Minimum Quality Threshold in Pre-Clinical Sepsis Studies (MQTiPSS): an international expert consensus initiative for improvement of animal modeling in sepsis.

PURPOSE: Pre-clinical animal studies precede the majority of clinical trials. While the clinical sepsis definitions and recommended treatments are regularly updated, a systematic review of pre-clinical models of sepsis has not been done and clear modeling guidelines are lacking. To address this deficit, a Wiggers-Bernard Conference on pre-clinical sepsis modeling was held in Vienna in May, 2017. The conference goal was to identify limitations of pre-clinical sepsis models and to propose a set of guidelines, defined as the "Minimum Quality Threshold in Pre-Clinical Sepsis Studies" (MQTiPSS), to enhance translational value of these models. METHODS: 31 experts from 13 countries participated and were divided into 6 thematic Working Groups (WG): (1) Study Design, (2) Humane modeling, (3) Infection types, (4) Organ failure/dysfunction, (5) Fluid resuscitation and (6) Antimicrobial therapy endpoints. As basis for the MQTiPSS discussions, the participants conducted a literature review of the 260 most highly cited scientific articles on sepsis models (2002-2013). RESULTS: Overall, the participants reached consensus on 29 points; 20 at "recommendation" (R) and 9 at "consideration" (C) strength. This Executive Summary provides a synopsis of the MQTiPSS consensus (Tables 1, 2 and 3). CONCLUSIONS: We believe that these recommendations and considerations will serve to bring a level of standardization to pre-clinical models of sepsis and ultimately improve translation of pre-clinical findings. These guideline points are proposed as "best practices" that should be implemented for animal sepsis models. In order to encourage its wide dissemination, this article is freely accessible in Shock, Infection and Intensive Care Medicine Experimental.

Correction to: Minimum Quality Threshold in Pre-Clinical Sepsis Studies (MQTiPSS): an international expert consensus initiative for improvement of animal modeling in sepsis.

The original version of this article unfortunately contained mistakes.

Organ-Specific Oxidative Events under Restrictive Versus Full Reperfusion Following Hemorrhagic Traumatic Shock in Rats.

Background aim: Reperfusion after hemorrhagic traumatic shock (HTS) is often associated with complications that are partly ascribed to the formation of reactive oxygen species (ROS). The aim of our study was to compare the effects of restrictive reperfusion (RR) to rapid full reperfusion (FR) on ROS formation and/or oxidative events. MATERIALS AND METHODS: Anesthetized male rats were randomly subjected to HTS followed by FR (75 mL/kg/h) or RR (30 mL/kg/h for 40 min, followed by 75 mL/kg/h) with Ringer's solution (n = 8/group). Compartment-specific ROS formation was determined by infusion of ROS scavenger 1-hydroxy-3-carboxy-2,2,5,5-tetramethyl-pyrrolidine hydrochloride (CP-H) during resuscitation, followed by electron paramagnetic resonance spectroscopy. Sham-operated animals (n = 8) served as controls. The experiment was terminated 100 min post-shock. RESULTS: Mean arterial pressure was significantly higher in the FR compared to the RR group during early reperfusion. Only RR animals, not FR animals, showed significantly higher ROS concentrations in erythrocytes (1951 ± 420 vs. 724 ± 75 AU) and in liver (474 ± 57 vs. 261 ± 21 AU) compared to sham controls. This was accompanied by elevated alanine aminotransferase and creatinine levels in RR animals compared to both shams and FR animals, while lipid peroxidation products (thiobarbituric acid reactive substances) were significantly increased only in the kidney in the FR group (p < 0.05). RR animals showed significantly higher plasma peroxiredoxin-4 values when compared to the FR group (20 ± 2 vs. 14 ± 0.5 RLU). CONCLUSION: Restrictive reperfusion after HTS is associated with increased ROS formation in erythrocytes and liver compared to sham controls. Moreover, the restrictive reperfusion is associated with a more pronounced injury to the liver and kidney, which is likely mediated by other than lipid peroxidation process and/or oxidative stress reactions.

Alterations in nitric oxide homeostasis during traumatic brain injury.

