HLJ1 amplifies endotoxin–induced sepsis severity by promoting IL-12 heterodimerization in macrophages

Heat shock protein (HSP) 40 has emerged as a key actor in both innate and adaptive immunity, whereas the role of HLJ1, a molecular chaperone in HSP40 family, in modulating endotoxin–induced sepsis severity is still unclear. Here, we use single-cell RNA sequencing to characterize mouse liver nonparenchymal cell populations under LPS (lipopolysaccharide) stimulation, and show that HLJ1 deletion affected IFN-γ-related gene signatures in distinct immune cell clusters. HLJ1 deficiency also leads to reduced serum levels of IL-12 in LPS-treated mice, contributing to dampened production of IFN-γ in natural killer cells but not CD4+ or CD8+ T cells, and subsequently to improved survival rate. Adoptive transfer of HLJ1-deleted macrophages into LPS-treated mice results in reduced IL-12 and IFN-γ levels and protects the mice from IFN-γ–dependent mortality. In the context of molecular mechanisms, HLJ1 is an LPS-inducible protein in macrophages and converts misfolded IL-12p35 homodimers to monomers, which maintains bioactive IL-12p70 heterodimerization and secretion. This study suggests HLJ1 causes IFN-γ–dependent septic lethality by promoting IL-12 heterodimerization, and targeting HLJ1 has therapeutic potential in inflammatory diseases involving activating IL-12/IFN-γ axis.

Vitamin D Deficiency among Children with Sepsis in a Tertiary Care Center in Eastern Nepal

INTRODUCTION: Vitamin D deficiency (VDD) is exceedingly predominant in children leading to dysregulation of the immune system and inflammation. Data on the prevalence of VDD in children with sepsis and its association with sepsis severity are limited from our part of the world. The primary aim of this study was to identify the burden of VDD in children with sepsis. MATERIAL AND METHODS: One hundred and five children (< 15 years) with sepsis were enrolled from April 15, 2017 to April 14, 2018 from a tertiary care center in Eastern Nepal. Demographic data including BMI, sequential organ failure assessment (SOFA) scores were recorded at the time of admission. Plasma 25-hydroxy vitamin D [25(OH)D] levels were measured by chemiluminescence immunoassay technique (CLIA) (MAGLUMI 25-OH Vitamin D; CLIA) within 24 hours of admission. Vitamin D concentrations of <20 ng/mL (50 nmol/L) were considered as deficient. RESULTS: Of the 105 children enrolled, the majority 74 (70.55%) had vitamin D deficiency. Vitamin D was deficient in 77, 65, and 66% of children in 1-5, 5-10, and 10-15 years of age group respectively. Vitamin D deficiency was maximum (80%) in underweight children. In the VDD group, 60% had severe sepsis, whereas only 32% had severe sepsis in vitamin D sufficient group with significant statistical association with sepsis severity and vitamin D deficiency. CONCLUSION: A high burden of VDD is present in children with sepsis which was found to be associated with greater severity of illness.

Neutrophil to Lymphocyte Ratio (NLR)—A Useful Tool for the Prognosis of Sepsis in the ICU

Sepsis is a life-threatening medical emergency induced by the body¢s extreme response to an infection. Despite well-defined and constantly updated criteria for diagnosing sepsis, it is still underdiagnosed worldwide. Among various markers studied over time, the neutrophil to lymphocyte ratio (NLR) recently emerged as a good marker to predict sepsis severity. Our study was a single-center prospective observational study performed in our ICU and included 114 patients admitted for sepsis or septic shock. Neutrophil to lymphocyte ratio (NLR) is easy to perform, CBC being one of the standard blood tests routinely performed upon admission for all ICU patients. We found that NLR was increased in all patients with sepsis and significantly raised in those with septic shock. NLR correlates significantly with sepsis severity evaluated by the SOFA score (R = 0.65) and also with extensively studied sepsis prognosis marker presepsin (R = 0.56). Additionally, NLR showed good sensitivity (47%) and specificity (78%) with AUC = 0.631 (p < 0.05). NLR is less expensive and easier to perform compared with other specific markers and may potentially become a good alternate option for evaluation of sepsis severity. Larger studies are needed in the future to demonstrate the prognosis value of NLR.

