Rethink analgo-sedation in digestive endoscopy: the role of scientific societies in tracing training path.

OBJECTIVE: The Italian Society of Anesthesia, Analgesia, Reanimation and Intensive Care Medicine (SIAARTI) and the Italian Society of Digestive Endoscopy (SIED) worked together to produce a joint Good Clinical Practice (GCP) on analgo-sedation in digestive endoscopy and launched a survey to support the document. The aim was to identify and describe the actual clinical practice of sedation in Italian digestive endoscopy units and offer material for a wider and more widespread discussion among anesthetists and endoscopists. SUBJECTS AND METHODS: A national survey was planned, in order to support the statements of the GCP. Twelve thousand and five hundred questionnaires were sent to the members of SIAARTI and SIED in June 2020. RESULTS: A total of 662 forms (5.3%) returned completed. Highly complex procedures are performed according to 70% of respondents; daily anesthesiologist's assistance is guaranteed in 26%, for scheduled sessions in 14.5% and as needed in 8%. 69% of respondents declared not to have a dedicated team of anesthesiologists, while just 5% reported an anesthesiologist in charge. A complete monitoring system was assured by 70% of respondents. Dedicated pathways for COVID-19-positive patients were confirmed in <40% of the answers. With regard to moderate/deep sedation, 90% of respondents stated that an anesthetist decides timing and doses. Propofol was exclusively administered by anesthetists according to 94% of answers, and for 6% of respondents the endoscopist is allowed to administer propofol in presence of a dedicated nurse, but with a readily available anesthetist. Only 32.8% of respondents reported institutional training courses on procedural analgo-sedation. CONCLUSIONS: The need to provide patients scheduled for endoscopy procedures with an adequate analgo-sedation is becoming an increasing concern, well-known in almost all countries, but many factors compromise the quality of patient care. Results of a national survey would give strength to the need for a shared GCP in gastrointestinal endoscopy. Training and certification of non-anesthetist professionals should be one of the main ways to center the objective.

Protocol for Probing Regulated Lysosomal Activity and Function in Living Cells.

Lysosomes are the catabolic center of the cell. Limitations of many lysosomal tracers include low specificity and lack of reliable physiological readouts for changes in growth factor-regulated lysosomal activity. The imaging-based protocols described here provide insights at the cellular level to quantify functions essential to lysosomal biology, including ß-glucosidase enzymatic cleavage, active Cathepsin D, and pH regulation in real time. These optimized protocols, applied in different cell types and pathophysiologic contexts, provide useful tools to study lysosome function in cultured living cells. For complete details on the use and execution of this protocol, please refer to Albrecht et al. (2020).

Effect Of α2-Adrenergic Agonists And Antagonists On Cytokine Release From Human Lung Macrophages Cultured In Vitro.

The most trusted hypothesis to explain how α2-adrenergic agonists may preserve pulmonary functions in critically ill patients is that they directly act on macrophages by interfering with an autocrine/paracrine adrenergic system that controls cytokine release through locally synthetized noradrenaline and α1- and α2-adrenoreceptors. We tested this hypothesis in primary cultures of resident macrophages from human lung (HLMs). HLMs were isolated by centrifugation on percoll gradients from macroscopically healthy human lung tissue obtained from four different patients at the time of lung resection for cancer. HLMs from these patients showed a significant expression of α2A, α2B and α2C adrenoreceptors both at the mRNA and at the protein level. To evaluate whether α2 adrenoreceptors controlled cytokine release from HMLs, we measured IL-6, IL-8 and TNF-α concentrations in the culture medium in basal conditions and after preincubation with several α2-adrenergic agonists or antagonists. Neither the pretreatment with the α2-adrenergic agonists clonidine, medetomidine or dexdemetomidine or with the α2-adrenergic antagonist yohimbine caused significant changes in the response of any of these cytokines to LPS. These results show that, different from what reported in rodents, clonidine and dexdemetomidine do not directly suppress cytokine release from human pulmonary macrophages. This suggests that alternative mechanisms such as effects on immune cells activation or the modulation of autonomic neurotransmission could be responsible for the beneficial effects of these drugs on lung function in critical patients.

Navigator® and SmartPilot® View are helpful in guiding anesthesia and reducing anesthetic drug dosing.