Changes in nitric oxide (NO) levels have been often associated with various forms of trauma, including secondary damage after traumatic brain injury (TBI). Several studies demonstrate the upregulation of NO synthase (NOS) enzymes, and concomitant increases in brain NO levels, which contribute to the TBI-associated glutamate cytotoxicity, including the pathogenesis of mitochondrial dysfunction. TBI is also associated with elevated NO levels in remote organs, indicating that TBI can induce systemic changes in NO regulation, which can be either beneficial or detrimental. Here we review the possible mechanisms responsible for changes in NO metabolism during TBI. Better understanding of the changes in NO homeostasis in TBI will be necessary to design rational therapeutic approaches for TBI. This article is part of a Special Issue entitled: Immune and Metabolic Alterations in Trauma and Sepsis edited by Dr. Raghavan Raju.

Concentrated lyophilized plasma used for reconstitution of whole blood leads to higher coagulation factor activity but unchanged thrombin potential compared with fresh-frozen plasma.

BACKGROUND: During massive hemorrhage, it is recommended to transfuse red blood cells, platelet concentrate, and fresh-frozen plasma in a ratio close to 1:1:1. To avoid the thawing process of fresh frozen plasma, lyophilized plasma (LP) is increasingly used. Evidence is limited on the activity of coagulation factors in reconstituted blood using LP and concentrated LP versions. STUDY DESIGN AND METHODS: Whole blood from ten healthy volunteers was separated into red blood cell, fresh frozen plasma, and platelet concentrate units. Aliquots of red blood cells and plasma concentrate were mixed with either fresh frozen plasma (200 mL) or LP at reconstitution ratios of 2:1:1, 1:1:1, and 1:1:2. LP was used either at the recommended standard volume of 200 mL (LP200) or was more concentrated at volumes of 100 and 50 mL (LP100 and LP50, respectively). The hemostatic capacity of each reconstituted whole blood sample was tested with blood cell counts, standard coagulation tests, factor activity, thrombin generation, and viscoelastic assays. RESULTS: Hematocrit, platelet counts, and fibrinogen levels of the three ratios were similar between FFP200 and LP200 units but were lower compared with the corresponding ratios in LP100 and LP50 units. The activity of procoagulant and anticoagulant factors increased linearly with the increasing plasmatic fraction and, at 1:1:2 ratio, was significantly higher in LP50 units compared with FFP200 and LP200 units. Thrombin generation was similar throughout the four plasma groups at any ratio. CONCLUSIONS: Decreasing the dilution volume of LP facilitates reaching higher hematocrit and coagulation protein levels without a relevant increase in thrombin generation. This is due to preserved balance between procoagulant and anticoagulant factors in the concentrated LP preparations.

Dual inhibition of thrombin and activated factor X attenuates disseminated intravascular coagulation and protects organ function in a baboon model of severe Gram-negative sepsis.

BACKGROUND: Inhibition of procoagulant pathways may improve outcome in sepsis. We examined whether a dual short-acting thrombin (factor II) and factor X (FX)a inhibitor (SATI) ameliorates sepsis-induced disseminated intravascular coagulation (DIC) and is organ-protective. METHODS: Escherichia coli were infused for 2 h in 22 anesthetized baboons. The control (CO) group (n = 8) received sterile isotonic solution only. In the treatment groups, SATI was administered starting 15 minutes after the end of the bacterial exposure. In the low-dose group (LD-SATI, n = 8), SATI was infused with 75 µg/kg/h for the first hour, followed by 23 µg/kg/h until the end of the study. In the high-dose SATI group (HD-SATI, n = 6), 225 µg/kg/h was administered for the first hour followed by continuous infusion of 69 µg/kg/h until termination of the study. RESULTS: Sepsis-induced DIC was attenuated, as reflected by lower peak thrombin-antithrombin complexes (threefold) and D-dimer levels (twofold) in both SATI groups compared to the CO. This coincided with strongly improved cell/organ protection assessed by decreased levels of lactate dehydrogenase (threefold), creatinine (twofold), aspartate aminotransferase (threefold), and amylase (twofold) compared to the CO group. Anuria, which started at 8 h in the CO group, was prevented in both SATI groups. Peak interleukin-6 release at 12 h was prevented in the treatment groups. In both SATI groups, fewer catecholamines were necessary and no bleeding complications were observed. CONCLUSIONS: Dual inhibition of thrombin and FXa preserved activation of coagulation, protected organ function and ameliorated inflammation in severe Gram-negative sepsis in baboons. SATI could be a novel therapeutic agent against sepsis-induced DIC.

