The pathophysiology of sepsis and precision-medicine-based immunotherapy.

Sepsis remains a major cause of morbidity and mortality in both low- and high-income countries. Antibiotic therapy and supportive care have significantly improved survival following sepsis in the twentieth century, but further progress has been challenging. Immunotherapy trials for sepsis, mainly aimed at suppressing the immune response, from the 1990s and 2000s, have largely failed, in part owing to unresolved patient heterogeneity in the underlying immune disbalance. The past decade has brought the promise to break this blockade through technological developments based on omics-based technologies and systems medicine that can provide a much larger data space to describe in greater detail the immune endotypes in sepsis. Patient stratification opens new avenues towards precision medicine approaches that aim to apply immunotherapies to sepsis, on the basis of precise biomarkers and molecular mechanisms defining specific immune endotypes. This approach has the potential to lead to the establishment of immunotherapy as a successful pillar in the treatment of sepsis for future generations.

B-cell immune deficiency in twin sisters expands the phenotype of MOPDI.

Microcephalic osteodysplastic primordial dwarfism type I (MOPDI) is a very rare and severe autosomal recessive disorder characterized by marked intrauterine growth retardation, skeletal dysplasia, microcephaly and brain malformations. MOPDI is caused by biallelic mutations in RNU4ATAC, a non-coding gene involved in U12-type splicing of 1% of the introns in the genome, which are recognized by their specific splicing consensus sequences. Here, we describe a unique observation of immunodeficiency in twin sisters with mild MOPDI, who harbor a novel n.108_126del mutation, encompassing part of the U4atac snRNA 3' stem-loop and Sm protein binding site, and the previously reported n.111G>A mutation. Interestingly, both twin sisters show mild B-cell anomalies, including low naive B-cell counts and increased memory B-cell and plasmablasts counts, suggesting partial and transitory blockage of B-cell maturation and/or excessive activation of naive B-cells. Hence, the localization of a mutation in stem II of U4atac snRNA, as observed in another RNU4ATAC-opathy with immunodeficiency, that is, Roifman syndrome (RFMN), is not required for the occurrence of an immune deficiency. Finally, we emphasize the importance of considering immunodeficiency in MOPDI management to reduce the risk of serious infectious episodes.

Interferon-Gamma-Release assay and absolute CD8 lymphocyte count for acquired immunosuppression monitoring in critically ill patients.

Guided biomarker-personalized immunotherapy is advancing rapidly as a means to rejuvenate immune function in injured patients who are the most immunosuppressed. A recent study introduced a fully automated interferon-γ release assay (IGRA) for monitoring the functionality of T lymphocytes in patients with septic shock. While a significant decrease in IFN-γ release capacity was observed, a significant correlation with CD8 lymphocyte absolute count was also reported, raising the question of whether ex-vivo IFN-γ production would be only a surrogate marker for lymphocyte count or if these two parameters conveyed distinct and complementary information. In a large cohort of more than 353 critically ill patients following various injuries (sepsis, trauma, major surgery), the primary objective of the present study was to simultaneously evaluate the association between ex vivo IFN-γ release and CD8 cell count with regard to adverse outcome. Our findings provide a clear-cut result, as they distinctly demonstrate that IGRA offers higher-quality information than CD8 count in terms of an independent association with the occurrence of an adverse outcome. These results strengthen the case for incorporating IGRA into the array of biomarkers of interest for defining endotypes in sepsis. This holds especially true given that fully automated tests are now readily available and could be used in routine clinical practice.

Tacrolimus Monitoring in Liver Transplant Recipients, Posttransplant Cholestasis: A Comparative Between 2 Commercial Immunoassays and a Liquid Chromatography-Tandem Mass Spectrometry Method.

