Effects of IFIH1 rs1990760 variants on systemic inflammation and outcome in critically ill COVID-19 patients in an observational translational study.

Background: Variants in IFIH1, a gene coding the cytoplasmatic RNA sensor MDA5, regulate the response to viral infections. We hypothesized that IFIH1 rs199076 variants would modulate host response and outcome after severe COVID-19. Methods: Patients admitted to an intensive care unit (ICU) with confirmed COVID-19 were prospectively studied and rs1990760 variants determined. Peripheral blood gene expression, cell populations, and immune mediators were measured. Peripheral blood mononuclear cells from healthy volunteers were exposed to an MDA5 agonist and dexamethasone ex-vivo, and changes in gene expression assessed. ICU discharge and hospital death were modeled using rs1990760 variants and dexamethasone as factors in this cohort and in-silico clinical trials. Results: About 227 patients were studied. Patients with the IFIH1 rs1990760 TT variant showed a lower expression of inflammation-related pathways, an anti-inflammatory cell profile, and lower concentrations of pro-inflammatory mediators. Cells with TT variant exposed to an MDA5 agonist showed an increase in IL6 expression after dexamethasone treatment. All patients with the TT variant not treated with steroids survived their ICU stay (hazard ratio [HR]: 2.49, 95% confidence interval [CI]: 1.29-4.79). Patients with a TT variant treated with dexamethasone showed an increased hospital mortality (HR: 2.19, 95% CI: 1.01-4.87) and serum IL-6. In-silico clinical trials supported these findings. Conclusions: COVID-19 patients with the IFIH1 rs1990760 TT variant show an attenuated inflammatory response and better outcomes. Dexamethasone may reverse this anti-inflammatory phenotype. Funding: Centro de Investigación Biomédica en Red (CB17/06/00021), Instituto de Salud Carlos III (PI19/00184 and PI20/01360), and Fundació La Marató de TV3 (413/C/2021).

Patients with severe COVID-19 often need mechanical ventilation to help them breathe and other types of intensive care. The outcome for many of these patients depends on how their immune system reacts to the infection. If the inflammatory response triggered by the immune system is too strong, this can cause further harm to the patient. One gene that plays an important role in inflammation is IFIH1 which encodes a protein that helps the body to recognize viruses. There are multiple versions of this gene which each produce a slightly different protein. It is possible that this variation impacts how the immune system responds to the virus that causes COVID-19. To investigate, Amado-Rodríguez, Salgado del Riego et al. analyzed the IFIH1 gene in 227 patients admitted to an intensive care unit in Spain for severe COVID-19 between March and December 2020. They found that patients with a specific version of the gene called TT experienced less inflammation and were more likely to survive the infection. Physicians typically treat patients with moderate to severe COVID-19 with corticosteroid drugs that reduce the inflammatory response. However, Amado-Rodríguez, Salgado del Riego et al. found that patients with the TT version of the IFIH1 gene were at greater risk of dying if they received corticosteroids. The team then applied the distribution of IFIH1 variants among different ethnic ancestries to data from a previous clinical trial, and simulated the effects of corticosteroid treatment. This 'mock' clinical trial supported their findings from the patient-derived data, which were also validated by laboratory experiments on immune cells from individuals with the TT gene. The work by Amado-Rodríguez, Salgado del Riego et al. suggests that while corticosteroids benefit some patients, they may cause harm to others. However, a real-world clinical trial is needed to determine whether patients with the TT version of the IFIH1 gene would do better without steroids.

N-terminal pro B-type natriuretic peptide and angiogenic biomarkers in the prognosis of adverse outcomes in women with suspected preeclampsia.