TNF-α release capacity is suppressed immediately after hemorrhage and resuscitation.

PURPOSE: It has been suggested that patients with traumatic insults are resuscitated into a state of an early systemic inflammatory response. We aimed to evaluate the influence of hemorrhagic shock and resuscitation (HSR) upon the inflammatory response capacity assessed by overall TNF-α secretion capacity of the host compared to its release from circulating leukocytes in peripheral circulation. METHODS: Rats (8/group) subjected to HS (MAP of 30-35 mmHg for 90 min followed by resuscitation over 50 min) were challenged with Lipopolysaccharide (LPS), 1 µg/kg intravenously at the end of resuscitation (HSR-LPS group) or 24 h later (HSR-LPS24 group). Control animals were injected with LPS without bleeding (LPS group). Plasma TNF-α was measured at 90 min after the LPS challenge. In addition, whole blood (WB) was obtained either from healthy controls (CON) immediately after resuscitation (HSR), or at 24 h post-shock (HSR 24). WB was incubated with LPS (100 ng/mL) for 2 h at 37 °C. TNF-α concentration and LPS binding capacity (LBC) was determined. RESULTS: Compared to LPS group, HSR followed by LPS challenge resulted in suppression of plasma TNF-α in HSR-LPS and HSR-LPS24 groups (1835 ± 478, 273 ± 77, 498 ± 200 pg/mL, respectively). Compared to CON the LPS-induced TNF-α release capacity of circulating leukocytes ex vivo was strongly declined both at the end of resuscitation (HSR) and 24 h later (HSR24) (1012 ± 259, 313 ± 154, 177 ± 63 ng TNF/mL, respectively). The LBC in WB was similar between CON and HSR and only moderately enhanced in HSR24 (57 ± 6, 56 ± 6, 71 ± 5 %, respectively). CONCLUSION: Our data suggest that the overall inflammatory response capacity is decreased immediately after HSR, persisting up to 24 h, and is independent of LBC.

A One-Nearest-Neighbor Approach to Identify the Original Time of Infection Using Censored Baboon Sepsis Data.

OBJECTIVES: Sepsis therapies have proven to be elusive because of the difficulty of translating biologically sound and effective interventions in animal models to humans. A part of this problem originates from the fact that septic patients present at various times after the onset of sepsis, whereas the exact time of infection is controlled in animal models. We sought to determine whether data mining longitudinal physiologic data in a nonhuman primate model of Escherichia coli-induced sepsis could help inform the time of onset of infection. DESIGN: A nearest-neighbor approach was used to back cast the time of onset of infection in animal models of sepsis. Animal data were censored to simulate prospective monitoring at any moment along the septic infection. This was compared against an uncensored database to find the most similar animal in order to estimate the infection onset time. Leave-one-out cross-validation was used for validation. Biomarker selection was performed based on the criteria of estimation accuracy and/or ease of measurement. SETTING: Computational experimental on existing experimental data. SUBJECTS: Retrospective data from 33 septic baboons (Papio ursinus) subjected to Escherichia coli infusion. Validation was performed using 14 pigs that were subjected to surgically induced fecal peritonitis and 22 pigs that were subjected to lipopolysaccharide infusion. MEASUREMENTS AND MAIN RESULTS: Longitudinal physiologic and serum markers, time of death. The presence of uniquely changing biomarkers during septic infection enabled the estimation of infection onset time in the datasets. Various combinations of temporal biomarkers, such as WBC, oxygen content, mean arterial pressure, and heart rate, yielded estimation accuracies of up to 97.8%. The use of temporal vital signs and a single measurement of serum biomarkers yielded highly accurate estimates without the need for invasive measurements. Validation in the pig data revealed similar results despite the heterogeneity of multiple experimental cohorts. This suggests that the method may be effective if sufficiently similar subjects are present in the database. CONCLUSIONS: One nearest-neighbor analysis showed promise in accurately identifying the onset of infection given a database of known infection times and of sufficient breadth. We suggest that this approach is ready for evaluation within the clinical setting using human data.

