Myocardial Ischemia and Reperfusion Leads to Transient CD8 Immune Deficiency and Accelerated Immunosenescence in CMV-Seropositive Patients

Rationale: There is mounting evidence of a higher incidence of coronary heart disease (CHD) in cytomegalovirus (CMV) seropositive individuals. Objective: The aim of this study was to investigate whether acute MI triggers an inflammatory T-cell response that might lead to accelerated immunosenescence in CMV-seropositive patients. Methods and Results: Thirty-four patients with acute MI undergoing primary PCI (PPCI) were longitudinally studied within 3 months following reperfusion (Cohort A). In addition, 54 patients with acute and chronic MI were analyzed in a cross-sectional study (Cohort B). CMV-seropositive patients demonstrated a greater fall in the concentration of terminally differentiated CD8 effector memory T cells (TEMRA) in peripheral blood during the first 30 min of reperfusion compared with CMV-seronegative patients (-192 vs. -63 cells/µl; p=0.008), correlating with the expression of programmed cell death-1 (PD-1) before PPCI (r=0.8; p=0.0002). A significant proportion of TEMRA cells remained depleted for at least 3 months in CMV-seropositive patients. Using high-throughput 13-parameter flow cytometry and HLA class I CMV-specific dextramers, we confirmed an acute and persistent depletion of terminally differentiated TEMRA and CMV-specific CD8+ cells in CMV-seropositive patients. Long-term reconstitution of the TEMRA pool in chronic CMV-seropositive post-MI patients was associated with signs of terminal differentiation including an increase in KLRG1 and shorter telomere length in CD8+ T cells (2225 bp vs. 3397 bp; p<0.001). Conclusions: Myocardial ischemia and reperfusion in CMV-seropositive patients undergoing PPCI leads to acute loss of antigen-specific, terminally differentiated CD8 T-cells, possibly through PD-1-dependent programmed cell death. Our results suggest that acute MI and reperfusion accelerate immunosenescence in CMV-seropositive patients

Homeobox gene expression in acute myeloid leukemia is linked to typical underlying molecular aberrations

Background: Although distinct patterns of homeobox (HOX) gene expression have been described in defined cytogenetic and molecular subsets of patients with acute myeloid leukemia (AML), it is unknown whether these patterns are the direct result of transcriptional alterations or rather represent the differentiation stage of the leukemic cell. Method: To address this question, we used qPCR to analyze mRNA expression of HOXA and HOXB genes in bone marrow (BM) samples of 46 patients with AML and sorted subpopulations of healthy BM cells. These various stages of myeloid differentiation represent matched counterparts of morphological subgroups of AML. To further study the transcriptional alterations of HOX genes in hematopoiesis, we also analyzed gene expression of epigenetic modifiers in the subpopluations of healthy BM and leukemic cells. Results: Unsupervised hierarchical clustering divided the AMLs into five clusters characterized by the presence of prevalent molecular genetic aberrations. Notably, the impact of genotype on HOX gene expression was significantly more pronounced than that of the differentiation stage of the blasts. This driving role of molecular aberrations was best exemplified by the repressive effect of the PML-RARa fusion gene on HOX gene expression, regardless of the presence of the FLT3/ITD mutation. Furthermore, HOX gene expression was positively correlated with mRNA levels of histone demethylases (JMJD3 and UTX) and negatively correlated with gene expression of DNA methyltranferases. No such relationships were observed in subpopulations of healthy BM cells. Conclusion: Our results demonstrate that specific molecular genetic aberrations, rather than differentiation per se, underlie the observed differences in HOX gene expression in AML. Moreover, the observed correlations between epigenetic modifiers and HOX ex pression that are specific to malignant hematopoiesis, suggest their potential causal relationships.</p

Homeobox gene expression in acute myeloid leukemia is linked to typical underlying molecular aberrations

