6N-mice were pre-treated with 1668-thioate, 1612-thioate (0.25 nmol/g, i.p.), or phosphate-buffered saline 16 h prior to TAC or sham surgery. Heart-weight/body-weight ratio (HW/BW), cardiomyocyte cell size, cellular macrophage accumulation, myofibroblast differentiation, and collagen deposition were investigated for up to 28 days. Cardiac function was monitored using a pressure-volume catheter and M-mode echocardiography. Inflammatory gene expression in the heart was analysed via gene array, while the time course of mRNA expression of key inflammatory mediators was assessed via RT-qPCR. TAC increased the HW/BW ratio and cardiomyocyte cell size and induced macrophage accumulation, myofibroblast differentiation, and collagen deposition. These changes were accompanied by cardiac inflammation and a significant loss of left ventricular function. Pre-treatment with cytosine-phosphate-guanine (CpG)-containing 1668-thioate attenuated the inflammatory response, the progression of cardiac hypertrophy, and cardiac remodelling, which resulted in a prolonged preservation of left ventricular function. These changes were induced to a smaller extent by the use of the non-CG-containing oligodeoxynucleotide 1612-thioate. CONCLUSION: Pre-treatment with 1668-thioate attenuated cardiac hypertrophy following pressure overload, possibly by modifying the hypertrophy-induced inflammatory response, thereby reducing cardiac growth and fibrosis as well as delaying loss of cardiac function.

Global changes in gene expression and synergistic interactions induced by TLR9 and TLR3.

The innate immune system is triggered when pathogen-associated molecular patterns (PAMPs) expressed by infectious microorganisms interact with toll-like receptors (TLR) present on immune cells. Individual TLRs signal through distinct molecular pathways. For example, TLR9 interacts with unmethylated CpG motifs expressed by bacterial DNA and triggers via a MyD88 dependent pathway whereas TLR3 recognizes viral RNA through a MyD88-independent pathway. Bioinformatic analysis of microarray data was used to identify the regulatory patterns underlying changes in gene expression induced when RAW 264.7 macrophages were stimulated via TLR9 by CpG oligonucleotides (ODN) and/or via TLR3 by poly (I:C). While the genes activated by each ligand mediated similar functions, poly (I:C) elicited a larger and more diverse change in gene expression. Co-stimulation with both ligands accelerated gene expression and synergistically activated genes primarily associated with immune function. This is the first work to compare global changes in gene regulation triggered by distinct TLR pathways and clarify their impact on gene expression.

Induction of Bim and Bid gene expression during accelerated apoptosis in severe sepsis.

INTRODUCTION: In transgenic animal models of sepsis, members of the Bcl-2 family of proteins regulate lymphocyte apoptosis and survival of sepsis. This study investigates the gene regulation of pro-apoptotic and anti-apoptotic members of the Bcl-2 family of proteins in patients with early stage severe sepsis. METHODS: In this prospective case-control study, patients were recruited from three intensive care units (ICUs) in a university hospital. Sixteen patients were enrolled when they fulfilled the criteria of severe sepsis. Ten critically ill but non-septic patients and 11 healthy volunteers served as controls. Blood samples were immediately obtained at inclusion. To confirm the presence of accelerated apoptosis in the patient groups, caspase-3 activation and phosphatidylserine externalisation in CD4+, CD8+ and CD19+ lymphocyte subsets were assessed using flow cytometry. Specific mRNAs of Bcl-2 family members were quantified from whole blood by real-time PCR. To test for statistical significance, Kruskal-Wallis testing with Dunn's multiple comparison test for post hoc analysis was performed. RESULTS: In all lymphocyte populations caspase-3 (p < 0.05) was activated, which was reflected in an increased phosphatidylserine externalisation (p < 0.05). Accordingly, lymphocyte counts were decreased in early severe sepsis. In CD4+ T-cells (p < 0.05) and B-cells (p < 0.001) the Bcl-2 protein was decreased in severe sepsis. Gene expression of the BH3-only Bim was massively upregulated as compared with critically ill patients (p < 0.001) and 51.6-fold as compared with healthy controls (p < 0.05). Bid was increased 12.9-fold compared with critically ill patients (p < 0.001). In the group of mitochondrial apoptosis inducers, Bak was upregulated 5.6-fold, while the expression of Bax showed no significant variations. By contrast, the pro-survival members Bcl-2 and Bcl-xl were both downregulated in severe sepsis (p < 0.001 and p < 0.05, respectively). CONCLUSIONS: In early severe sepsis a gene expression pattern with induction of the pro-apoptotic Bcl-2 family members Bim, Bid and Bak and a downregulation of the anti-apoptotic Bcl-2 and Bcl-xl proteins was observed in peripheral blood. This constellation may affect cellular susceptibility to apoptosis and complex immune dysfunction in sepsis.

Synthetic oligonucleotides as modulators of inflammation.

Synthetic oligodeoxynucleotides (ODN) containing unmethylated CpG motifs mimic the immunostimulatory activity of bacterial DNA. CpG ODN directly stimulate human B cells and plasmacytoid dendritic cells, promote the production of Th1 and proinflammatory cytokines, and trigger the maturation/activation of professional APC. CpG ODN are finding use in the treatment of cancer, allergy, and infection. In contrast, ODN containing multiple TTAGGG motifs mimic the immunosuppressive activity of self-DNA, down-regulating the production of proinflammatory and Th1 cytokines. Preclinical studies suggest that "suppressive" ODN may slow or prevent diseases characterized by pathologic immune stimulation, including autoimmunity and septic shock. Extensive studies in animal models suggest that the therapeutic value of CpG and TTAGGG ODN may be optimized by early administration.

Oxidoreductase Macrophage Migration Inhibitory Factor is simultaneously increased in leukocyte subsets of patients with severe sepsis.

The oxidoreductase Macrophage Migration Inhibitory Factor (MIF) is discussed as a promising target for immunomodulatory therapy in patients with severe sepsis. Moreover, MIF expresses tautomerase as well as thiol-protein oxidoreductase activities and has a potential role in cellular redox homeostasis, apoptosis inhibition, endotoxin responsiveness as well as regulation of nuclear transcription factors. To further elucidate a potential role of intracellular MIF in severe sepsis, we assessed alterations of intracellular MIF content in peripheral blood leukocytes of patients with severe sepsis in comparison to healthy controls and non-septic patients after major surgery. Intracellular MIF was significantly elevated simultaneously in lymphocytes, B-cells, macrophages and granulocytes of patients with severe sepsis when compared to healthy control individuals (p < 0.05) and increased when compared to non-septic patients after major surgery. In parallel, plasma MIF levels were elevated in severe sepsis (p < 0.05). There was no difference of intracellular MIF in lymphocytes, B-cells, macrophages or granulocytes between surviving and non-surviving patients with severe sepsis (p > 0.05). However, in survivors LPS ex vivo stimulation increased MIF secretion but not in non-survivors of sepsis (p < 0.05). This finding underlines the role of intracellular MIF in inflammatory diseases. It suggests monitoring of intracellular MIF in further clinical and non-clinical research valuable.