Sensitive detection of C-reactive protein in serum by immunoprecipitation-microchip capillary gel electrophoresis.

Sepsis represents a significant cause of mortality in intensive care units. Early diagnosis of sepsis is essential to increase the survival rate of patients. Among others, C-reactive protein (CRP) is commonly used as a sepsis marker. In this work we introduce immune precipitation combined with microchip capillary gel electrophoresis (IP-MCGE) for the detection and quantification of CRP in serum samples. First high-abundance proteins (HSA, IgG) are removed from serum samples using affinity spin cartridges, and then the remaining proteins are labeled with a fluorescence dye and incubated with an anti-CRP antibody, and the antigen/antibody complex is precipitated with protein G-coated magnetic beads. After precipitation the complex is eluted from the beads and loaded onto the MCGE system. CRP could be reliably detected and quantified, with a detection limit of 25 ng/µl in serum samples and 126 pg/µl in matrix-free samples. The overall sensitivity (LOQ = 75 ng/µl, R(2) = 0.9668) of the method is lower than that of some specially developed methods (e.g., immune radiometric assay) but is comparable to those of clinically accepted ELISA methods. The straightforward sample preparation (not prone to mistakes), reduced sample and reagent volumes (including the antibodies), and high throughput (10 samples/3 h) are advantages and therefore IP-MCGE bears potential for point-of-care diagnosis.

Mouse model of posttraumatic abdominal sepsis: survival advantage of females over males does not depend on the cecum size.

BACKGROUND: Regardless of whether alone or in combination, cecal ligation and puncture (CLP) is the most commonly used model to emulate human polymicrobial sepsis. Numerous CLP studies have shown that female mice survive better than males. In adult mice, this effect may be partly due to the unequal cecum mass (larger in males), as cecum ligation is frequently not size standardized. Comparing two ligation approaches, we investigated whether cecum size influences gender-specific outcome differences. METHODS: 15-month-old (middle-aged) female and male CD-1 mice underwent sublethal trauma/hemorrhage followed by CLP 48 h later. The evaluation of cecum size (in centimeters) and filling (score) was immediately followed by two ligation protocols: (1) ileocecal ligation (IC-L; n = 28/each gender) below the ileocecal valve, and (2) apex ligation (AP-L; n = 31 males, n = 24 females) 2 cm from the apex ceci - all followed by an identical double puncture and 10-day monitoring. RESULTS: Cecum length (3.4 cm in males vs. 2.65 cm in females) and filling (2.5 score points in males vs. 1.7 in females; both p < 0.05) were always greater in male than female mice (27 vs. 44%, respectively). Compared to AP-L, the 10-day CLP mortality was higher with IC-L (86% in males and 61% in females), and this exacerbation was similar in both genders (by 21% in male and 23% in female mice). Finally, the intergender comparison demonstrated that female mice displayed a significant and similar survival advantage regardless of the ligation approach: the difference reached 25% with IC-L and 24% with AP-L. CONCLUSIONS: Standardization by AP-L did not eliminate the gender-specific difference in mortality due to posttraumatic sepsis. The greater cecum length/filling in middle-aged male mice was not responsible for their inferior survival compared to females.

Decellularized Extracellular Matrix Scaffolds for Soft Tissue Augmentation: From Host-Scaffold Interactions to Bottlenecks in Clinical Translation.

