Myeloid-derived suppressor cells (MDSCs) expand during mouse and human sepsis, but the mechanism responsible for this is unclear. We previously reported that nuclear transport of S100A9 protein programs Gr1CD11b myeloid precursors into MDSCs in septic mice. Here, we show that long non-coding RNA Hotairm1 converts MDSCs from an activator to a repressor state. Mechanistically, increased Hotairm1 expression in MDSCs in mice converted S100A9 from a secreted proinflammatory mediator to an immune repressor by binding to and shuttling it from cytosol to nucleus during late sepsis. High Hotairm1 levels were detected in exosomes shed from MDSCs from late septic mice. These exosomes inhibited lipopolysaccharide-stimulated secretion of S100A9 from early sepsis Gr1CD11b cells. Importantly, Hotairm1 knockdown in late sepsis Gr1CD11b MDSCs prevented S100A9 cytosol to nuclear transfer and decreased repression of proimmune T cells. Notably, ectopic expression of Hotairm1 in early sepsis Gr1CD11b cells shuttled S100A9 to the nucleus and promoted the MDSC repressor phenotype. In support of translating the mechanistic concept to human sepsis, we found that Hotairm1 binds S100A9 protein in CD33CD11bHLA-DR MDSCs during established sepsis. Together, these data support that Hotairm1 is a plausible molecular target for treating late sepsis immune suppression in humans and its immune repressor mechanism may be cell autonomous.

Sepsis, a life-threatening response to infection, represents a substantial global health concern. Each year, about 48,9 million people are affected by sepsis. While survival rates have improved, sepsis survivors often experience numerous challenges after discharge from the hospital. Therefore, the aim of this study was to describe patients’ experience of their life situation after surviving sepsis. A literature review was carried out using Friberg’s four step model for analyzing quantitative and qualitative research. The data were collected from seven quantitative and four qualitative studies. The analysis revealed two themes and eight subthemes: (1) Difficulties adjusting to new living conditions, (2) A challenging recovery process. The first theme had four subthemes: Changed quality of life after sepsis, Physical limitations in everyday life, The challenge of becoming dependent on others and The importance of social relationships. The second theme had four subthemes: A mental struggle to return to a normal life, Traumatization and fears of relapse, The importance of recovery, and Educational and informational gaps. Among sepsis survivors, various problems and challenges have been identified that impact their daily life. Numerous survivors suffered from persistent residual symptoms primarily related to physical, psychological, and cognitive domains. Survivors were widely dissatisfied with the information and education about sepsis and its aftermath provided by the healthcare system. Further research is needed to clarify the long-term consequences of sepsis survivorship, and to improve the healthcare guidelines and interventions aimed at improving the well-being of this patient group

Inflammation is a crucial physiological defense mechanism of the human body to injury or infection. However, dysregulation of the magnitude or duration of inflammation response underlies multiple disease pathologies and may cause organ damage. Sepsis is a severe inflammatory disease now known as a clinical syndrome defined as life-threatening organ dysfunction caused by a dysregulated host response to infection. Sepsis patients often die of organ failure and the endothelium and neutrophil-endothelial cell (EC) interactions play an active role in the regulation of the systemic inflammatory response. Systemic inflammatory disease often results in alterations in vascular endothelium barrier function, increased permeability, excessive leukocyte trafficking, and reactive oxygen species production, leading to organ damage. While neutrophils are critical to host defense, neutrophil dysregulation has a critical role in organ damage through release of proteases, neutrophil extracellular traps (NETs), and reactive oxygen species (ROS), which can damage host tissue leading to organ failure. To date therapeutic approaches are largely supportive and therapeutics targeting endothelium inflammation and immune cell dysregulation are urgently needed. However, strong concerns regarding the level of phenotypic heterogeneity of microvascular ECs between different organs have been expressed. Microvascular EC heterogeneity in different organs and organ-specific variations in EC structure and function are regulated by intrinsic signals that are differentially expressed across organs and species, as a result of which neutrophil recruitment to discrete organs may be regulated differently. In addition, therapeutic development is hindered due to the heterogeneous nature of sepsis and the presence of multiple distinct immune phenotypes that can impact function and response to infection. In fact, clinically sepsis is a heterogeneous syndrome and diagnosis is complicated due to the broad spectrum of non-specific clinical features. Patients with similar clinical symptoms can be associated with distinct immune cell phenotypes ranging from excessive immune activation to immunosuppression, which means different therapeutics are required. In this work, the morphological and functional variations of differently originated microvascular endothelium are discussed and how these variances affect systemic function in response to inflammation. Emerging in vivo and in vitro models and techniques including microphysiological devices, proteomics, and RNA-Sequencing used to study the cellular and molecular heterogeneity of endothelium from different organs will also be discussed. Our group have developed a novel Organ-on-Chip, the biomimetic microfluidic assay (bMFA) that mimics physiological conditions, allowing us to observe real-time neutrophil-endothelial interactions, including rolling, adhesion, and migration, and to study endothelial barrier function under physiologically relevant conditions including the effect of shear forces and vascular geometry. The bMFA enables the quantification of leukocyte-EC interactions, including rolling velocity, number of adhered leukocytes in response to different shear rates, number of migrated leukocytes, EC permeability, adhesion molecule expression and other important variables. Furthermore, by using human related samples, such as human ECs and leukocytes, bMFA provides a tool for rapid screening of potential therapeutics to increase their clinical translatability. In this work, a protocol was developed to study endothelium function and neutrophil-endothelial interactions during inflammation in the bMFA. Lastly, to develop targeted therapeutics, immunophenotyping is needed to identify distinct immune cell functional phenotypes. We have developed a methodology to classify ICU sepsis patients into three phenotypes using patient data, Organ-on-Chip-based neutrophil functional analysis and proteomics. The findings of the study will help identify different sepsis patient immune-phenotypes and personalize treatment accordingly. / Mechanical Engineering