BACKGROUND: This study compares the performance of the soluble fms-like tyrosine kinase 1 to placental growth factor (sFlt-1/PlGF) ratio and the cardiac biomarker N-terminal pro-B type natriuretic peptide (NT-proBNP) in the prediction of adverse outcomes in women with suspicion of PE. METHODS: A retrospective cohort study was conducted on women admitted at triage with signs and/or symptoms of PE (n=340). Serum levels of sFlt-1, PlGF and NT-proBNP were determined by an electrochemiluminescence immunoassay (Roche Diagnostics). The main outcomes were early- or late-onset PE and development of adverse outcome, defined as delivery within the first week since clinical presentation or fetal/early neonatal death. RESULTS: NT-proBNP concentrations (ng/L) were significantly increased in PE versus non-PE women, both at <34 (169 versus 34) and ≥34weeks of gestation (101 versus 49) (p<0.001). A cut-point of 70 showed sensitivities/specificities of 78/74% for early-, and 70/62% for late-onset PE; slightly lower than those offered by the sFlt-1/PlGF ratio or uric acid. The respective cut-points of 178 and 219 for sFlt-1/PlGF ratio and NT-proBNP, demonstrated similar performance in the prediction of adverse outcome, with sensitivity/specificity of 95/84% and 94/76%, respectively. CONCLUSION: NT-proBNP and sFlt-1/PlGF ratio can be used to predict the development of an adverse outcome.

Interleukin-6 in cerebrospinal fluid as a biomarker of acute meningitis.

BACKGROUND: Microbiological culture of cerebrospinal fluid is the gold standard to differentiate between aseptic and bacterial meningitis, but this method has low sensitivity. A fast and reliable new marker would be of interest in clinical practice. OBJECTIVE: Interleukin-6, secreted by T cells in response to meningeal pathogens and quickly delivered into cerebrospinal fluid, was evaluated as a marker of acute meningitis. DESIGN AND METHODS: A total of 150 cerebrospinal fluid samples were analysed by an electrochemiluminescence method, selected according to patient diagnosis: (a) bacterial meningitis confirmed by positive culture (n = 26); (b) bacterial meningitis with negative culture or not performed (n = 15); (c) viral meningitis confirmed by polymerase chain reaction or immunoglobulin G determination (n = 23); (d) viral meningitis with polymerase chain reaction negative or not performed (n = 42); and (e) controls (n = 44). RESULTS: Cerebrospinal fluid interleukin-6 concentration showed significant differences between all pathologic groups and the control group (P < 0.001). As a diagnostic tool for bacterial meningitis, interleukin-6 showed an area under the curve of 0.937 (95% confidence intervals: 0.895-0.978), significantly higher than those of classical biomarkers. An interleukin-6 cutoff of 1418 pg/mL showed 95.5% sensitivity and 77.5% specificity, whereas a value of 15,060 pg/mL showed 63.6% sensitivity and 96.7% specificity, for diagnosis of bacterial meningitis. CONCLUSION: Interleukin-6 measured by electrochemiluminescence method is a promising marker for early differentiation between aseptic and bacterial meningitis. More studies are needed to validate clinical implications for future practice in an emergency laboratory.

Role of vitamin D and sFlt-1/PlGF ratio in the development of early- and late-onset preeclampsia.

BACKGROUND: The imbalanced production of placental biomarkers and vitamin D deficiency have been proposed as risk factors for the development of preeclampsia (PE). However, little is known about the relationship between them and their role in early- versus late-onset PE. The objectives were to assess the role of 25-hydroxyvitamin D [25(OH)D] concentrations and the soluble fms-like tyrosine kinase 1 (sFlt-1) to placental growth factor (PlGF) ratio in the development of early- and late-onset PE; and to evaluate the relationship between 25(OH)D and the biomarkers. METHODS: A retrospective, full-blinded cohort study was conducted at the Obstetric Emergency Service of a tertiary care hospital. Pregnant women (n=257) attending obstetric triage with suspicion of PE were included. sFlt-1, PlGF and 25(OH)D concentrations were measured by electrochemoluminescence (ECLIA) immunoassay and pregnancy outcome (development of PE) was registered from patients records. RESULTS: PE women showed lower 25(OH)D concentrations at clinical presentation than non-PE women (median: 35.0 nmol/L and 39.6 nmol/L, respectively; p=0.027). Women with 25(OH)D levels <50 nmol/L experienced an increased risk of developing late-onset PE [odds ratio (OR) 4.6, 95% confidence interval (CI) 1.4-15], but no association was found for early-onset PE. However, a sFlt-1/PlGF ratio above the corresponding cutpoints increased the risk of developing both early- and late-onset PE [ORs 58 (95% CI 11-312) and 12 (95% CI 5.0-27), respectively]. No association was found between 25(OH)D levels and sFlt-1/PlGF ratio. CONCLUSIONS: Low vitamin D status in women with suspected late-onset PE increases the risk of imminent development of the disease.

