Tujana Boldanova

Incremental value of copeptin for rapid rule out of acute myocardial infarction.

OBJECTIVES The purpose of this study was to examine the incremental value of copeptin for rapid rule out of acute myocardial infarction (AMI). BACKGROUND The rapid and reliable exclusion of AMI is a major unmet clinical need. Copeptin, the C-terminal part of the vasopressin prohormone, as a marker of acute endogenous stress may be useful in this setting. METHODS In 487 consecutive patients presenting to the emergency department with symptoms suggestive of AMI, we measured levels of copeptin at presentation, using a novel sandwich immunoluminometric assay in a blinded fashion. The final diagnosis was adjudicated by 2 independent cardiologists using all available data. RESULTS The adjudicated final diagnosis was AMI in 81 patients (17%). Copeptin levels were significantly higher in AMI patients compared with those in patients having other diagnoses (median 20.8 pmol/l vs. 6.0 pmol/l, p < 0.001). The combination of troponin T and copeptin at initial presentation resulted in an area under the receiver-operating characteristic curve of 0.97 (95% confidence interval: 0.95 to 0.98), which was significantly higher than the 0.86 (95% confidence interval: 0.80 to 0.92) for troponin T alone (p < 0.001). A copeptin level <14 pmol/l in combination with a troponin T < or =0.01 microg/l correctly ruled out AMI with a sensitivity of 98.8% and a negative predictive value of 99.7%. CONCLUSIONS The additional use of copeptin seems to allow a rapid and reliable rule out of AMI already at presentation and may thereby obviate the need for prolonged monitoring and serial blood sampling in the majority of patients. (Advantageous Predictors of Acute Coronary Syndromes Evaluation [APACE]; NCT00470587).

IFN-λ receptor 1 expression is induced in chronic hepatitis C and correlates with the IFN-λ3 genotype and with nonresponsiveness to IFN-α therapies

Liver biopsies from patients with chronic hepatitis C reveal high IFN-λR1 expression, which can be induced with IFN-α and is associated with elevated ISG expression, the IFN-λ3 minor alleles, and nonresponsiveness to peg-IFN-α and ribavirin therapy.

QRS and QTc interval prolongation in the prediction of long-term mortality of patients with acute destabilised heart failure

Objectives: To quantify the prognostic utility of QRS and QTc interval prolongation in patients presenting with acute destabilised heart failure (ADHF) to the emergency department (ED). Design: Prospective cohort study among patients enrolled in the B-Type Natriuretic Peptide for Acute Shortness of Breath Evaluation (BASEL) study. QRS and QT intervals were measured in 173 consecutive patients with ADHF. QT interval was corrected using the Bazett formula. The primary end point was all-cause mortality during the 720-day follow-up. Results: QRS interval was prolonged (⩾120 ms) in 27% of patients, and QTc interval was prolonged (⩾440 ms) in 72% of patients. Baseline demographic and clinical characteristics were comparable in patients with normal and prolonged QRS or QTc intervals. A total of 78 patients died during follow-up. Interestingly, the 720-day mortality was similar in patients with prolonged and normal QTc (44% vs 42%, p = 0.546), but was significantly higher in patients with prolonged QRS interval than in those with normal QRS (59% vs 37%, p = 0.004). In Cox proportional hazards analysis, prolonged QRS interval was associated with a nearly twofold increase in mortality (HR 1.94, 95% CI 1.22 to 3.07; p = 0.005). This association persisted after adjustment for variables routinely available in the ED. Conclusions: Prolonged QRS interval, but not prolonged QTc interval, is associated with increased long-term mortality in patients with ADHF.

Interferon-inducible cholesterol-25-hydroxylase restricts hepatitis C virus replication through blockage of membranous web formation.

Hepatitis C virus (HCV) is a positive‐strand RNA virus that primarily infects human hepatocytes. Infections with HCV constitute a global health problem, with 180 million people currently chronically infected. Recent studies have reported that cholesterol 25‐hydroxylase (CH25H) is expressed as an interferon‐stimulated gene and mediates antiviral activities against different enveloped viruses through the production of 25‐hydroxycholesterol (25HC). However, the intrinsic regulation of human CH25H (hCH25H) expression within the liver as well as its mechanistic effects on HCV infectivity remain elusive. In this study, we characterized the expression of hCH25H using liver biopsies and primary human hepatocytes. In addition, the antiviral properties of this protein and its enzymatic product, 25HC, were further characterized against HCV in tissue culture. Levels of hCH25H messenger RNA were significantly up‐regulated both in HCV‐positive liver biopsies and in HCV‐infected primary human hepatocytes. The expression of hCH25H in primary human hepatocytes was primarily and transiently induced by type I interferon. Transient expression of hCH25H in human hepatoma cells restricted HCV infection in a genotype‐independent manner. This inhibition required the enzymatic activity of CH25H. We observed an inhibition of viral membrane fusion during the entry process by 25HC, which was not due to a virucidal effect. Yet the primary effect by 25HC on HCV was at the level of RNA replication, which was observed using subgenomic replicons of two different genotypes. Further analysis using electron microscopy revealed that 25HC inhibited formation of the membranous web, the HCV replication factory, independent of RNA replication. Conclusion: Infection with HCV causes up‐regulation of interferon‐inducible CH25H in vivo, and its product, 25HC, restricts HCV primarily at the level of RNA replication by preventing formation of the viral replication factory. (Hepatology 2015;62:702–714)