BACKGROUND: The recently introduced Navigator® (GE Healthcare, Helsinki, Finland) and SmartPilot® View (Dräger Medical, Lübeck, Germany) show the concentrations and predicted effects of combined anesthetic drugs, and should facilitate more precisely their titration. Our aim was to evaluate if Navigator® or SmartPilot® View guided anesthesia was associated with a good quality of analgesia, depth of hypnosis and may reduce anesthetic requirements. METHODS: We performed a prospective non-randomized study. Sixty ASA I-II patients undergoing balanced general anesthesia for abdominal and plastic surgery were enrolled. Patients were divided in 4 groups. Group 1 (N. 15) and group 3 (N. 15) were cases in whom anesthesia was performed with standard monitoring plus the aid of Navigator® (Nav) or SmartPilot® View (SPV) display. Group 2 (N. 15) and group 4 (N. 15) were controls in whom anesthesia was performed with standard monitoring (heart rate, NIBP, SpO2, end-tidal CO2, end-expired sevoflurane concentration, train of four, Bispectral Index [Aspect Medical Systems, Natick, MA, USA] or Entropy [GE Healthcare]). Patients' vital parameters and end-expired sevoflurane concentration were recorded during anesthesia. RESULTS: All patients recovered uneventfully and showed hemodynamic stability. End-tidal sevoflurane concentrations values [median (min-max)], during maintenance of anesthesia, were significantly (P<0.05) lower in SPV [1.1% (0.8-1.5)] and Nav [1%(0.8-1.8)] groups compared to SPV-control group [1.5%(1-2.5)] and Nav-control group [1.5%(0.8-2)]. BIS and entropy values were respectively higher in the SPV group [53 (46-57)] compared to the control group [43 (37-51)] (P<0.05) and Nav group [53 (43-60)] compared to the control group [41 (35-51)] (P<0.05). No significant differences in Remifentanil dosing were observed in the four groups. CONCLUSION: Navigator® and SmartPilot® View may be of clinical use in monitoring adequacy of anesthesia. Both displays can optimize the administration and monitoring of anesthetic drugs during general anesthesia and may reduce the consumption of volatile anesthetic agents.

Coagulopathy induced by acidosis, hypothermia and hypocalcaemia in severe bleeding.

Acidosis, hypothermia and hypocalcaemia are determinants for morbidity and mortality during massive hemorrhages. However, precise pathological mechanisms of these environmental factors and their potential additive or synergistic anticoagulant and/or antiplatelet effects are not fully elucidated and are at least in part controversial. Best available evidences from experimental trials indicate that acidosis and hypothermia progressively impair platelet aggregability and clot formation. Considering the cell-based model of coagulation physiology, hypothermia predominantly prolongs the initiation phase, while acidosis prolongs the propagation phase of thrombin generation. Acidosis increases fibrinogen breakdown while hypothermia impairs its synthesis. Acidosis and hypothermia have additive effects. The effect of hypocalcaemia on coagulopathy is less investigated but it appears that below the cut-off of 0.9 mmol/L, several enzymatic steps in the plasmatic coagulation system are blocked while above that cut-off effects remain without clinical sequalae. The impact of environmental factor on hemostasis is underestimated in clinical practice due to our current practice of using routine coagulation laboratory tests such as partial thromboplastin time or prothrombin time, which are performed at standardized test temperature, after pH correction, and upon recalcification. Temperature-adjustments are feasible in viscoelastic point-of-care tests such as thrombelastography and thromboelastometry which may permit quantification of hypothermia-induced coagulopathy. Rewarming hypothermic bleeding patients is highly recommended because it improves patient outcome. Despite the absence of high-quality evidence, calcium supplementation is clinical routine in bleeding management. Buffer administration may not reverse acidosis-induced coagulopathy but may be essential for the efficacy of coagulation factor concentrates such as recombinant activated factor VII.

S100B induces the release of pro-inflammatory cytokines in alveolar type I-like cells.