Proinflammatory Cytokine Levels in Sepsis and in Health and TNFα Association with Sepsis Mortality and patient characteristics: a Systematic Review and Meta-analysis

Background: Sepsis is a global health problem associated with significant morbidity and mortality. Detrimental sepsis effects are attributed to a "cytokine storm." However, anti-cytokine therapies have failed to lower sepsis mortality. We aim to characterize levels of key cytokines in sepsis patients and healthy controls and relate TNFα levels to patient characteristics and outcomes. Methods: We performed a systematic review and meta-analysis. Medline, Embase, Cochrane Library, and Web of Science Core Collection databases were searched from 1985 to May 2020 for studies in English. We included randomized controlled trials (RCTs), controlled trials, cohort studies, case series, and cross-sectional studies that reported mean levels of cytokines in the circulation thought to be relevant for sepsis pathogenesis. We also evaluated concentrations of these cytokines in healthy persons. Quality in Prognosis Studies tool was used to assess the methodological quality of included studies. We extracted summary data from published reports. Data analyses were performed using a random-effects model to estimate pooled odds ratios (OR) with 95% confidence intervals for cytokine levels and mortality. This systematic review is registered in PROSPERO (CRD42020179800). Findings: We identified 3654 records, and 104 studies were included with a total of 3250 participants. The pooled estimated mean TNFα concentration in sepsis patients was 58.4 pg/ml (95% Confidence Interval or CI 39.8-85.8 pg/ml) and 5.5 pg/ml (95% CI 3.8-8.0 pg/ml) in healthy controls. Pooled estimate means for IL-1β and IFNγ in sepsis patients were 21.8 pg/ml and 63.3 pg/ml, respectively. Elevated TNFα concentrations associated with increased 28-day sepsis mortality (p=0.001). In subgroup analyses, TNFα levels did not relate to sepsis source, sepsis severity, or sequential organ failure assessment (SOFA) score. Interpretation: TNFα concentration in sepsis is increased approximately 10-fold compared to healthy persons, and TNFα associated with sepsis mortality but not with sepsis severity. The concept that elevated cytokines cause sepsis should be revisited in the context of these data.

Comparison of enzyme-linked fluorescent assay and electrochemiluminescence immune assay in procalcitonin measurement

Background Procalcitonin (PCT) measurement is required for intensive care patients with systemic inflammation symptoms, early diagnosis of possible infections, and evaluation of sepsis severity and prognosis. Objectives We aimed to determine the analytical performance of PCT measurement in a Roche Modular E170 (ECLIA) analyzer and compare the performance with VIDAS (BRAHMS/ELFA) analyzer findings. Material and methods Within-day and between-day precision value, linearity was determined, and two methods were compared with regression and Bland–Altman analysis. Results Both ECLIA and ELFA assays indicated excellent precision, where within-day precision varied between 1.18% and 3.97% CV, and between-day precision varied between 1.77% and 3.93% CV. The ECLIA method was linear up to 62.15 ng/mL. The arithmetic mean was 6.02 ng/mL with the ECLIA method and 8.02 ng/mL with the ELFA method. The correlation coefficient was r=0.996 and p=0.001. The correlation was linear between the two methods. Regression equation was found y=0.78x − 0.23. The Bland–Altman figure was revealed the difference between the methods was specifically in lower concentrations (<0.15 ng/mL). Conclusions Both methods show good precision and correlation. It was determined that the difference between methods was significant, especially at <0.15 ng/mL concentration.

1003. Cytokine Levels in Sepsis and TNFα Association with Mortality but not Sepsis Severity or Infection Source: a Systematic Review and Meta-analysis

Background Sepsis is a global health problem associated with significant morbidity and mortality and is attributed to a “cytokine storm.”. However, anti-cytokine therapies have failed to lower sepsis mortality in clinical trials. Linking cytokine excess to sepsis pathogenesis requires quantification of cytokine levels in sepsis. This systematic review and meta-analysis characterizes levels of key cytokines in the circulation of sepsis patients and relates TNFα levels to mortality and patient characteristics. Methods Medline, Embase, Cochrane Library, and Web of Science Core Collection databases were searched from 1946 to May 2020 for studies in English disclosing cytokine levels in sepsis. Keywords included sepsis, septic shock, purpura fulminans, and tumor necrosis factor (TNF)α. We related cytokine amounts to 28-day mortality. Data analyses were performed using a random-effects model to estimate pooled odds ratios (OR) and 95% confidence intervals (CI). This systematic review is registered in PROSPERO under number CRD42020179800. Results A total of 3656 records were identified. After exclusions, 103 studies were included. Among these studies, 72 disclosed TNFα levels, 25 showed interleukin (IL)-1β levels, and 6 presented interferon (IFN)γ levels. The pooled estimate mean TNFα concentration in sepsis patients was 58.4 pg/ml (95% CI, 39.8-85.8 pg.ml; I2 = 99.4%). Pooled estimate means for IL-1α and IFNγ in sepsis patients were 21.8 pg/ml (95% CI, 12.6-37.8 pg.ml; I2 =99.8%) and 63.3 pg/ml (95% CI, 19.4-206.6 pg/ml; I2 = 99.7%), respectively. Elevated TNFα concentrations were associated with increased 28-day mortality (P=0.001). In a subgroup analysis, TNFα levels did not relate to sepsis source, sepsis severity, or sequential organ failure assessment (SOFA) score (figure 1). In a metaregression, TNFα associated with age, percentage of females and mortality at 28 days. Figure 1: A: TNFa levels according to sepsis source. B: TNFa levels according to measurement technique. C: TNFa levels according to presence or absence of cardiovascular disease. D: TNFa levels according to presence or absence of malignancy. E: TNFa levels according to sepsis severity. F: TNFa levels in fungal compared to other causes of sepsis (Yes=fungal sepsis; No= Other types of sepsis). G: TNFa levels according to SOFA score. H: TNFa levels and mortality at 28 days. Conclusion We presented levels of TNFα, IL-1β, and IFNγ in human sepsis and showed that TNFα elevations are associated with sepsis mortality. TNFα concentrations did not correlate with sepsis severity. We believe the concept that elevated cytokines cause sepsis should be revisited in the context of these data. Disclosures All Authors: No reported disclosures