BACKGROUND: Cholestasis commonly occurs after orthotopic liver transplantation. It can be extrahepatic because of mechanical obstruction or intrahepatic because of various causes. During cholestasis episodes, blood concentrations of tacrolimus (TAC) metabolites may increase, potentially affecting TAC concentrations measured by immunoassays. This study aimed to simultaneously evaluate the analytical performance of 2 TAC immunoassays, a quantitative microsphere system (QMS) immunoassay, and chemiluminescence microparticle immunoassay, using liquid chromatography-tandem mass spectrometry (LC-MS/MS) as a reference method in liver transplant recipients. METHODS: This single-center study included 265 patients who underwent orthotopic liver transplantation. In total, 942 blood samples were collected. TAC trough concentrations were measured using LC-MS/MS and 2 immunoassays in parallel. The plasma concentrations of conjugated bilirubin were measured in all samples. The results were analyzed using Bland-Altman plots and Passing-Bablok regressions. RESULTS: The Bland-Altman plot analysis showed that the TAC QMS immunoassay has a significant bias (+37%) compared with LC-MS/MS, and this bias was higher in patients with cholestasis with hyperbilirubinemia (≤+70% in patients with conjugated bilirubin >150 µmol/L). In comparison, the chemiluminescence microparticle immunoassay showed acceptable analytical performance in patients with hyperbilirubinemia (bias <10%). CONCLUSIONS: In agreement with previous findings, the TAC QMS immunoassay showed a positive bias compared with LC-MS/MS. This bias is remarkably high in patients with cholestasis and hyperbilirubinemia, suggesting the cross-reactivity of TAC metabolites with the monoclonal antibody used in the QMS immunoassay.

Immune profiling of critically ill patients with acute kidney injury during the first week after various types of injuries: the REALAKI study.

BACKGROUND: Acute kidney injury (AKI) is common in hospitalized patients and results in significant morbidity and mortality. The objective of the study was to explore the systemic immune response of intensive care unit patients presenting with AKI, especially the association between immune profiles and persistent AKI during the first week after admission following various types of injuries (sepsis, trauma, surgery, and burns). METHODS: REALAKI is an ancillary analysis of the REAnimation Low Immune Status Marker (REALISM) cohort study, in which 359 critically ill patients were enrolled in three different intensive care units. Patients with end-stage renal disease were excluded from the REALAKI study. Clinical samples and data were collected three times after admission: at day 1 or 2 (D1-2), day 3 or 4 (D3-4) and day 5, 6 or 7 (D5-7). Immune profiles were compared between patients presenting with or without AKI. Patients with AKI at both D1-2 and D5-7 were defined as persistent AKI. A multivariable logistic regression model was performed to determine the independent association between AKI and patients' immunological parameters. RESULTS: Three hundred and fifty-nine patients were included in this analysis. Among them, 137 (38%) were trauma patients, 103 (29%) post-surgery patients, 95 (26%) sepsis patients, and 24 (7%) were burn patients. One hundred and thirty-nine (39%) patients presented with AKI at D1-2 and 61 (20%) at D5-7. Overall, 94% presented with persistent AKI at D5-7. Patients with AKI presented with increased pro and anti-inflammatory cytokines and altered innate and adaptive immune responses. The modifications observed in the immune profiles tended to be more pronounced with increasing KDIGO stages. In the logistic regression model, a statistically significant association was observed at D1-2 between AKI and CD10lowCD16low immature neutrophils (OR 3.03 [1.7-5.5]-p < 0.001). At D5-7, increased interleukin-10 (IL-10) levels and reduced ex vivo TNF-α production after LPS stimulation were significantly associated with the presence of AKI (OR 1.38 [1.12-1.71]-p = 0.001 and 0.51 [0.27-0.91]-p = 0.03, respectively). Patients who recovered from AKI between D1-2 and D5-7 compared to patients with persistent AKI at D5-7, tended to correct these alterations. CONCLUSION: Following various types of severe injuries, early AKI is associated with the initial inflammatory response. Presence of AKI at the end of the first week after injury is associated with injury-induced immunosuppression.

Integrated clustering of multiple immune marker trajectories reveals different immunotypes in severely injured patients.

BACKGROUND: The immune response of critically ill patients, such as those with sepsis, severe trauma, or major surgery, is heterogeneous and dynamic, but its characterization and impact on outcomes are poorly understood. Until now, the primary challenge in advancing our understanding of the disease has been to concurrently address both multiparametric and temporal aspects. METHODS: We used a clustering method to identify distinct groups of patients, based on various immune marker trajectories during the first week after admission to ICU. In 339 severely injured patients, we initially longitudinally clustered common biomarkers (both soluble and cellular parameters), whose variations are well-established during the immunosuppressive phase of sepsis. We then applied this multi-trajectory clustering using markers composed of whole blood immune-related mRNA. RESULTS: We found that both sets of markers revealed two immunotypes, one of which was associated with worse outcomes, such as increased risk of hospital-acquired infection and mortality, and prolonged hospital stays. This immunotype showed signs of both hyperinflammation and immunosuppression, which persisted over time. CONCLUSION: Our study suggest that the immune system of critically ill patients can be characterized by two distinct longitudinal immunotypes, one of which included patients with a persistently dysregulated and impaired immune response. This work confirms the relevance of such methodology to stratify patients and pave the way for further studies using markers indicative of potential immunomodulatory drug targets.