S100B, a 21kDa cytosolic calcium-binding protein of the EF-hand type, present in high abundance in the brain, stimulates inflammatory responses in different cellular types inside and outside the central nervous system. Most of extracellular S100B effects are mediated by Receptor for Advanced Glycation End-products (RAGE). RAGE is highly expressed in lung by Alveolar Type-I (AT-I) cells and its activation contributes to ALI/ARDS pathogenesis. In this in-vitro study, we tested the hypothesis that S100B stimulates an ATI-derived cell line (R3/1) to secrete inflammatory mediators involved in lung inflammation. Our main result is that S100B stimulates R3/1 cells to secrete TNF-alpha and IL-6 (well-known pro-inflammatory cytokines in lung inflammation and neurogenic pulmonary edema), but not sICAM-1, CINC-1 or CINC-3. Soluble RAGE (sRAGE) reduced S100B-dependent secretion of TNF-alpha but did not decrease S100B-dependent secretion of IL-6. Moreover, in absence of S100B, sRAGE enhanced IL-6 release. This study demonstrates that in vitro S100B dose-dependently stimulated R3/1 cells, to enhance the secretion of TNF-alpha and IL-6; S100B pro-inflammatory activity might be mediated at least in part by RAGE. Besides acting as decoy receptor, sRAGE could have pro-inflammatory properties.

Protective ventilation in experimental acute respiratory distress syndrome after ventilator-induced lung injury: a randomized controlled trial.

BACKGROUND: Low tidal volume (V(T)), PEEP, and low plateau pressure (P(PLAT)) are lung protective during acute respiratory distress syndrome (ARDS). This study tested the hypothesis that the aspiration of dead space (ASPIDS) together with computer simulation can help maintain gas exchange at these settings, thus promoting protection of the lungs. METHODS: ARDS was induced in pigs using surfactant perturbation plus an injurious ventilation strategy. One group then underwent 24 h protective ventilation, while control groups were ventilated using a conventional ventilation strategy at either high or low pressure. Pressure-volume curves (P(el)/V), blood gases, and haemodynamics were studied at 0, 4, 8, 16, and 24 h after the induction of ARDS and lung histology was evaluated. RESULTS: The P(el)/V curves showed improvements in the protective strategy group and deterioration in both control groups. In the protective group, when respiratory rate (RR) was ≈ 60 bpm, better oxygenation and reduced shunt were found. Histological damage was significantly more severe in the high-pressure group. There were no differences in venous oxygen saturation and pulmonary vascular resistance between the groups. CONCLUSIONS: The protective ventilation strategy of adequate pH or PaCO2 with minimal V(T), and high/safe P(PLAT) resulting in high PEEP was based on the avoidance of known lung-damaging phenomena. The approach is based upon the optimization of V(T), RR, PEEP, I/E, and dead space. This study does not lend itself to conclusions about the independent role of each of these features. However, dead space reduction is fundamental for achieving minimal V(T) at high RR. Classical physiology is applicable at high RR. Computer simulation optimizes ventilation and limiting of dead space using ASPIDS. Inspiratory P(el)/V curves recorded from PEEP or, even better, expiratory P(el)/V curves allow monitoring in ARDS.

The Helsinki Declaration on Patient Safety in Anesthesiology: a way forward with the European Board and the European Society of Anesthesiology.

Anesthesiology, which includes anaesthesia, perioperative care, intensive care medicine, emergency medicine and pain therapy, is acknowledged as the leading medical specialty in addressing issues of patient safety, but there is still a long way to go. Several factors pose hazards in Anesthesiology, like increasingly older and sicker patients, more complex surgical interventions, more pressure on throughput, as well as new drugs and devices. To better design educational and research strategies to improve patient safety, the European Board of Anesthesiology (EBA) and the European Society of Anesthesiology (ESA) have produced a blueprint for patient safety in Anesthesiology. This document, to be known as the Helsinki Declaration on Patient Safety in Anesthesiology, was endorsed together with the World Health Organization (WHO), the World Federation of Societies of Anesthesiologists (WFSA), and the European Patients' Federation (EPF) at the Euroanaesthesia meeting in Helsinki in June 2010. It was signed by several Presidents of National Anesthesiology Societies as well as other stakeholders. The Helsinki Declaration on Patient Safety in Anesthesiology represents a shared European view of what is necessary to improve patient safety, recommending practical steps that all anesthesiologists can include in their own clinical practice. The Italian Society of Anaesthesia, Analgesia, Reanimation and Intensive Care (SIAARTI) is looking forward to continuing work on "patient safety" issues in Europe, and to cooperating with the ESA in the best interest of European patients.

Different patterns of lung recruitment maneuvers in primary acute respiratory distress syndrome: effects on oxygenation and central hemodynamics.