Along with a paradigm shift in looking at soft tissue fillers from space-filling to bioactive materials, decellularized extracellular matrix (DEM) fillers have gained more attention considering their superior bioactivity. However, the complex mechanisms that govern the interaction between host tissues and DEMs have been partially understood. This review first covers the mechanisms that determine immunogenicity, angiogenesis and vasculogenesis, and recellularization and remodeling after DEM implantation into host tissue, with a particular focus on related findings from filler materials. Accordingly, the review delves into the dual role of macrophages and their M1/M2 polarization paradigm to form both constructive and destructive immune responses to DEM implants. Moreover, the contribution of macrophages in angiogenesis has been elucidated, which includes but is not limited to the secretion of angiogenic growth factors and extracellular matrix (ECM) remodeling. The findings challenge the traditional view of immune cells as solely destructive entities in biomaterials and indicate their multifaceted roles in tissue regeneration. Furthermore, the review discusses how the compositional factors of DEMs, such as the presence of growth factors and matrikines, can influence angiogenesis, cell fate, and differentiation during the recellularization process. It is also shown that the biomechanical properties of DEMs, including tissue stiffness, modulate cell responses through mechanotransduction pathways, and the structural properties of DEMs, such as scaffold porosity, impact cell-cell and cell-ECM interactions. Finally, we pointed out the current clinical applications, the bottlenecks in the clinical translation of DEM biomaterials into soft tissue fillers, as well as the naïve research areas of the field.

Angiogenesis and Re-endothelialization in decellularized scaffolds: Recent advances and current challenges in tissue engineering.

Decellularization of tissues and organs has recently become a promising approach in tissue engineering and regenerative medicine to circumvent the challenges of organ donation and complications of transplantations. However, one main obstacle to reaching this goal is acellular vasculature angiogenesis and endothelialization. Achieving an intact and functional vascular structure as a vital pathway for supplying oxygen and nutrients remains the decisive challenge in the decellularization/re-endothelialization procedure. In order to better understand and overcome this issue, complete and appropriate knowledge of endothelialization and its determining variables is required. Decellularization methods and their effectiveness, biological and mechanical characteristics of acellular scaffolds, artificial and biological bioreactors, and their possible applications, extracellular matrix surface modification, and different types of utilized cells are factors affecting endothelialization consequences. This review focuses on the characteristics of endothelialization and how to optimize them, as well as discussing recent developments in the process of re-endothelialization.

Exosomal Cargo: Pro-angiogeneic, anti-inflammatory, and regenerative effects in ischemic and non-ischemic heart diseases - A comprehensive review.

Heart diseases are the primary cause of mortality and morbidity worldwide which inflict a heavy social and economic burden. Among heart diseases, most deaths are due to myocardial infarction (MI) or heart attack, which occurs when a decrement in blood flow to the heart causes injury to cardiac tissue. Despite several available diagnostic, therapeutic, and prognostic approaches, heart disease remains a significant concern. Exosomes are a kind of small extracellular vesicles released by different types of cells that play a part in intercellular communication by transferring bioactive molecules important in regenerative medicine. Many studies have reported the diagnostic, therapeutic, and prognostic role of exosomes in various heart diseases. Herein, we reviewed the roles of exosomes as new emerging agents in various types of heart diseases, including ischemic heart disease, cardiomyopathy, arrhythmia, and valvular disease, focusing on pathogenesis, therapeutic, diagnostic, and prognostic roles in different areas. We have also mentioned different routes of exosome delivery to target tissues, the effects of preconditioning and modification on exosome's capability, exosome production in compliance with good manufacturing practice (GMP), and their ongoing clinical applications in various medical contexts to shed light on possible clinical translation.

Immunogenicity of decellularized extracellular matrix scaffolds: a bottleneck in tissue engineering and regenerative medicine.

Tissue-engineered decellularized extracellular matrix (ECM) scaffolds hold great potential to address the donor shortage as well as immunologic rejection attributed to cells in conventional tissue/organ transplantation. Decellularization, as the key process in manufacturing ECM scaffolds, removes immunogen cell materials and significantly alleviates the immunogenicity and biocompatibility of derived scaffolds. However, the application of these bioscaffolds still confronts major immunologic challenges. This review discusses the interplay between damage-associated molecular patterns (DAMPs) and antigens as the main inducers of innate and adaptive immunity to aid in manufacturing biocompatible grafts with desirable immunogenicity. It also appraises the impact of various decellularization methodologies (i.e., apoptosis-assisted techniques) on provoking immune responses that participate in rejecting allogenic and xenogeneic decellularized scaffolds. In addition, the key research findings regarding the contribution of ECM alterations, cytotoxicity issues, graft sourcing, and implantation site to the immunogenicity of decellularized tissues/organs are comprehensively considered. Finally, it discusses practical solutions to overcome immunogenicity, including antigen masking by crosslinking, sterilization optimization, and antigen removal techniques such as selective antigen removal and sequential antigen solubilization.