Immune Profiling Panel Gene Set Identifies Critically Ill Patients With Low Monocyte Human Leukocyte Antigen-DR Expression: Preliminary Results From the REAnimation Low Immune Status Marker (REALISM) Study.

OBJECTIVES: There is a crucial unmet need for biomarker-guided diagnostic and prognostic enrichment in clinical trials evaluating immune modulating therapies in critically ill patients. Low monocyte expression of human leukocyte antigen-DR (mHLA-DR), considered as a reference surrogate to identify immunosuppressed patients, has been proposed for patient stratification in immunostimulation approaches. However, its widespread use in clinic has been somewhat hampered by technical constraints inherent to flow cytometry technology. The objective of the present study was to evaluate the ability of a prototype multiplex polymerase chain reaction tool (immune profiling panel [IPP]) to identify immunosuppressed ICU patients characterized by a low mHLA-DR expression. DESIGN: Retrospective observational cohort study. SETTING: Adult ICU in a University Hospital, Lyon, France. PATIENTS: Critically ill patients with various etiologies enrolled in the REAnimation Low Immune Status Marker study (NCT02638779). INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: mHLA-DR and IPP data were obtained from 1,731 blood samples collected from critically ill patients with various etiologies and healthy volunteers. A partial least square regression model combining the expression levels of IPP markers was trained and used for the identification of samples from patients presenting with evidence of immunosuppression, defined here as mHLADR less than 8,000 antibodies bound per cell (AB/C). The IPP gene set had an area under the receiver operating characteristic curve (AUC) of 0.86 (95% CI 0.83-0.89) for the identification of immunosuppressed patients. In addition, when applied to the 123 patients still in the ICU at days 5-7 after admission, IPP similarly enriched the number of patients with ICU-acquired infections in the immunosuppressed group (26%), in comparison with low mHLA-DR (22%). CONCLUSIONS: This study reports on the potential of the IPP gene set to identify ICU patients presenting with mHLA-DR less than 8,000 AB/C. Upon further optimization and validation, this molecular tool may help in the stratification of patients that could benefit from immunostimulation in the context of personalized medicine.

Fully automated interferon-γ release assay to monitor antigen-independent T cell functionality: A proof of concept study in septic shock.

In sepsis, personalized immunotherapy is being evaluated as a means of restoring immune function in the most severely affected patients. Biomarkers play a crucial role in this process, as there are no clear clinical indicators of immune dysfunction. Functional testing is considered a gold standard for assessing immune function, but this approach faces analytical challenges in terms of clinical implementation. The use of technician-dependent, time-consuming, home-made protocols often leads to poor standardization. This study represents the first beta testing of a fully automated interferon-γ release assay (IGRA) for monitoring the functionality of antigen-independent T lymphocytes. We observed a significant decrease in IFN-γ release capacity, which was associated with typical alterations in immunological cellular parameters (such as low mHLA-DR expression and decreased CD8 T lymphocyte count), in 22 patients with septic shock. Since the test is performed using whole blood and requires no technician intervention, with results available within 4 h, it may offer new possibilities for monitoring patients with immune alterations in routine clinical conditions. Further investigations in larger cohorts of patients are now needed to validate its clinical potential.

The double sides of NLRP3 inflammasome activation in sepsis.