BACKGROUND: The aim of this study was to test if different recruitment maneuver (RM) patterns, that achieve the same maximum pressure for the same length of time in humans, have a similar efficacy on alveolar recruitment, intrathoracic vascular pressures and flows, and on cardiac function and ventricular filling. METHODS: Forty patients were randomly allocated to undergo different RM patterns: sustained inflation (SI) or pressure controlled ventilation (PCV). The RM methods tested are as follows: SI was achieved by raising peak inspiratory pressure to 45 cmH(2)O and sustaining it for 40 seconds. The PCV was set to obtain a 45 cmH(2)O peak inspiratory pressure for 2 minutes, I:E 1:2, PEEP 16 RR 8/min. During the study period, patients were mechanically ventilated to obtain a volume of 6 mL/kg, FiO(2) 0.7, PEEP 14, RR 14, Pplateau < or =30 cmH(2)O according to the ARDSnet trial. All patients were sedated and paralyzed during the study period. All patients were given i.v. norepinephrine. Heart rate, pulse oxymetry, blood pressure, pulmonary artery catheter data (C.I., PVRI, MPAP, PAOP, SvO(2), CVP), and arterial and right heart side venous blood gas analysis data (ph, PaO(2), PaCO(2), SatO(2), HCO(3)(-), SvO(2)) were recorded before and immediately after the lung recruitment maneuver. The static compliance of the respiratory system (CRS) was recorded. Echocardiographic spot evaluations before and after RM were obtained in all cases. RESULTS: Central venous pressure increased during RM. Mean pulmonary artery pressure, pulmonary capillary wedge pressure and pulmonary vascular resistance index were reduced during PCV RM compared to SI RM (P<0.05). The right ventricle stroke work index decreased to a major extent during PCV RM (P<0.05). The P/F ratio was significantly increased after PCV RM compared to SI RM (P<0.05). PaCO(2) levels were similar in the two groups. Compared to baseline, the Qs/Qt decreased significantly after the PCV recruitment maneuver. Ventricular end-diastolic and end-systolic areas decreased during both RM protocols, but they were decreased to a greater extent after SI RM than after PCV RM (P<0.05). The eccentricity index increased from baseline after the SI RM (P<0.05). CONCLUSION: Given its comparable, or even superior, performance over the SI RM, we favor the PCV technique over the time-honored SI maneuver.

Crossveinless-2 is required for the relocalization of Chordin protein within the vertebral field in mouse embryos.

Bone morphogenetic proteins (BMPs), as well as the BMP-binding molecules Chordin (Chd), Crossveinless-2 (CV2) and Twisted Gastrulation (Tsg), are essential for axial skeletal development in the mouse embryo. We previously reported a strong genetic interaction between CV2 and Tsg and proposed a role for this interaction in the shaping of the BMP morphogenetic field during vertebral development. In the present study we investigated the roles of CV2 and Chd in the formation of the vertebral morphogenetic field. We performed immunostainings for CV2 and Chd protein on wild-type, CV2(-/-) or Chd(-/-) mouse embryo sections at the stage of onset of the vertebral phenotypes. By comparing mRNA and protein localizations we found that CV2 does not diffuse away from its place of synthesis, the vertebral body. The most interesting finding of this study was that Chd synthesized in the intervertebral disc accumulates in the vertebral body. This relocalization does not take place in CV2(-/-) mutants. Instead, Chd was found to accumulate at its site of synthesis in CV2(-/-) embryos. These results indicate a CV2-dependent flow of Chd protein from the intervertebral disc to the vertebral body. Smad1/5/8 phosphorylation was decreased in CV2(-/-)vertebral bodies. This impaired BMP signaling may result from the decreased levels of Chd/BMP complexes diffusing from the intervertebral region. The data indicate a role for CV2 and Chd in the establishment of the vertebral morphogenetic field through the long-range relocalization of Chd/BMP complexes. The results may have general implications for the formation of embryonic organ-forming morphogenetic fields.

The RNA-binding protein bicaudal C regulates polycystin 2 in the kidney by antagonizing miR-17 activity.