Wet-spinnability and crosslinked Fiber properties of alginate/hydroxyethyl cellulose with varied proportion for potential use in tendon tissue engineering.

Researchers have examined different bio-inspired materials in tissue engineering and regenerative medicine to fabricate scaffolds to address tendon regeneration requirements. We developed fibers based on alginate (Alg) and hydroxyethyl cellulose (HEC) by wet-spinning technique to mimic the fibrous sheath of ECM. Various proportions (25:75, 50:50, 75:25) of 1 % Alg and 4 % HEC were blended to this aim. Two steps of crosslinking with different concentrations of CaCl2 (2.5 and 5 %) and glutaraldehyde (2.5 %) were used to improve physical and mechanical properties. The fibers were characterized by FTIR, SEM, swelling, degradation, and tensile tests. The in vitro proliferation, viability, and migration of tenocytes on the fibers were also evaluated. Moreover, the biocompatibility of implanted fibers was investigated in an animal model. The results showed ionic and covalent molecular interactions between the components. In addition, by properly maintaining surface morphology, fiber alignment, and swelling, lower concentrations of HEC in the blending provided good degradability and mechanical features. The mechanical strength of fibers was in the range of collagenous fibers. Increasing the crosslinking led to significantly different mechanical behaviors in terms of tensile strength and elongation at break. Because of good in vitro and in vivo biocompatibility, tenocyte proliferation, and migration, the biological macromolecular fibers could serve as desirable tendon substitutes. This study provides more practical insight into tendon tissue engineering in translational medicine.

The Cross-Talks Among Bone Morphogenetic Protein (BMP) Signaling and Other Prominent Pathways Involved in Neural Differentiation.

The bone morphogenetic proteins (BMPs) are a group of potent morphogens which are critical for the patterning, development, and function of the central nervous system. The appropriate function of the BMP pathway depends on its interaction with other signaling pathways involved in neural differentiation, leading to synergistic or antagonistic effects and ultimately favorable biological outcomes. These opposite or cooperative effects are observed when BMP interacts with fibroblast growth factor (FGF), cytokines, Notch, Sonic Hedgehog (Shh), and Wnt pathways to regulate the impact of BMP-induced signaling in neural differentiation. Herein, we review the cross-talk between BMP signaling and the prominent signaling pathways involved in neural differentiation, emphasizing the underlying basic molecular mechanisms regarding the process of neural differentiation. Knowing these cross-talks can help us to develop new approaches in regenerative medicine and stem cell based therapy. Recently, cell therapy has received significant attention as a promising treatment for traumatic or neurodegenerative diseases. Therefore, it is important to know the signaling pathways involved in stem cell differentiation toward neural cells. Our better insight into the cross-talk of signaling pathways during neural development would improve neural differentiation within in vitro tissue engineering approaches and pre-clinical practices and develop futuristic therapeutic strategies for patients with neurological disease.

Recent advances on small molecules in osteogenic differentiation of stem cells and the underlying signaling pathways.

Bone-related diseases are major contributors to morbidity and mortality in elderly people and the current treatments result in insufficient healing and several complications. One of the promising areas of research for healing bone fractures and skeletal defects is regenerative medicine using stem cells. Differentiating stem cells using agents that shift cell development towards the preferred lineage requires activation of certain intracellular signaling pathways, many of which are known to induce osteogenesis during embryological stages. Imitating embryological bone formation through activation of these signaling pathways has been the focus of many osteogenic studies. Activation of osteogenic signaling can be done by using small molecules. Several of these agents, e.g., statins, metformin, adenosine, and dexamethasone have other clinical uses but have also shown osteogenic capacities. On the other hand, some other molecules such as T63 and tetrahydroquinolines are not as well recognized in the clinic. Osteogenic small molecules exert their effects through the activation of signaling pathways known to be related to osteogenesis. These pathways include more well-known pathways including BMP/Smad, Wnt, and Hedgehog as well as ancillary pathways including estrogen signaling and neuropeptide signaling. In this paper, we review the recent data on small molecule-mediated osteogenic differentiation, possible adjunctive agents with these molecules, and the signaling pathways through which each small molecule exerts its effects.