Impact of history of heart failure on diagnostic and prognostic value of BNP: Results from the B-type Natriuretic Peptide for Acute Shortness of Breath Evaluation (BASEL) Study ☆

OBJECTIVES This study aimed to examine the influence of history of heart failure (HF) on circulating levels, diagnostic accuracy and prognostic value of B-type natriuretic peptide (BNP) in patients presenting with all cause dyspnea at the emergency department. BACKGROUND BNP has been shown to be very helpful in diagnosis and prognosis of HF. Due to chronically elevated cardiac filling pressures, patients with a history of HF might have higher BNP levels and therefore diagnostic and prognostic properties of BNP may be affected. METHODS We analyzed circulating levels, diagnostic accuracy and prognostic value of BNP in 388 patients without a previous history of HF and compared these to data to 64 patients with a history of HF included in the B-type Natriuretic Peptide for Acute Shortness of Breath Evaluation (BASEL) Study. RESULTS Baseline BNP levels were higher in patients with a history of HF (median 814 pg/ml [353-1300 pg/ml] vs. 216 pg/ml [45-801 pg/ml], p<0.001). Diagnostic accuracy of BNP to identify HF was comparable in patients with (AUC=0.804; 95% CI 0.628-0.980) and in patients without history of HF (AUC=0.883; 95% CI 0.848-0.919, p=0.389). Prognostic ability of BNP to predict one-year mortality was lower in overall patients with history of HF (AUC=0.458; 95%CI 0.294-0.622) compared to patients without history of HF (AUC=0.710; 95% CI 0.653-0.768, p<0.05). CONCLUSIONS In patients with history of HF, BNP levels retain diagnostic accuracy. Ability to predict one-year mortality was decreased in unselected patients, but not in patients with acute HF-induced dyspnea.

Gene expression analysis of biopsy samples reveals critical limitations of transcriptome-based molecular classifications of hepatocellular carcinoma.

Molecular classification of hepatocellular carcinomas (HCC) could guide patient stratification for personalized therapies targeting subclass‐specific cancer ‘driver pathways’. Currently, there are several transcriptome‐based molecular classifications of HCC with different subclass numbers, ranging from two to six. They were established using resected tumours that introduce a selection bias towards patients without liver cirrhosis and with early stage HCCs. We generated and analyzed gene expression data from paired HCC and non‐cancerous liver tissue biopsies from 60 patients as well as five normal liver samples. Unbiased consensus clustering of HCC biopsy profiles identified 3 robust classes. Class membership correlated with survival, tumour size and with Edmondson and Barcelona Clinical Liver Cancer (BCLC) stage. When focusing only on the gene expression of the HCC biopsies, we could validate previously reported classifications of HCC based on expression patterns of signature genes. However, the subclass‐specific gene expression patterns were no longer preserved when the fold‐change relative to the normal tissue was used. The majority of genes believed to be subclass‐specific turned out to be cancer‐related genes differentially regulated in all HCC patients, with quantitative rather than qualitative differences between the molecular subclasses. With the exception of a subset of samples with a definitive β‐catenin gene signature, biological pathway analysis could not identify class‐specific pathways reflecting the activation of distinct oncogenic programs. In conclusion, we have found that gene expression profiling of HCC biopsies has limited potential to direct therapies that target specific driver pathways, but can identify subgroups of patients with different prognosis.

Quantitative proteomics and phosphoproteomics on serial tumor biopsies from a sorafenib-treated HCC patient.

Significance Elucidation of evasive resistance to targeted therapies is a major challenge in cancer research. As a proof-of-concept study, we describe quantitative proteomics and phosphoproteomics on serial tumor biopsies from a sorafenib-treated hepatocellular carcinoma (HCC) patient. This approach reveals signaling pathway activity in a tumor and how it evades therapy. The described method will allow precision medicine based on phenotypic data. In particular, application of the method to a patient cohort will potentially identify new biomarkers, drug targets, and signaling pathways that mediate evasive resistance. Compensatory signaling pathways in tumors confer resistance to targeted therapy, but the pathways and their mechanisms of activation remain largely unknown. We describe a procedure for quantitative proteomics and phosphoproteomics on snap-frozen biopsies of hepatocellular carcinoma (HCC) and matched nontumor liver tissue. We applied this procedure to monitor signaling pathways in serial biopsies taken from an HCC patient before and during treatment with the multikinase inhibitor sorafenib. At diagnosis, the patient had an advanced HCC. At the time of the second biopsy, abdominal imaging revealed progressive disease despite sorafenib treatment. Sorafenib was confirmed to inhibit MAPK signaling in the tumor, as measured by reduced ribosomal protein S6 kinase phosphorylation. Hierarchical clustering and enrichment analysis revealed pathways broadly implicated in tumor progression and resistance, such as epithelial-to-mesenchymal transition and cell adhesion pathways. Thus, we describe a protocol for quantitative analysis of oncogenic pathways in HCC biopsies and obtained first insights into the effect of sorafenib in vivo. This protocol will allow elucidation of mechanisms of resistance and enable precision medicine.