Interleukin-6 (IL-6) and C-reactive protein (CRP) are being used for diagnosis of sepsis. However, studies have reported varying cut-off levels and diagnostic performance. This study aims to investigate the optimal cut-off levels and performance of IL-6 and CRP for the diagnosis of neonatal sepsis. The study was conducted at the University Hospital of Leipzig, Germany from November 2012 to June 2020. A total of 899 neonates: 104 culture proven sepsis, 160 clinical sepsis, and 625 controls were included. Blood culture was performed using BacT/ALERT 3D system. IL-6 and CRP were analyzed by electrochemiluminescent immunoassay and immunoturbidimetric assay, respectively. Data were analyzed using SPSS 20 statistical software. Among neonates with proven sepsis, the optimal cut-off value of IL-6 was 313.5 pg/mL. The optimal cut-off values for CRP in 5 days serial measurements (CRP1, CRP2, CRP3, CRP4, and CRP5) were 2.15 mg/L, 8.01 mg/L, 6.80 mg/L, 5.25 mg/L, and 3.72 mg/L, respectively. IL-6 showed 73.1% sensitivity, 80.2% specificity, 37.6% PPV, and 94.8% NPV. The highest performance of CRP was observed in the second day with 89.4% sensitivity, 97.3% specificity, 94.5% PPV, and 98.3% NPV. The combination of IL-6 and CRP showed increase in sensitivity with decrease in specificity. In conclusion, this study defines the optimal cut-off values for IL-6 and CRP. The combination of IL-6 and CRP demonstrated increased sensitivity. The CRP 2 at cut-off 8.01 mg/L showed the highest diagnostic performance for identification of culture negative clinical sepsis cases. We recommend the combination of IL-6 (≥313.5 pg/mL) and CRP1 (≥2.15 mg/L) or IL-6 (≥313.5 pg/mL) and CRP2 (≥8.01 mg/L) for early and accurate diagnosis of neonatal sepsis. The recommendation is based on increased sensitivity, that is, to minimize the risk of any missing cases of sepsis. The CRP2 alone at cut-off 8.01 mg/L might be used to identify clinical sepsis cases among culture negative sepsis suspected neonates in hospital settings.

Neonatal sepsis caused by resistant bacteria is a worldwide concern due to the associated high mortality and increased hospitals costs. Bacterial pathogens causing neonatal sepsis and their antibiotic resistance patterns vary among hospital settings and at different points in time. This study aimed to determine the antibiotic resistance patterns of pathogens causing neonatal sepsis and to assess trends in antibiotic resistance. The study was conducted among neonates with culture proven sepsis at the University Hospital of Leipzig between November 2012 and September 2020. Blood culture was performed by BacT/ALERT 3D system. Antimicrobial susceptibility testing was done with broth microdilution method based on ISO 20776-1 guideline. Data were analyzed by SPSS version 20 software. From 134 isolates, 99 (74%) were gram positive bacteria. The most common gram positive and gram negative bacteria were S. epidermidis, 51 (38%) and E. coli, 23 (17%), respectively. S. epidermidis showed the highest resistance to penicillin G and roxithromycin (90% each) followed by cefotaxime, cefuroxime, imipenem, oxacillin, and piperacillin-tazobactam (88% each), ampicillin-sulbactam (87%), meropenem (86%), and gentamicin (59%). Moreover, S. epidermidis showed raising levels of resistance to amikacin, gentamicin, ciprofloxacin, levofloxacin, moxifloxacin, and cotrimoxazol. Gram positive bacteria showed less or no resistance to daptomycin, linezolid, teicoplanin, and vancomycin. E. coli showed the highest resistance to ampicillin (74%) followed by ampicillin-sulbactam (52%) and piperacillin (48%). Furthermore, increasing levels in resistance to ampicillin, ampicillin-sulbactam, piperacillin, and cefuroxime were observed over the years. Encouragingly, E. coli showed significantly declining trends of resistance to ciprofloxacin and levofloxacin, and no resistance to amikacin, colistin, fosfomycin, gentamicin, imipenem, piperacillin-tazobactam, and tobramycin. In conclusion, this study demonstrates that gram positive bacteria were the leading causes of neonatal sepsis. Bacterial isolates were highly resistant to first and second-line empiric antibiotics used in this hospital. The high levels of antibiotic resistance patterns highlight the need for modifying empiric treatment regimens considering the most effective antibiotics. Periodic surveillance in hospital settings to monitor changes in pathogens, and antibiotic resistance patterns is crucial in order to implement optimal prevention and treatment strategies.

Identifying antibiotic resistance in blood infections requires separating bacteria from whole blood. A hollow spinning disk rapidly removes suspended red blood cells by leveraging hydrodynamic differences between bacteria and whole blood components in a centrifugal field. Once the red cells are removed, the supernatant plasma which contains bacteria is collected for downstream antibiotic testing. This work improves upon previous work by modifying the disk design to maximize fractional plasma recovery and minimize fractional red cell recovery. V-shaped channels induce plasma flow and increase fractional plasma recovery. Additionally, diluting a blood sample spiked with bacteria prior to spinning it increased the fractional bacteria recovery. A numerical model for red cell sedimentation shows that red cells are removed from solution more rapidly as the blood is diluted. Diluting blood is beneficial but may create too much biological waste. The benefits of diluting are formulated as an optimization problem subject to the end user’s needs.

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