IntroductionSeptic syndromes remain the leading cause of mortality in intensive care units (ICU). Septic patients rapidly develop immune dysfunctions, the intensity and duration of which have been linked with deleterious outcomes. Decreased mRNA expressions of major histocompatibility complex (MHC) class II-related genes have been reported after sepsis. We investigated whether their mRNA levels in whole blood could predict mortality in septic shock patients.MethodsA total of 93 septic shock patients were included. On the third day after shock, the mRNA expressions of five MHC class II-related genes (CD74, HLA-DRA, HLA-DMB, HLA-DMA, CIITA) were measured by qRT-PCR and monocyte human leukocyte antigen-DR (mHLA-DR) by flow cytometry.ResultsA significant correlation was found among MHC class II related gene expressions. Among mRNA markers, the best prognostic value was obtained for CD74 (HLA-DR antigen-associated invariant chain). For this parameter, the area under the receiver operating characteristic curve (AUC) was calculated (AUC = 0.67, 95% confidence interval (CI) = 0.55 to 0.79; P = 0.01) as well as the optimal cut-off value. After stratification based on this threshold, survival curves showed that a decreased CD74 mRNA level was associated with increased mortality after septic shock (Log rank test, P = 0.0043, Hazard Ratio = 3.0, 95% CI: 1.4 to 6.5). Importantly, this association remained significant after multivariate logistic regression analysis including usual clinical confounders (that is, severity scores, P = 0.026, Odds Ratio = 3.4, 95% CI: 1.2 to 9.8).ConclusionDecreased CD74 mRNA expression significantly predicts 28-day mortality after septic shock. After validation in a larger multicentric study, this biomarker could become a robust predictor of death in septic patients.

IntroductionIt is now well accepted that sepsis is associated with the development of a pronounced immunosuppressive state, characterized by severe immune alterations (e.g. reduced proliferative capacity, endotoxin tolerance, apoptosis) participating in increased mortality and susceptibility to nosocomial infections. Efforts are currently aimed at restoring a functional immune response in septic patients. Successful therapy depends on the identification of appropriate immunostimulatory drugs and on the development of suitable biomarkers that could be used to stratify patients and to follow response to treatment.MethodsIn this study, we evaluated the ex vivo effect of recombinant interferon gamma (rIFN-γ) in restoring monocyte functionality (endotoxin-induced Tumor Necrosis Factor-α production) in a two-hit model of endotoxin tolerance (ET) with peripheral blood mononuclear cells from healthy volunteers and in whole blood of septic shock patients. Importantly, we used quantitative-reverse transcription polymerase-chain reaction to monitor the effect of rIFN-γ on the expression of seven genes known to participate in ET (TNF-α, IL-10, HLA-DRA, CIITA, IRAK-M, ABIN-3 and LY64).ResultsExpression analysis of those genes confirmed the presence of an immunosuppression state and the ex vivo restoration of immune functions by rIFN-γ. We show for the first time that rIFN-γ is able to bypass, at the mRNA level, the effect of negative regulators of the LPS signalling pathway such as IRAK-M, ABIN-3 and LY64.ConclusionsOverall, mRNA expressions of a panel of genes could represent promising candidates for the ex vivo evaluation of rIFN-γ effect on monocyte functionality. This ex vivo translational research study demonstrates the potential of a mRNA-based approach to successfully monitor drug efficacy.

BackgroundHuman Endogenous Retroviruses (HERVs) and Mammalian apparent LTR-retrotransposons (MaLRs) represent the 8% of our genome and are distributed among our 46 chromosomes. These LTR-retrotransposons are thought to be essentially silent except in cancer, autoimmunity and placental development. Their Long Terminal Repeats (LTRs) constitute putative promoter or polyA regulatory sequences. In this study, we used a recently described high-density microarray which can be used to study HERV/MaLR transcriptome including 353,994 HERV/MaLR loci and 1559 immunity-related genes.ResultsWe described, for the first time, the HERV transcriptome in peripheral blood mononuclear cells (PBMCs) using a cellular model mimicking inflammatory response and monocyte anergy observed after septic shock. About 5.6% of the HERV/MaLR repertoire is transcribed in PBMCs. Roughly one-tenth [5.7–13.1%] of LTRs exhibit a putative constitutive promoter or polyA function while one-quarter [19.5–27.6%] may shift from silent to active. Evidence was given that some HERVs/MaLRs and genes may share similar regulation control under lipopolysaccharide (LPS) stimulation conditions. Stimulus-dependent response confirms that HERV expression is tightly regulated in PBMCs. Altogether, these observations make it possible to integrate 62 HERVs/MaLRs and 26 genes in 11 canonical pathways and suggest a link between HERV expression and immune response. The transcriptional modulation of HERVs located close to genes such as OAS2/3 and IFI44/IFI44L or at a great distance from genes was discussed.ConclusionThis microarray-based approach revealed the expression of about 47,466 distinct HERV loci and identified 951 putative promoter LTRs and 744 putative polyA LTRs in PBMCs. HERV/MaLR expression was shown to be tightly modulated under several stimuli including high-dose and low-dose LPS and Interferon-γ (IFN-γ). HERV incorporation at the crossroads of immune response pathways paves the way for further functional studies and analyses of the HERV transcriptome in altered immune responses in vivo such as in sepsis.Electronic supplementary materialThe online version of this article (10.1186/s12864-018-4901-9) contains supplementary material, which is available to authorized users.