Hepatitis B Virus Does Not Interfere With Innate Immune Responses in the Human Liver

BACKGROUND & AIMS Most viruses are detected at early stages of cell infection and induce an innate immune response mediated by production of interferons (IFNs). IFNs induce expression of hundreds of IFN-stimulated genes (ISGs). Infection of chimpanzees with hepatitis C virus, but not hepatitis B virus (HBV), induces ISG expression in the liver. HBV might not induce an innate immune response because it is not detected by pattern recognition receptors (the stealth properties of HBV) or because HBV suppresses IFN production or signaling despite detection by pattern recognition receptors. We studied innate immune signaling in liver biopsies from patients with different stages of chronic HBV infection and uninfected individuals (controls). METHODS We obtained liver within 10 minutes after collection from 30 patients with chronic HBV infection (hepatitis B e antigen-positive or -negative, with or without hepatitis) and 42 controls (most with fatty liver disease). The liver tissues were analyzed by histology, immunohistochemistry, quantitative reverse-transcription polymerase chain reaction, in situ hybridization, HBV RNA quantification, and HBV genotyping; some specimens were incubated with toll-like receptor (TLR) ligands (polyinosinic-polycytidylic acid) or infected with Sendai virus and then analyzed. RESULTS Liver specimens from patients with HBV infection were not expressing more IFN or ISGs than those from control patients, indicating that chronic HBV infection did not activate an innate immune response. However, liver specimens from patients with HBV infection did produce IFN and induce expression of ISGs following activation of TLR3 with poly(I:C) or Sendai virus infections, so the innate immune response is not suppressed in these tissues. CONCLUSION Liver tissues from patients with chronic HBV infection do not have induction of an innate immune response, but this response can be activated by other factors (TLR3 binding, Sendai virus infection) in HBV-infected liver tissue. These findings support the hypothesis that HBV is invisible to pattern recognition receptors.

Incremental value of multiplex real-time PCR for the early diagnosis of sepsis in the emergency department

BACKGROUND Delayed recognition of sepsis and inappropriate initial antibiotic therapy are associated with increased mortality and morbidity. The early detection of the causative organism in sepsis is an unmet clinical need. A novel multiplex real-time polymerase chain reaction (MRT-PCR) (SeptiFast®) technique may provide the microbiological diagnosis within six hours. METHODS We assessed the diagnostic accuracy of blood cultures and MRT-PCR in a comparative diagnostic cohort study in 110 consecutive adult patients presenting to the emergency department (ED) with suspected sepsis. RESULTS We collected 205 corresponding PCR samples and blood culture (BC) pairs from the 110 patients. There was moderate to high concordance between PCR and BC with 181 (88%) matching and 24 (12%) mismatching samples. The diagnostic accuracy of MRT-PCR in detecting sepsis and its causative organism was comparable to that of BCs. The additional use of MRT-PCR significantly reduced the time to microbiological diagnosis as compared to the use of conventional microbiological methods alone (mean time gained 3.9 hours, range 0-66 hours, p <0.001). CONCLUSION Diagnostic accuracy of BCs and MRT-PCR in the early diagnosis of sepsis and its causative organism in the ED are comparable. However, MRT-PCR reduces the time to microbiological diagnosis. Whether a more rapid detection of the organism by MRT-PCR could improve the outcome of patients has to be assessed in large prospective randomised trials.

Transcriptional response to hepatitis C virus infection and interferon‐alpha treatment in the human liver

Hepatitis C virus (HCV) is widely used to investigate host–virus interactions. Cellular responses to HCV infection have been extensively studied in vitro. However, in human liver, interferon (IFN)‐stimulated gene expression can mask direct transcriptional responses to infection. To better characterize the direct effects of HCV infection in vivo, we analyze the transcriptomes of HCV‐infected patients lacking an activated endogenous IFN system. We show that expression changes observed in these patients predominantly reflect immune cell infiltrates rather than cell‐intrinsic pathways. We also investigate the transcriptomes of patients with endogenous IFN activation, which paradoxically cannot eradicate viral infection. We find that most IFN‐stimulated genes are induced by both recombinant IFN therapy and the endogenous IFN system, but with lower induction levels in the latter, indicating that the innate immune response in chronic hepatitis C is too weak to clear the virus. We show that coding and non‐coding transcripts have different expression dynamics following IFN treatment. Several microRNA primary transcripts, including that of miR‐122, are significantly down‐regulated in response to IFN treatment, suggesting a new mechanism for IFN‐induced expression fine‐tuning.