Sepsis is defined as a life-threatening organ dysfunction induced by a dysregulated host immune response to infection. Immune response induced by sepsis is complex and dynamic. It is schematically described as an early dysregulated systemic inflammatory response leading to organ failures and early deaths, followed by the development of persistent immune alterations affecting both the innate and adaptive immune responses associated with increased risk of secondary infections, viral reactivations, and late mortality. In this review, we will focus on the role of NACHT, leucin-rich repeat and pyrin-containing protein 3 (NLRP3) inflammasome in the pathophysiology of sepsis. NLRP3 inflammasome is a multiproteic intracellular complex activated by infectious pathogens through a two-step process resulting in the release of the pro-inflammatory cytokines IL-1ß and IL-18 and the formation of membrane pores by gasdermin D, inducing a pro-inflammatory form of cell death called pyroptosis. The role of NLRP3 inflammasome in the pathophysiology of sepsis can be ambivalent. Indeed, although it might protect against sepsis when moderately activated after initial infection, excessive NLRP3 inflammasome activation can induce dysregulated inflammation leading to multiple organ failure and death during the acute phase of the disease. Moreover, this activation might become exhausted and contribute to post-septic immunosuppression, driving impaired functions of innate and adaptive immune cells. Targeting the NLRP3 inflammasome could thus be an attractive option in sepsis either through IL-1ß and IL-18 antagonists or through inhibition of NLRP3 inflammasome pathway downstream components. Available treatments and results of first clinical trials will be discussed.

Identification of a sub-group of critically ill patients with high risk of intensive care unit-acquired infections and poor clinical course using a transcriptomic score.

BACKGROUND: The development of stratification tools based on the assessment of circulating mRNA of genes involved in the immune response is constrained by the heterogeneity of septic patients. The aim of this study is to develop a transcriptomic score based on a pragmatic combination of immune-related genes detected with a prototype multiplex PCR tool. METHODS: As training cohort, we used the gene expression dataset obtained from 176 critically ill patients enrolled in the REALISM study (NCT02638779) with various etiologies and still hospitalized in intensive care unit (ICU) at day 5-7. Based on the performances of each gene taken independently to identify patients developing ICU-acquired infections (ICU-AI) after day 5-7, we built an unweighted score assuming the independence of each gene. We then determined the performances of this score to identify a subgroup of patients at high risk to develop ICU-AI, and both longer ICU length of stay and mortality of this high-risk group were assessed. Finally, we validated the effectiveness of this score in a retrospective cohort of 257 septic patients. RESULTS: This transcriptomic score (TScore) enabled the identification of a high-risk group of patients (49%) with an increased rate of ICU-AI when compared to the low-risk group (49% vs. 4%, respectively), with longer ICU length of stay (13 days [95% CI 8-30] vs. 7 days [95% CI 6-9], p < 0.001) and higher ICU mortality (15% vs. 2%). High-risk patients exhibited biological features of immune suppression with low monocytic HLA-DR levels, higher immature neutrophils rates and higher IL10 concentrations. Using the TScore, we identified 160 high-risk patients (62%) in the validation cohort, with 30% of ICU-AI (vs. 18% in the low-risk group, p = 0.06), and significantly higher mortality and longer ICU length of stay. CONCLUSIONS: The transcriptomic score provides a useful and reliable companion diagnostic tool to further develop immune modulating drugs in sepsis in the context of personalized medicine.

Immune Profiling Demonstrates a Common Immune Signature of Delayed Acquired Immunodeficiency in Patients With Various Etiologies of Severe Injury.

OBJECTIVES: The host response plays a central role in the pathophysiology of sepsis and severe injuries. So far, no study has comprehensively described the overtime changes of the injury-induced immune profile in a large cohort of critically ill patients with different etiologies. DESIGN: Prospective observational cohort study. SETTING: Adult ICU in a University Hospital in Lyon, France. PATIENTS: Three hundred fifty-three septic, trauma, and surgical patients and 175 healthy volunteers were included in the REAnimation Low Immune Status Marker study. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Extensive immune profiling was performed by assessing cellular phenotypes and functions, protein, and messenger RNA levels at days 1-2, 3-4, and 5-7 after inclusion using a panel of 30 standardized immune markers. Using this immunomonitoring panel, no specificity in the immune profile was observed among septic, trauma, and surgical patients. This common injury-induced immune response was characterized by an initial adaptive (i.e., physiologic) response engaging all constituents of the immune system (pro- and anti-inflammatory cytokine releases, and innate and adaptive immune responses) but not associated with increased risk of secondary infections. In contrary, the persistence in a subgroup of patients of profound immune alterations at the end of the first week after admission was associated with increased risk of secondary infections independently of exposure to invasive devices. The combined monitoring of markers of pro-/anti-inflammatory, innate, and adaptive immune responses allowed a better enrichment of patients with risk of secondary infections in the selected population. CONCLUSIONS: Using REAnimation Low Immune Status Marker immunomonitoring panel, we detected delayed injury-acquired immunodeficiency in a subgroup of severely injured patients independently of primary disease. Critically ill patients' immune status could be captured through the combined monitoring of a common panel of complementary markers of pro-/anti-inflammatory, innate, and adaptive immune responses. Such immune monitoring needs to be incorporated in larger study cohorts with more extensive immune surveillance to develop specific hypothesis allowing for identification of biological systems affecting altered immune function related to late infection in the setting of acute systemic injury.