Presence of Candida tropicalis on Staphylococcus epidermidis Biofilms Facilitated Biofilm Production and Candida Dissemination: An Impact of Fungi on Bacterial Biofilms

While Staphylococcus epidermidis (SE) is a common cause of infections in implanted prostheses and other indwelling devices, partly due to the biofilm formation, Candida tropicalis (CT) is an emerging Candida spp. with a potent biofilm-producing property. Due to the possible coexistence between SE and CT infection in the same patient, characteristics of the polymicrobial biofilms from both organisms might be different from those of the biofilms of each organism. Then, the exploration on biofilms, from SE with or without CT, and an evaluation on l-cysteine (an antibiofilm against both bacteria and fungi) were performed. As such, Candida incubation in preformed SE biofilms (SE &gt; CT) produced higher biofilms than the single- (SE or CT) or mixed-organism (SE + CT) biofilms as determined by crystal violet staining and fluorescent confocal images with z-stack thickness analysis. In parallel, SE &gt; CT biofilms demonstrated higher expression of icaB and icaC than other groups at 20 and 24 h of incubation, suggesting an enhanced matrix polymerization and transportation, respectively. Although organism burdens (culture method) from single-microbial biofilms (SE or CT) were higher than multi-organism biofilms (SE + CT and SE &gt; CT), macrophage cytokine responses (TNF-α and IL-6) against SE &gt; CT biofilms were higher than those in other groups in parallel to the profound biofilms in SE &gt; CT. Additionally, sepsis severity in mice with subcutaneously implanted SE &gt; CT catheters was more severe than in other groups as indicated by mortality rate, fungemia, serum cytokines (TNF-α and IL-6), and kidney and liver injury. Although CT grows upon preformed SE-biofilm production, the biofilm structures interfered during CT morphogenesis leading to the frailty of biofilm structure and resulting in the prominent candidemia. However, l-cysteine incubation together with the organisms in catheters reduced biofilms, microbial burdens, macrophage responses, and sepsis severity. In conclusion, SE &gt; CT biofilms prominently induced biofilm matrix, fungemia, macrophage responses, and sepsis severity, whereas the microbial burdens were lower than in the single-organism biofilms. All biofilms were attenuated by l-cysteine.

Relation of Procollagen Type III Amino Terminal Propeptide Level to Sepsis Severity in Pediatrics

Background: Sepsis is still the main etiology of mortality in pediatric intensive care units (PICUs). Therefore, we performed this study to evaluate the value of procollagen Type III amino-terminal propeptide (PIIINP) as a biomarker for sepsis severity diagnosis and mortality. Method: A prospective study was carried out on 170 critically ill children admitted into the PICU and 100 controls. The performed clinical examinations included calculation of the pediatric risk of mortality. Serum PIIINP was withdrawn from patients at admission and from the controls. Results: PIIINP level was significantly more increased in sepsis, severe sepsis, and septic shock than among the controls (p < 0.001). PIIINP was significantly higher in severe sepsis and septic shock (568.3 (32.5–1304.7) and 926.2 (460.6–1370), respectively) versus sepsis (149.5 (29.6–272.9)) (p < 0.001). PIIINP was significantly increased in non-survivors (935.4 (104.6–1370)) compared to survivors (586.5 (29.6–1169)) (p < 0.016). ROC curve analysis exhibited an area under the curve (AUC) of 0.833 for PIIINP, which is predictive for sepsis, while the cut-off point of 103.3 ng/mL had a sensitivity of 88% and specificity of 82%. The prognosis of the AUC curve for PIIINP to predict mortality was 0.651; the cut-off of 490.4 ng/mL had a sensitivity of 87.5% and specificity of 51.6%. Conclusions: PIIINP levels are increased in sepsis, with significantly higher levels in severe sepsis, septic shock, and non-survivors, thus representing a promising biomarker for pediatric sepsis severity and mortality.