Junín Virus Infection of Human Hematopoietic Progenitors Impairs In Vitro Proplatelet Formation and Platelet Release via a Bystander Effect Involving Type I IFN Signaling

Argentine hemorrhagic fever (AHF) is an endemo-epidemic disease caused by Junín virus (JUNV), a member of the arenaviridae family. Although a recently introduced live attenuated vaccine has proven to be effective, AHF remains a potentially lethal infection. Like in other viral hemorrhagic fevers (VHF), AHF patients present with fever and hemorrhagic complications. Although the causes of the bleeding are poorly understood, impaired hemostasis, endothelial cell dysfunction and low platelet counts have been described. Thrombocytopenia is a common feature in VHF syndromes, and it is a major sign for its diagnosis. However, the underlying pathogenic mechanism has not yet been elucidated. We hypothesized that thrombocytopenia results from a viral-triggered alteration of the megakaryo/thrombopoiesis process. Therefore, we evaluated the impact of JUNV on megakaryopoiesis using an in vitro model of human CD34+ cells stimulated with thrombopoietin. Our results showed that CD34+ cells are infected with JUNV in a restricted fashion. Infection was transferrin receptor 1 (TfR1)-dependent and the surface expression of TfR1 was higher in infected cultures, suggesting a novel arenaviral dissemination strategy in hematopoietic progenitor cells. Although proliferation, survival, and commitment in JUNV-infected cultures were normal, viral infection impaired thrombopoiesis by decreasing in vitro proplatelet formation, platelet release, and P-selectin externalization via a bystander effect. The decrease in platelet release was also TfR1-dependent, mimicked by poly(I:C), and type I interferon (IFN α/β) was implicated as a key paracrine mediator. Among the relevant molecules studied, only the transcription factor NF-E2 showed a moderate decrease in expression in megakaryocytes from either infected cultures or after type I IFN treatment. Moreover, type I IFN-treated megakaryocytes presented ultrastructural abnormalities resembling the reported thrombocytopenic NF-E2−/− mouse phenotype. Our study introduces a potential mechanism for thrombocytopenia in VHF and other diseases associated with increased bone marrow type I IFN levels

Increased Levels of Leukocyte-Derived MMP-9 in Patients with Stable Angina Pectoris

Objective: There is a growing interest for matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) in plasma as novel biomarkers in coronary artery disease (CAD). We aimed to identify the sources of MMP-8, MMP-9, TIMP-1 and TIMP-2 among peripheral blood cells and further explore whether gene expression or protein release was altered in patients with stable angina pectoris (SA). Methods: In total, plasma MMP-9 was measured in 44 SA patients and 47 healthy controls. From 10 patients and 10 controls, peripheral blood mononuclear cells (PBMC) and neutrophils were isolated and stimulated ex vivo. MMPs, TIMPs and myeloperoxidase were measured in plasma and supernatants by ELISA. The corresponding gene expression was measured by real-time PCR. Results: Neutrophils were the dominant source of MMP-8 and MMP-9. Upon moderate stimulation with IL-8, the neutrophil release of MMP-9 was higher in the SA patients compared with controls (p,0.05). In PBMC, the TIMP-1 and MMP-9 mRNA expression was higher in SA patients compared with controls, p,0.01 and 0.05, respectively. There were no differences in plasma levels between patients and controls except for TIMP-2, which was lower in patients, p,0.01. Conclusion: Measurements of MMPs and TIMPs in plasma may be of limited use. Despite similar plasma levels in SA patients and controls, the leukocyte-derived MMP-9 and TIMP-1 are significantly altered in patients. The findings indicate that th

Diffusion-weighted Imaging Reversibility In Stroke After Successful Mechanical Recanalization. Case report

Increased use of Diffusion-weighted imaging (DWI) in acute stroke has led to observations of early diffusion normalization in lesions thatinitially show diffusion slowing. The “renormalization” of DWI may be spontaneous or the result of thrombolytic therapy, thus, acuteslowing of diffusion is not necessarily an indicator of irreversible tissue damage. The perfusion-diffusion mismatch concept is attractiveas it assumes that DWI lesion size reflects the infarct core whilst the mismatch area reflects the penumbra. However, this concept maybe an oversimplification. This paper shows a case with Diffusion Lesion Reversal after successful neuroendovascular treatment andexcellent clinical outcome, and discuss the imaging characteristics associated with this phenomenon.</jats:p