The rapid development in septic patients of features of marked immunosuppression associated with increased risk of nosocomial infections and mortality represents the rational for the initiation of immune targeted treatments in sepsis. However, as there is no clinical sign of immune dysfunctions, the current challenge is to develop biomarkers that will help clinicians identify the patients that would benefit from immunotherapy and monitor its efficacy. Using an in vitro model of endotoxin tolerance (ET), a pivotal feature of sepsis-induced immunosuppression in monocytes, we identified using gene expression profiling by microarray a panel of transcripts associated with the development of ET which expression was restored after immunostimulation with interferon-gamma (IFN-γ). These results were confirmed by qRT-PCR. Importantly, this short-list of markers was further evaluated in patients. Of these transcripts, six (TNFAIP6, FCN1, CXCL10, GBP1, CXCL5 and PID1) were differentially expressed in septic patients’ blood compared to healthy blood upon ex vivo LPS stimulation and were restored by IFN-γ. In this study, by combining a microarray approach in an in vitro model and a validation in clinical samples, we identified a panel of six new transcripts that could be used for the identification of septic patients eligible for IFNg therapy. Along with the previously identified markers TNFa, IL10 and HLA-DRA, the potential value of these markers should now be evaluated in a larger cohort of patients. Upon favorable results, they could serve as stratification tools prior to immunostimulatory treatment and to monitor drug efficacy.

Patients that suffer from sepsis exhibit an early hyper-inflammatory immune response which can lead to organ failure and death. In our study, we assessed the immune modulation in the human in vivo endotoxemia model and compared it to ex vivo LPS stimulation using 38 transcriptomic markers. Blood was collected before and after 4 hours of LPS challenge and tested with the Immune Profiling Panel (IPP) using the FilmArray system. The use of IPP showed that markers from the innate immunity dominated the response to LpS in vivo, mainly markers related to monocytes and neutrophils. Comparing the two models, in vivo and ex vivo, revealed that most of the markers were modulated in a similar pattern (68%). Some cytokine markers such as TNF, IFN-γ and IL-1β were under-expressed ex vivo compared to in vivo. T-cell markers were either unchanged or up-modulated ex vivo, compared to a down-modulation in vivo. Interestingly, markers related to neutrophils were expressed in opposite directions, which might be due to the presence of cell recruitment and feedback loops in vivo. the ipp tool was able to capture the early immune response in both the human in vivo endotoxemia model, a translational model mimicking the immune response observed in septic patients.

ObjectivesHospital-acquired infections (HAI) are associated with significant mortality and morbidity and prolongation of hospital stay, adding strain on limited hospital resources. Despite stringent infection control practices some children remain at high risk of developing HAI. The development of biomarkers which could identify these patients would be useful. In this study our objective was to evaluate mRNA candidate biomarkers for HAI prediction in a pediatric intensive care unit.DesignSerial blood samples were collected from patients admitted to pediatric intensive care unit between March and June 2012. Candidate gene expression (IL1B, TNF, IL10, CD3D, BCL2, BID) was quantified using RT-qPCR. Comparisons of relative gene expression between those that did not develop HAI versus those that did were performed using Mann Whitney U-test.PatientsExclusion criteria were: age <28 days or ≥16 years, expected length of stay < 24 hours, expected survival < 28 days, end-stage renal disease and end-stage liver disease. Finally, 45 children were included in this study.Main ResultsThe overall HAI rate was 30% of which 62% were respiratory infections. Children who developed HAI had a three-fold increase in hospital stay compared to those who did not (27 days versus 9 days, p<0.001). An increased expression of cytokine genes (IL1B and IL10) was observed in patients who developed HAI, as well as a pro-apoptosis pattern (higher expression of BID and lower expression of BCL2). CD3D, a key TCR co-factor was also significantly down-modulated in patients who developed HAI.ConclusionsTo our knowledge, this is the first study of mRNA biomarkers of HAI in the paediatric population. Increased mRNA expressions of anti-inflammatory cytokine and modulation of apoptotic genes suggest the development of immunosuppression in critically ill children. Immune monitoring using a panel of genes may offer a novel stratification tool to identify HAI risk.

Although human endogenous retroviruses (HERVs) expression is a growing subject of interest, no study focused before on specific endogenous retroviruses loci activation in severely injured patients. Yet, HERV reactivation is observed in immunity compromised settings like some cancers and auto-immune diseases. Our objective was to assess the transcriptional modulation of HERVs in burn, trauma and septic shock patients. We analyzed HERV transcriptome with microarray data from whole blood samples of a burn cohort (n = 30), a trauma cohort (n = 105) and 2 septic shock cohorts (n = 28, n = 51), and healthy volunteers (HV, n = 60). We described expression of the 337 probesets targeting HERV from U133 plus 2.0 microarray in each dataset and then we compared HERVs transcriptional modulation of patients compared to healthy volunteers. Although all 4 cohorts contained critically ill patients, the majority of the 337 HERVs was not expressed (around 74% in mean). Each cohort had differentially expressed probesets in patients compared to HV (from 19 to 46). Strikingly, 5 HERVs were in common in all types of severely injured patients, with 4 being up-modulated in patients. We highlighted co-expressed profiles between HERV and nearby CD55 and CD300LF genes as well as autonomous HERV expression. We suggest an inflammatory-specific HERV transcriptional response, and importantly, we introduce that the HERVs close to immunity-related genes might have a role on its expression.

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