__Background:__ Although distinct patterns of homeobox (HOX) gene expression have been described in defined cytogenetic and molecular subsets of patients with acute myeloid leukemia (AML), it is unknown whether these patterns are the direct result of transcriptional alterations or rather represent the differentiation stage of the leukemic cell. __Method:__ To address this question, we used qPCR to analyze mRNA expression of HOXA and HOXB genes in bone marrow (BM) samples of 46 patients with AML and sorted subpopulations of healthy BM cells. These various stages of myeloid differentiation represent matched counterparts of morphological subgroups of AML. To further study the transcriptional alterations of HOX genes in hematopoiesis, we also analyzed gene expression of epigenetic modifiers in the subpopluations of healthy BM and leukemic cells. __Results:__ Unsupervised hierarchical clustering divided the AMLs into five clusters characterized by the presence of prevalent molecular genetic aberrations. Notably, the impact of genotype on HOX gene expression was significantly more pronounced than that of the differentiation stage of the blasts. This driving role of molecular aberrations was best exemplified by the repressive effect of the PML-RARa fusion gene on HOX gene expression, regardless of the presence of the FLT3/ITD mutation. Furthermore, HOX gene expression was positively correlated with mRNA levels of histone demethylases (JMJD3 and UTX) and negatively correlated with gene expression of DNA methyltranferases. No such relationships were observed in subpopulations of healthy BM cells. __Conclusion:__ Our results demonstrate that specific molecular genetic aberrations, rather than differentiation per se, underlie the observed differences in HOX gene expression in AML. Moreover, the observed correlations between epigenetic modifiers and HOX ex pression that are specific to malignant hematopoiesis, suggest their potential causal relationships

CD maps—dynamic profiling of CD1–CD100 surface expression on human leukocyte and lymphocyte subsets

CD molecules are surface molecules expressed on cells of the immune system that play key roles in immune cell-cell communication and sensing the microenvironment. These molecules are essential markers for the identification and isolation of leukocytes and lymphocyte subsets. Here, we present the results of the first phase of the CD Maps study, mapping the expression of CD1–CD100 (n = 110) on 47 immune cell subsets from blood, thymus, and tonsil using an eight-color standardized EuroFlow approach and quantification of expression. The resulting dataset included median antibody binding capacities (ABCs) and percentage of positivity for all markers on all subsets and was developed into an interactive CD Maps web resource. Using the resource, we examined differentially expressed proteins between granulocyte, monocyte, and dendritic cell subsets, and profiled dynamic expression of markers during thymocyte differentiation, T-cell maturation, and between functionally distinct B-cell subset clusters. The CD Maps resource will serve as a benchmark of antibody reactivities ensuring improved reproducibility of flow cytometry-based research. Moreover, it will provide a full picture of the surfaceome of human immune cells and serves as a useful platform to increase our understanding of leukocyte biology, as well as to facilitate the identification of new biomarkers and therapeutic targets of immunological and hematological diseases

Interactive Dendrograms: The R Packages idendro and idendr0

Hierarchical cluster analysis is a valuable tool for exploring data by describing their structure using a dendrogram. However, proper visualization and interactive inspection of the dendrogram are needed to unlock the information in the data. We describe a new R package, idendro, that enables the user to inspect dendrograms interactively: to select and color clusters, to zoom and pan the dendrogram, and to visualize the clustered data not only in a built-in heat map, but also in any interactive plot implemented in the cranvas package. A lightweight version idendr0 with reduced dependencies is also available from the Comprehensive R Archive Network

Interactive Dendrograms: The R Packages idendro and idendr0

Hierarchical cluster analysis is a valuable tool for exploring data by describing their structure using a dendrogram. However, proper visualization and interactive inspection of the dendrogram are needed to unlock the information in the data. We describe a new R package, idendro, that enables the user to inspect dendrograms interactively: to select and color clusters, to zoom and pan the dendrogram, and to visualize the clustered data not only in a built-in heat map, but also in any interactive plot implemented in the cranvas package. A lightweight version idendr0 with reduced dependencies is also available from the Comprehensive R Archive Network

What matters in chronic <i>Burkholderia cenocepacia</i> infection in cystic fibrosis: Insights from comparative genomics