Expert consensus on the monitoring and treatment of sepsis-induced immunosuppression.

Emerged evidence has indicated that immunosuppression is involved in the occurrence and development of sepsis. To provide clinical practice recommendations on the immune function in sepsis, an expert consensus focusing on the monitoring and treatment of sepsis-induced immunosuppression was developed. Literature related to the immune monitoring and treatment of sepsis were retrieved from PubMed, Web of Science, and Chinese National Knowledge Infrastructure to design items and expert opinions were collected through an online questionnaire. Then, the Delphi method was used to form consensus opinions, and RAND appropriateness method was developed to provide consistency evaluation and recommendation levels for consensus opinions. This consensus achieved satisfactory results through two rounds of questionnaire survey, with 2 statements rated as perfect consistency, 13 as very good consistency, and 9 as good consistency. After summarizing the results, a total of 14 strong recommended opinions, 8 weak recommended opinions and 2 non-recommended opinions were produced. Finally, a face-to-face discussion of the consensus opinions was performed through an online meeting, and all judges unanimously agreed on the content of this consensus. In summary, this expert consensus provides a preliminary guidance for the monitoring and treatment of immunosuppression in patients with sepsis.

Thromboelastometry (TEM) findings in disseminated intravascular coagulation in a pig model of endotoxinemia.

Standard coagulation tests have a low specificity and sensitivity for diagnosing disseminated intravascular coagulation. The aim of this study was to determine whether whole blood thromboelastometry (TEM) detects lipopolysaccharide (LPS)-induced changes in coagulation. Blood samples from 10 pigs were drawn at baseline, before and at the end of LPS infusion and 2, 3, 4 and 5 h after the start of endotoxinemia. Simultaneous to TEM, standard coagulation tests and extended coagulation analysis including tissue plasminogen activator (t-PA) and plasminogen activator inhibitor 1 (PAI-1) were performed. Endotoxinemia resulted in a significant acceleration of the nonactivated TEM (NATEM) clotting time 2 h after the end of LPS infusion; in contrast, the changes in international normalized ratio and activated partial thromboplastin time suggested delayed initiation of coagulation. NATEM maximum clot firmness (MCF) and fibrin-based thromboelastometry test (FIBTEM)-MCF decreased significantly from baseline until the last time point (from 64.6 ± 7.8 and 35.1 ± 12.8 mm to 52.8 ± 4.6 and 21.4 ± 11.8 mm, respectively; P = 0.01 for both parameters). A sharp, transient increase of t-PA had no effect on maximum lysis in the NATEM test. PAI-1 increased significantly 3 h after the start of LPS infusion, paralleled by a decrease in maximum lysis. In conclusion, TEM was superior to standard coagulation tests in reflecting initial activation of coagulation during endotoxinemia. TEM further suggested consumption of coagulation substrate; at the same time, inhibition of plasminogen activation was accompanied by improved clot stability. Further investigations are necessary to establish the clinical relevance of these findings.

Experimentally approaching the ICU: monitoring outcome-based responses in the two-hit mouse model of posttraumatic sepsis.

To simulate and monitor the evolution of posttraumatic sepsis in mice, we combined a two-hit model of trauma/hemorrhage (TH) followed by polymicrobial sepsis with repetitive blood sampling. Anesthetized mice underwent femur fracture/sublethal hemorrhage and cecal ligation and puncture (CLP) 48 h later. To monitor outcome-dependent changes in circulating cells/biomarkers, mice were sampled daily (facial vein) for 7 days and retrospectively divided into either dead (DIE) or surviving (SUR) by post-CLP day 7. Prior to CLP, AST was 3-fold higher in DIE, while all other post-TH changes were similar between groups. There was a significant post-CLP intergroup separation. In SUR, RBC and Hb were lower, platelets and neutrophils higher, and lymphocytes mixed compared to DIE. In DIE, all organ function markers except glucose (decrease) were few folds higher compared to SUR. In summary, the combination of daily monitoring with an adequate two-hit model simulates the ICU setting, allows insight into outcome-based responses, and can identify biomarkers indicative of death in the acute posttraumatic sepsis in mice.