The RNA-binding protein Bicaudal C is an important regulator of embryonic development in C. elegans, Drosophila and Xenopus. In mouse, bicaudal C (Bicc1) mutants are characterized by the formation of fluid-filled cysts in the kidney and by expansion of epithelial ducts in liver and pancreas. This phenotype is reminiscent of human forms of polycystic kidney disease (PKD). Here, we now provide data that Bicc1 functions by modulating the expression of polycystin 2 (Pkd2), a member of the transient receptor potential (TRP) superfamily. Molecular analyses demonstrate that Bicc1 acts as a post-transcriptional regulator upstream of Pkd2. It regulates the stability of Pkd2 mRNA and its translation efficiency. Bicc1 antagonized the repressive activity of the miR-17 microRNA family on the 3'UTR of Pkd2 mRNA. This was substantiated in Xenopus, in which the pronephric defects of bicc1 knockdowns were rescued by reducing miR-17 activity. At the cellular level, Bicc1 protein is localized to cytoplasmic foci that are positive for the P-body markers GW182 and HEDLs. Based on these data, we propose that the kidney phenotype in Bicc1(-/-) mutant mice is caused by dysregulation of a microRNA-based translational control mechanism.

Extracellular regulation of BMP signaling.

In the developing organism, cells differentiate, divide and die as part of groups of hundreds or thousands of cells called 'morphogenetic fields'. Fields have the remarkable property of self-regulation: for example, if the forelimb field is bisected, each half can give rise to a complete limb after transplantation, as discovered by Ross Harrison in 1918. Therefore, cells in the morphogenetic field are capable of long-range communication with each other in order to ascertain their position [1]. This positional information is relayed in the extracellular space in the form of concentration gradients of specific classes of extracellular molecules called 'morphogens' that trigger cellular responses by binding and activating cell surface receptors. Here, we focus on a family of morphogens called 'Bone Morphogenetic Proteins' (BMPs), which has provided a new paradigm for signaling regulation in the extracellular space.

Integration of BMP and Wnt signaling via vertebrate Smad1/5/8 and Drosophila Mad.

BMPs pattern the dorsal-ventral axis of vertebrate embryos. Smad1/5/8 transduces the BMP signal, and receives phosphorylation inputs from both MAPK and GSK3. Phosphorylation of Smad1 by MAPK and GSK3 result in its polyubiquitination and transport to the centrosome where it is degraded by the proteasome. These linker phosphorylations inhibit BMP/Smad1signaling by shortening its duration. Wnt, which negatively regulates GSK3 activity, prolongs the BMP/Smad1 signal. Remarkably, linker-phosphorylated Smad1 has been shown to be inherited asymmetrically during cell division. Drosophila contains a single Smad1/5/8 homologue, Mad, and is stabilized by phosphorylation-resistant mutations at GSK3 sites, causing Wingless-like effects. We summarize here the significance of linker-phosphorylated Smad1/Mad in relation to signal intensity and duration, and how this integrates the Wnt and BMP pathways during cell differentiation.

Spemann's organizer and the self-regulation of embryonic fields.

Embryos and developing organs have the remarkable ability of self-regenerating after experimental manipulations. In the Xenopus blastula half-embryos can regenerate the missing part, producing identical twins. Studies on the molecular nature of Spemann's organizer have revealed that self-regulation results from the battle between two signaling centers under reciprocal transcriptional control. Long-range communication between the dorsal and ventral sides is mediated by the action of growth factor antagonists - such as the BMP antagonist Chordin - that regulate the flow of BMPs within the embryonic morphogenetic field. BMPs secreted by the dorsal Spemann organizer tissue are released by metalloproteinases of the Tolloid family, which cleave Chordin at a distance of where they were produced. The dorsal center secretes Chordin, Noggin, BMP2 and ADMP. The ventral center of the embryo secretes BMP4, BMP7, Sizzled, Crossveinless-2 and Tolloid-related. Crossveinless-2 binds Chordin/BMP complexes, facilitating their flow towards the ventral side, where BMPs are released by Tolloid allowing peak BMP signaling. Self-regulation occurs because transcription of ventral genes is induced by BMP while transcription of dorsal genes is repressed by BMP signals. This assures that for each action of Spemann's organizer there is a reaction in the ventral side of the embryo. Because both dorsal and ventral centers express proteins of similar biochemical activities, they can compensate for each other. A novel biochemical pathway of extracellular growth factor signaling regulation has emerged from these studies in Xenopus. This remarkable dorsal-ventral positional information network has been conserved in evolution and is ancestral to all bilateral animals.