<div><p><i>Burkholderia cenocepacia</i> causes severe pulmonary infections in cystic fibrosis (CF) patients. Since the bacterium is virtually untreatable by antibiotics, chronic infections persist for years and might develop into fatal septic pneumonia (cepacia syndrome, CS). To devise new strategies to combat chronic <i>B</i>. <i>cenocepacia</i> infections, it is essential to obtain comprehensive knowledge about their pathogenesis. We conducted a comparative genomic analysis of 32 Czech isolates of epidemic clone <i>B</i>. <i>cenocepacia</i> ST32 isolated from various stages of chronic infection in 8 CF patients. High numbers of large-scale deletions were found to occur during chronic infection, affecting preferentially genomic islands and nonessential replicons. Recombination between insertion sequences (IS) was inferred as the mechanism behind deletion formation; the most numerous IS group was specific for the ST32 clone and has undergone transposition burst since its divergence. Genes functionally related to transition metal metabolism were identified as hotspots for deletions and IS insertions. This functional category was also represented among genes where nonsynonymous point mutations and indels occurred parallelly among patients. Another category exhibiting parallel mutations was oxidative stress protection; mutations in catalase KatG resulted in impaired detoxification of hydrogen peroxide. Deep sequencing revealed substantial polymorphism in genes of both categories within the sputum <i>B</i>. <i>cenocepacia</i> ST32 populations, indicating extensive adaptive evolution. Neither oxidative stress response nor transition metal metabolism genes were previously reported to undergo parallel evolution during chronic CF infection. Mutations in <i>katG</i> and copper metabolism genes were overrepresented in patients where chronic infection developed into CS. Among professional phagocytes, macrophages use both hydrogen peroxide and copper for their bactericidal activity; our results thus tentatively point to macrophages as suspects in pathogenesis towards the fatal CS.</p></div

Parallel mutations in <i>B</i>. <i>cenocepacia</i> ST32 sputum populations from 12 chronically infected, non-CS patients.

<p>Parallel mutations in <i>B</i>. <i>cenocepacia</i> ST32 sputum populations from 12 chronically infected, non-CS patients.</p

Overview of sequenced <i>B</i>. <i>cenocepacia</i> ST32 isolates.

<p>(A) Sample collection chart. Bacteria were isolated at indicated time points and named by patient IDs (1 to 8) and chronology of isolation A to D (i.e., isolates 1A to 1D, 2A to 2D etc.). Isolates 2C and 2D, 5C and 5D, 7C and 7D, and 8C and 8D overlapped due to their concurrent isolation at the time of CS (see <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006762#ppat.1006762.s004" target="_blank">S1 Table</a> for details). (B) Whole-genome phylogeny. Consensus genomic sequences obtained by mapping sequencing reads onto the complete reference genome were used for tree inference. The Maximum-Likelihood tree was constructed from 2,754 variant nucleotide positions using the CSIPhylogeny pipeline [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006762#ppat.1006762.ref074" target="_blank">74</a>]. (C) SNP accumulation during chronic infection. SNPs informative of within-patient evolution (see <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006762#sec012" target="_blank">Materials and Methods</a>) were plotted against time elapsed from the first Bcc culture positivity to the point of bacterial isolation.</p

Localization of mutations in proteins under parallel evolution during chronic infection.

<p>Protein chains under parallel evolution are denoted in light blue, their functional domains are denoted in dark blue (DHp in CusS, βi4 in RpoB). Cofactors are denoted as forest green spheres, catalytic and active site amino acids are denoted as lime green spheres. Amino acids homologous to residues affected by mutations during chronic Bcc infection (<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006762#ppat.1006762.s013" target="_blank">S10 Table</a>) are denoted as spheres and colored as explained in the legend (for ST32 WGS data, see <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006762#ppat.1006762.g003" target="_blank">Fig 3</a>; for ST32 DPS data, see <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006762#ppat.1006762.t001" target="_blank">Table 1</a>; for IIIA WGS data, see <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006762#ppat.1006762.s012" target="_blank">S9 Table</a>). Visualizations were carried out in Chimera [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006762#ppat.1006762.ref083" target="_blank">83</a>].</p