Platypnea-Orthodeoxia Syndrome: Diagnostic Challenge and the Importance of Heightened Clinical Suspicion

Platypnea-orthodeoxia syndrome is an uncommon condition of positional dyspnea and hypoxemia; symptoms occur when the patient is upright and resolve with recumbency. Causes can be broadly categorized into 4 groups: intracardiac shunting, pulmonary shunting, ventilation-perfusion mismatch, or a combination of these. Platypnea-orthodeoxia syndrome should be suspected when normal arterial oxygen saturations are recorded while an individual is supine, followed by abrupt declines in those saturations when upright. Further investigations with use of imaging and cardiac catheterization aid in the evaluation. When platypnea-orthodeoxia syndrome is due to intracardiac shunting without pulmonary hypertension, intracardiac shunt closure can be curative. In this article, we report a case of platypnea-orthodeoxia syndrome in an 83-year-old woman who was successfully treated by means of percutaneous transcatheter closure of an atrial septal defect.</jats:p

Differences Between Normal and Leukemic Stem Cell-Specific Methylome Indicates Aberrantly Silenced Genes Involved in the Pathogenesis of Malignant Evolution

Abstract Due to technical limitations of methods used to study aberrant DNA hypermethylation, only a small numbers of promoters of empirically-selected genes have been investigated historically. Nevertheless, these studies demonstrated that aberrant methylation constitutes an important pathophysiologic mechanism of malignant evolution in MDS and AML. Novel methods utilizing genomic arrays allow for analysis of very large number of CpG islands and establishment of disease or tissues specific methylomes. In this study we hypothesized that a concordant methylation patterns exist that characterize hematopoietic progenitor and stem cells and genome-wide analysis of methylation patterns will allow for identification of a stem cell methylome that is distinct from that found in leukemia. To investigate this, chromatin immunoprecipitation promoter array-based profiling (ChIP-DSL 20,000 promoter array, Aviva Systems Biology) was used to identify genomewide methylated promoters in CD34 cells. We first sought to identify the normal stem cell methylome, using a pooled set of purified CD34 cells. We identified a relatively small set of 534 hypermethylated promoters (2.7% of genes analyzed with a cutoff of 2.6-fold methylation enrichment). Cytoskeleton remodeling, transcription regulation and nucleotide catabolism genes were significantly overrepresented (p=4.2E−5, p=2.2E−4, P=3.6E−3, Benjamini correction [BC] p=.008; p=.016; p=.069, respectively, using the DAVID database). When pooled CD34 cells were treated with decitabine; hypomethylation of 195 previously methylated genes (37%) was observed. The hypomethylated genes primarily functioned in basic metabolic processes (p=1.1E−4) and DNA repair/replication (p=3.0E−5). Subsequently, we analyzed triplicate samples of both immortalized AML cell lines (UT7, TF1, U937, HL60, KG1, K562 and Kazumi) and primary AML cells. Globally, the prevalence of hypermethylated sites in AML was infrequent: on average 2.8% or 552±181 of CpG islands were hypermethylated in cell lines compared to CD34 cells. Only 2431 (12%) promoters were hypermethylated in any AML cell line. Functional classification of hypermethylated genes showed that these genes were significantly enriched for cell development/differentiation (p=4.1E−5, BC p=.069), transcription regulation (p=4.8E−5, BC p=.027) and apoptosis (p=.01, BC p=.78) The methylation signature across several AML cell lines was variable: 115 genes were concordantly hypermethylated across the 7 cell lines and showed preponderance of transcription factor promoters (p=.0073) ); however, a functionally cancer-specific pattern was not obvious, suggesting that the establishment of phenotype in individual AML cell lines results from unique methylation events. Decitabine treatment of Kasumi cells resulted in hypomethylation of 75 genes involved in terminal differentiation (p=8.1E−4), transcription regulation (p=1.5E−2) and proliferation (p=4.3E−2). Analysis of the methylation pattern in AML patients produced similar results; 117 genes were concordantly hypermethylated in at least 60% of AML patients and were involved in mRNA transcription (p=.00073), leukocyte regulation (p=.007) and the Wnt signaling pathway (p=.01). Concordant hypermethylation of genes within Wnt pathways suggests involvement in leukemic evolution. In conclusion, the AML methylome is characterized by a highly variable hypermethylation of genes with consistent cellular functions including transcription, cell differentiation, apoptosis and leukocyte regulation.</jats:p