Pathogenic Mechanisms of Secondary Organic Aerosols.

Air pollution represents a major health problem and an economic burden. In recent years, advances in air pollution research has allowed particle fractionation and identification of secondary organic aerosol (SOA). SOA is formed from either biogenic or anthropogenic emissions, through a mass transfer from the gaseous mass to the particulate phase in the atmosphere. They can have deleterious impact on health and the mortality of individuals with chronic inflammatory diseases. The pleiotropic effects of SOA could involve different and interconnected pathogenic mechanisms ranging from oxidative stress, inflammation, and immune system dysfunction. The purpose of this review is to present recent findings about SOA pathogenic roles and potential underlying mechanisms focusing on the lungs; the latter being the primary exposed organ to atmospheric pollutants.

Monocyte CD169 expression in COVID-19 patients upon intensive care unit admission.

During the second surge of COVID-19 in France (fall 2020), we assessed the expression of monocyte CD169 (i.e., Siglec-1, one of the numerous IFN-stimulated genes) upon admission to intensive care units of 45 patients with RT-PCR-confirmed SARS-CoV2 pulmonary infection. Overall, CD169 expression was strongly induced on circulating monocytes of COVID-19 patients compared with healthy donors and patients with bacterial sepsis. Beyond its contribution at the emergency department, CD169 testing may be also helpful for patients' triage at the ICU to rapidly reinforce suspicion of COVID-19 etiology in patients with acute respiratory failure awaiting for PCR results for definitive diagnosis.

Clinical significance of a single cerebrospinal fluid immunoglobulin band: A retrospective study.

BACKGROUND: To demonstrate an inflammatory process in the central nervous system, the presence of at least two immunoglobulin (Ig) bands in the cerebrospinal fluid (CSF) is required. So far, the presence of a single abnormal Ig band is considered as negative. OBJECTIVE: The objective was to assess retrospectively the significance of a single CSF Ig band in clinical practice. METHODS AND RESULTS: Out of 10,286 CSF analyses, we retained 214 results with single Ig. An inflammatory neurological disorder was diagnosed in 41% of patients. CONCLUSION: Despite a modest sensitivity, the presence of a single CSF Ig band may be a biomarker of an inflammatory mechanism and, as such, may prompt the clinician to repeat the analysis when the clinical context remains suggestive.

Longitudinal assessment of IFN-I activity and immune profile in critically ill COVID-19 patients with acute respiratory distress syndrome.

BACKGROUND: Since the onset of the pandemic, only few studies focused on longitudinal immune monitoring in critically ill COVID-19 patients with acute respiratory distress syndrome (ARDS) whereas their hospital stay may last for several weeks. Consequently, the question of whether immune parameters may drive or associate with delayed unfavorable outcome in these critically ill patients remains unsolved. METHODS: We present a dynamic description of immuno-inflammatory derangements in 64 critically ill COVID-19 patients including plasma IFNα2 levels and IFN-stimulated genes (ISG) score measurements. RESULTS: ARDS patients presented with persistently decreased lymphocyte count and mHLA-DR expression and increased cytokine levels. Type-I IFN response was initially induced with elevation of IFNα2 levels and ISG score followed by a rapid decrease over time. Survivors and non-survivors presented with apparent common immune responses over the first 3 weeks after ICU admission mixing gradual return to normal values of cellular markers and progressive decrease of cytokines levels including IFNα2. Only plasma TNF-α presented with a slow increase over time and higher values in non-survivors compared with survivors. This paralleled with an extremely high occurrence of secondary infections in COVID-19 patients with ARDS. CONCLUSIONS: Occurrence of ARDS in response to SARS-CoV2 infection appears to be strongly associated with the intensity of immune alterations upon ICU admission of COVID-19 patients. In these critically ill patients, immune profile presents with similarities with the delayed step of immunosuppression described in bacterial sepsis.