Vitamin D deficiency at pediatric intensive care admission / Deficiência de vitamina D em internações na unidade de terapia intensiva pediátrica

OBJECTIVE: to assess whether 25hydroxivitaminD or 25(OH)vitD deficiency has a high prevalence at pediatric intensive care unit (PICU) admission, and whether it is associated with increased prediction of mortality risk scores. METHOD: prospective observational study comparing 25(OH)vitD levels measured in 156 patients during the 12 hours after critical care admission with the 25(OH)vitD levels of 289 healthy children. 25(OH)vitD levels were also compared between PICU patients with pediatric risk of mortality III (PRISM III) or pediatric index of mortality 2 (PIM 2) > p75 [(group A; n = 33) vs. the others (group B; n = 123)]. Vitamin D deficiency was defined as < 20 ng/mL levels. RESULTS: median (p25-p75) 25(OH)vitD level was 26.0 ng/mL (19.2-35.8) in PICU patients vs. 30.5 ng/mL (23.2-38.6) in healthy children (p = 0.007). The prevalence of 25(OH)vitD < 20 ng/mL was 29.5% (95% CI: 22.0-37.0) vs. 15.6% (95% CI: 12.2-20.0) (p = 0.01). Pediatric intensive care patients presented an odds ratio (OR) for hypovitaminosis D of 2.26 (CI 95%: 1.41-3.61). 25(OH)vitD levels were 25.4 ng/mL (CI 95%: 15.5-36.0) in group A vs. 26.6 ng/mL (CI 95%: 19.3-35.5) in group B (p = 0.800). CONCLUSIONS: hypovitaminosis D incidence was high in PICU patients. Hypovitaminosis D was not associated with higher prediction of risk mortality scores. .

OBJETIVO: avaliar se a deficiência da 25-hidroxivitamina D, ou 25 (OH) vitD, tem prevalência elevada em internações na unidade de terapia intensiva pediátrica, e se estaria relacionada à previsão de escores de risco de mortalidade. MÉTODO: estudo observacional prospectivo comparando níveis de 25 (OH) vitD de 156 pacientes, mensurados nas primeiras 12 horas da internação em terapia intensiva, com níveis de 25 (OH) vitD de 289 crianças saudáveis. Os níveis de 25 (OH) vitD também foram comparados entre pacientes na UTIP com escore PRISM III ou PIM 2 > p75 (Grupo A; n = 33), e o restante, (Grupo B; n = 123). A deficiência de vitamina D foi definida como níveis < 20 ng/mL. RESULTADOS: o nível médio (p25-p75) de 25 (OH) vitD foi 26,0 ng/mL (19,2-35,8) em pacientes internados na UTIP, em comparação a 30,5 ng/mL (23,2-38,6) em crianças saudáveis (p = 0,007). A prevalência de 25 (OH) vitD < 20 ng/mL foi de 29,5% (IC 95%, 22,0-37,0), em comparação a 15,6% (IC 95%,12,2-20,0) (p = 0,01). Os pacientes em terapia intensiva pediátrica apresentaram uma razão de chance (RC) para hipovitaminose D de 2,26 (IC 95%, 1,41-3,61). Os níveis de 25 (OH) vitD foram 25,4 ng/mL (IC 95%, 15,5-36,0) no grupo A, em comparação a 26,6 ng/mL (IC 95%, 19,3-35,5) no grupo B (p = 0,800). CONCLUSÕES: a incidência de hipovitaminose D foi elevada em pacientes em terapia intensiva pediátrica, mas não foi associada à maior previsão de escores de risco de mortalidade. .

New biomarkers in diagnosis of early onset preeclampsia and imminent delivery prognosis.

BACKGROUND: Several studies have revealed a high soluble fms-like tyrosine kinase 1 (sFlt-1) to placental growth factor (PlGF) ratio in preeclamptic women. However, its role in patients with suspected preeclampsia (PE) at triage in the emergency department remains an issue and a controversial unique cutpoint of 85 has been proposed regardless of gestational age. A new cutpoint for sFlt-1/PlGF ratio was investigated to rule out PE at obstetric triage, and to assess its prognostic value for risk of imminent delivery. METHODS: Blood samples from 257 pregnant women with suspected PE were obtained at obstetric triage admission. Serum PlGF and sFlt-1 were measured by an electrochemoluminiscence immunoassay (ECLIA) on the immunoanalyzer Cobas e601 (Roche Diagnostics) and the corresponding ratio was calculated. Final outcomes (mainly development of PE) were reviewed and time between clinical presentation and delivery was calculated. RESULTS: The best ratio cutpoint to diagnose PE changed according to gestational age: 23 (92.0% sensitivity, 81.1% specificity) and 45 (83.7% sensitivity, 72.6% specificity) for women <34 and ≥ 34 weeks' gestation, respectively. Furthermore, sFlt-1/PlGF ratio inversely correlated with time elapsed between clinical presentation and delivery, and a cutpoint of 178 could predict complications such as imminent delivery or fetal/neonatal death with a sensitivity of 70.6% and a specificity of 97.8%. CONCLUSIONS: The new cut-off values for the sFlt-1/PlGF ratio adjusted by the gestational age at clinical presentation can be used to rule out PE at obstetric triage and to predict imminent delivery with better accuracy than the cutpoint currently accepted.

New quantitative electrochemiluminescence method (ECLIA) for interleukin-6 (IL-6) measurement.

BACKGROUND: A new interleukin-6 (IL-6) electrochemiluminescent immunoassay (ECLIA, Roche Diagnostics) was evaluated and compared to a previous semiquantitative immunoassay (Milenia Biotec). Reference ranges for cord blood plasma were also calculated. METHODS: The new IL-6 ECLIA test was performed using a Cobas E601 analyzer (Roche Diagnostics). The comparison method (Milenia Biotec) was a semiquantitative lateral flow immunoassay, coupled to a digital image capture system (PicoScan). Healthy at term newborns were recruited to establish reference ranges for IL-6 in cord blood plasma. RESULTS: Total imprecision ranged from 3.7% to 8.0% depending on the IL-6 concentrations. The calculated limit of detection for IL-6 measured by ECLIA was 2.63 pg/mL, almost twice as high as that claimed by the manufacturer (1.5 pg/mL). The linearity of the method was verified to 5000 pg/mL. The IL-6 reference limit, obtained from 148 cord blood samples, was 30 pg/mL (90% confidence interval: 19-48 pg/mL). CONCLUSIONS: The new quantitative ECLIA method showed good reproducibility, linearity and functional sensitivity. Additional clinically based studies are needed to elucidate the usefulness of the IL-6 reference limit, calculated here for the first time by ECLIA in cord blood plasma, to aid in the diagnosis of vertical transmission of sepsis.

Clinical utility of NT-proBNP levels in late heart transplantation patients.