Decreased Human Leukocyte Antigen DR on Circulating Monocytes Expression After Severe Pediatric Trauma: An Exploratory Report.

OBJECTIVES: Major trauma in adults induces immune dysfunction, with diminished expression of human leukocyte antigen-DR on circulating monocytes. No pediatric data are available. This study described the kinetics of human leukocyte antigen-DR on circulating monocytes following major pediatric trauma and relationships between human leukocyte antigen-DR on circulating monocytes and outcomes. DESIGN: Prospective observational study. SETTING: PICU and trauma unit at a tertiary-care university hospital in South Africa. PATIENTS: Children between 1 month and 13 years hospitalized for severe brain trauma or trauma with an Injury Severity Score greater than or equal to 16, from November 2016 to March 2017. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: We included 36 children. Median (interquartile range) age and Injury Severity Score were 7 years (4.9-10.5 yr) and 25 years (22.7-30 yr), respectively. Blood samples (n = 83) for standardized human leukocyte antigen-DR on circulating monocytes measurement were collected at days 1-2, 3-4, and 8-9 after injury (D1, D3, and D8, respectively). On D1, median (interquartile range) human leukocyte antigen-DR on circulating monocytes was markedly reduced relative to normal values (7,031 [5,204-11,201] antibodies per cell). There was a significant increase in human leukocyte antigen-DR on circulating monocytes from D1 to D8. Although all patients with secondary infections (n = 8; 22%) had human leukocyte antigen-DR on circulating monocytes less than 15,000 antibodies per cell at D3, human leukocyte antigen-DR on circulating monocytes levels were not associated with the occurrence of secondary infections (p = 0.22). At D3, human leukocyte antigen-DR on circulating monocytes was significantly higher in patients discharged home (n = 21) by Day 30 after trauma compared with those who died or were still hospitalized (n = 14) (p = 0.02). CONCLUSIONS: Pediatric severe trauma induced an early and dramatic decrease in human leukocyte antigen-DR on circulating monocytes expression. This alteration of innate immunity was not associated with the occurrence of secondary infection, possibly due to a lack of statistical power. However, human leukocyte antigen-DR on circulating monocytes at Day 3 is a potential indicator of those at high risk of secondary infection and worse outcomes.

Immune Profiling Panel: A Proof-of-Concept Study of a New Multiplex Molecular Tool to Assess the Immune Status of Critically Ill Patients.

BACKGROUND: Critical illness such as sepsis is a life-threatening syndrome defined as a dysregulated host response to infection and is characterized by patients exhibiting impaired immune response. In the field of diagnosis, a gap still remains in identifying the immune profile of critically ill patients in the intensive care unit (ICU). METHODS: A new multiplex immune profiling panel (IPP) prototype was assessed for its ability to semiquantify messenger RNA immune-related markers directly from blood, using the FilmArray System, in less than an hour. Samples from 30 healthy volunteers were used for the technical assessment of the IPP tool. Then the tool was clinically assessed using samples from 10 healthy volunteers and 20 septic shock patients stratified using human leukocyte antigen-DR expression on monocytes (mHLA-DR). RESULTS: The IPP prototype consists of 16 biomarkers that target the immune response. The majority of the assays had a linear expression with different RNA inputs and a coefficient of determination (R2) > 0.8. Results from the IPP pouch were comparable to standard quantitative polymerase chain reaction and the assays were within the limits of agreement in Bland-Altman analysis. Quantification cycle values of the target genes were normalized against reference genes and confirmed to account for the different cell count and technical variability. The clinical assessment of the IPP markers demonstrated various gene modulations that could distinctly differentiate 3 profiles: healthy volunteers, intermediate mHLA-DR septic shock patients, and low mHLA-DR septic shock patients. CONCLUSIONS: The use of IPP showed great potential for the development of a fully automated, rapid, and easy-to-use immune profiling tool. The IPP tool may be used in the future to stratify critically ill patients in the ICU according to their immune status. Such stratification will enable personalized management of patients and guide treatments to avoid secondary infections and lower mortality.

Monocyte HLA-DR Measurement by Flow Cytometry in COVID-19 Patients: An Interim Review.

Several months after the sudden emergence of SARS-CoV-2 and COVID-19, the understanding of the appropriate host immune response to a virus totally unknown of human immune surveillance is still of major importance. By international definition, COVID-19 falls in the scope of septic syndromes (organ dysfunction due to dysregulated host response to an infection) in which immunosuppression is a significant driver of mortality. Sepsis-induced immunosuppression is mostly defined and monitored by the measurement of decreased expression of HLA-DR molecules on circulating monocytes (mHLA-DR). In this interim review, we summarize the first mHLA-DR results in COVID-19 patients. In critically ill patients, results homogenously indicate a decreased mHLA-DR expression, which, along with profound lymphopenia and other functional alterations, is indicative of a status of immunosuppression. © 2020 International Society for Advancement of Cytometry.

Septic Shock Shapes B Cell Response toward an Exhausted-like/Immunoregulatory Profile in Patients.

Septic shock is accompanied by the development of immune dysfunctions whose intensity and duration are associated with increased risk of secondary infections and mortality. Although B lymphocytes play a pivotal role in the immune response to infections, no comprehensive exploration of circulating B cell status has been performed during the immunosuppressive phase of septic shock. Thus, our aim was to extensively characterize the phenotype and function of B cells in septic shock, including IL-10 production. Circulating B lymphocyte phenotype and function were evaluated by flow cytometry on fresh whole blood and after ex vivo stimulation in adult septic shock patients sampled at day 1, 3, and 6 after the onset of shock. The circulating B cell number was reduced in septic shock patients, whereas the B cell proportion among total lymphocytes was increased. The remaining circulating B lymphocytes presented with decreased MHC class II expression and increased CD21low CD95high exhausted-like phenotype but showed no change in maturation status. Circulating B cell functions were markedly altered after sepsis with reduced ex vivo activation and proliferation capacities. Finally, B cell response after septic shock was characterized by a clear plasmacytosis and an increased IL-10 production in remaining B cells from patients after ex vivo stimulation. During the sepsis-induced immunosuppression phase, B cell response is altered and is oriented toward an exhausted-like/immunoregulatory profile. Further studies are now needed to confirm the immunoregulatory properties of B lymphocytes and evaluate their role in sepsis-induced immunosuppression.

Monocytic HLA-DR expression kinetics in septic shock patients with different pathogens, sites of infection and adverse outcomes.

BACKGROUND: Decreased monocytic (m)HLA-DR expression is the most studied biomarker of sepsis-induced immunosuppression. To date, little is known about the relationship between sepsis characteristics, such as the site of infection, causative pathogen, or severity of disease, and mHLA-DR expression kinetics. METHODS: We evaluated mHLA-DR expression kinetics in 241 septic shock patients with different primary sites of infection and pathogens. Furthermore, we used unsupervised clustering analysis to identify mHLA-DR trajectories and evaluated their association with outcome parameters. RESULTS: No differences in mHLA-DR expression kinetics were found between groups of patients with different sites of infection (abdominal vs. respiratory, p = 0.13; abdominal vs. urinary tract, p = 0.53) and between pathogen categories (Gram-positive vs. Gram-negative, p = 0.54; Gram-positive vs. negative cultures, p = 0.84). The mHLA-DR expression kinetics differed between survivors and non-survivors (p < 0.001), with an increase over time in survivors only. Furthermore, we identified three mHLA-DR trajectories ('early improvers', 'delayed or non-improvers' and 'decliners'). The probability for adverse outcome (secondary infection or death) was higher in the delayed or non-improvers and decliners vs. the early improvers (delayed or non-improvers log-rank p = 0.03, adjusted hazard ratio 2.0 [95% CI 1.0-4.0], p = 0.057 and decliners log-rank p = 0.01, adjusted hazard ratio 2.8 [95% CI 1.1-7.1], p = 0.03). CONCLUSION: Sites of primary infection or causative pathogens are not associated with mHLA-DR expression kinetics in septic shock patients. However, patients showing delayed or no improvement in or a declining mHLA-DR expression have a higher risk for adverse outcome compared with patients exhibiting a swift increase in mHLA-DR expression. Our study signifies that changes in mHLA-DR expression over time, and not absolute values or static measurements, are of clinical importance in septic shock patients.