BACKGROUND: The present work was aimed to describe NT-proBNP levels in heart transplant patients after the first year postsurgery. METHODS: NT-proBNP concentration was measured in 1231 samples from 142 patients when a routine four-month follow-up was carried out, including other biochemical and clinical examinations. Endomyocardial biopsies were performed only upon clinical suspicion of acute rejection. RESULTS: NT-proBNP concentrations were not significantly correlated to post-transplantation time, though differences were observed according to clinical symptoms (Kruskal-Wallis test, p<0.001). Although multivariate analysis revealed statistically significant association between NT-proBNP concentration and some qualitative (cardiac allograft vasculopathy-CAV-, sex, diabetes) and quantitative (creatinine, hematocrit, age) variables, only moderately relevant contribution was observed for creatinine and CAV. Patients with rejection showed noticeable increases in serum NT-proBNP concentrations, above more than 2.5 times the reference change value (97%). NT-proBNP concentrations higher than 1000ng/L increased in 3.5 times (95%CI: 2.4-5) the risk of death in less than one year. CONCLUSIONS: After the first year from surgery, NT-proBNP concentrations were not associated to post-transplantation time and NT-proBNP could be a useful diagnostic marker for rejection, whenever serial measurements are made. A NT-proBNP cutoff value of 1000ng/L was identified for classifying patients at risk of death after one year.

Plasma procalcitonin measured by time-resolved amplified cryptate emission (TRACE) in liver transplant patients. A prognosis marker of early infectious and non-infectious postoperative complications.

BACKGROUND: Elevated procalcitonin (PCT) levels are observed after major surgery, such as orthotopic liver transplantation (OLTx). The aim of this observational study was to evaluate PCT kinetics during the first 5 following days after surgery to establish the prognostic value of PCT changes in the outcome of OLTx, and to predict medical, technical and infectious complications. PCT was also evaluated in the differential diagnosis of infection vs. rejection. METHODS: A total of 64 OLTx were performed in 58 patients; they were split into two groups: with and without complications. Out of these patients, 18 developed infection, and nine rejection. PCT was measured before and during surgery, 12 h after transplantation and daily for the 5 following days. PCT was also measured the day when infection or rejection was diagnosed, and on the previous day. PCT was determined by time-resolved amplified cryptate emission (TRACE) technology. RESULTS: PCT elevation began at 12 h after surgery, reaching a peak on the 1st day in both groups. Significantly higher PCT concentrations were found in the group of patients developing complications, on the 5 postoperative days. It was found that a 24 h PCT value higher than 1.92 microg/L increased by 9.1-time-fold the risk of complications. When infection was diagnosed, a second peak of PCT was observed, but no PCT elevation was shown in rejection. CONCLUSIONS: Daily monitored PCT provides valuable information about the early outcome of OLTx.

A monoclonal antibody with potential for aiding non-invasive prenatal diagnosis: utility in screening of pregnant women at risk of preeclampsia.

The development of a non-invasive method of prenatal diagnosis in maternal blood has been the goal of our investigations during the last years. We have developed several anti-CD71 monoclonal antibodies and optimized a protocol for the isolation of nucleated red blood cells (NRBC) from peripheral maternal blood. The enhanced traffic of fetal erythroblasts into the maternal circulation in preeclampsia has been investigated by several groups. In this study, we compared one of our antibodies, 2F6.3, with a commercial anti-CD71 antibody in blood samples from pregnant women suffering pregnancy-induced hypertension (PIH) and in a control group of pregnant women without clinical features suggestive of an increased risk of developing preeclampsia. The mAb 2F6.3, developed by our group, has succeeded in isolating a significantly higher number of erythroblasts with less maternal cell contamination than the commercial antibody in both women with PIH and in the control group (p<0.01; Wilcoxon Signed Ranks Test). Fluorescence in situ hybridization analysis also demonstrated that 2F6.3 is a better antibody for the isolation of fetal NRBC in maternal blood than the commercial anti-CD71 antibody.