Unknown,Transcriptomics,Genomics,Proteomics

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Fever-range temperature alters continual efferocytosis mediated by mouse pro-inflammatory macrophages

ABSTRACT: Fever, a cardinal sign of inflammation, has been shown to modulate macrophage functions. Here, we wonder whether fever affects macrophage efferocytosis. This process is critical for the resolution of inflammation and the return to homeostasis with the reprogramming of macrophages toward a pro-resolving phenotype. Using primary mouse bone marrow-derived macrophages (BMDM) stimulated by LPS and IFN-γ (M1-like BMDM), we first validated that exposure to febrile-range temperature (39.5°C) induced a heat shock protein response. This was done by RNA sequencing (RNAseq), quantitative RT-PCR (RT-qPCR), and intracellular flow cytometry (FC). Then, we observed that febrile-range temperature decreased pro-inflammatory macrophage efferocytosis assessed by two different FC-based assays and by IncuCyte® live-cell imaging. This reduced efferocytic capacity of macrophages exposed to febrile-range temperature resulted from a decrease capacity to interact with apoptotic cells and to internalise these dying cells. This real-time image-based efferocytosis assay showed also that the heat reduced cell motility of macrophages in response to apoptotic cells. In our RNAseq dataset, we found an upregulation of the Adam17 gene and confirmed this increase by RT-qPCR analysis. Since this gene encodes a protease shedding the efferocytic receptor Mer, we determined Mer expression by FC and soluble Mer in the culture supernatants by ELISA. Febrile-range hyperthermia induced the cleavage of Mer from the cell surface of pro-inflammatory macrophages. Overall, decreased efferocytosis induced by fever-range hyperthermia may result from reduced Mer expression. This suggests that fever, by acting on Mer expression, decreases the efferocytic capacity of macrophages to prolong the pro-inflammatory state.

INSTRUMENT(S): DNBSEQ-G400

ORGANISM(S): Mus musculus

SUBMITTER: Mathieu VETTER

PROVIDER: E-MTAB-14408 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:<p>Definition of the human microbiome is an important scientific priority. This study will expand the scope of the investigation to include viruses, which account for a substantial proportion of infectious disease morbidity and mortality, especially in children. The long-term goal of this project is to describe the human virome in children and to investigate its relevance to febrile illnesses in children. The project will also seek to understand the relationship of the immune system to the composition of the virome. Thus, the project's specific aims are 1) To elucidate the spectrum of viruses that can be detected using non-biased, high throughput sequencing on samples of blood, respiratory, and gastrointestinal secretions from healthy children and to use this information as a basis for understanding the role of viruses in acute febrile illnesses without an obvious source, and 2) to investigate the effect of various forms of immunosuppression on the spectrum of viruses detected in children, and to use this information as a basis for understanding the role of viruses in acute febrile illnesses occurring in these children. Our preliminary studies show that diverse viruses can be detected in children having undiagnosed fever. To carry out the specific aims, well children will be enrolled prior to having elective surgery, and febrile otherwise well children will be enrolled from the Emergency Department at St. Louis Children's Hospital. Immunocompromised children will be recruited from hematopoietic stem cell and solid organ transplant clinics, the HIV/AIDS clinic, and the rheumatology/immunology clinic from the same hospital. Children with fever will have samples obtained at the time of the febrile illness and at 1 and 6-month follow-up visits. Selected samples from each study group will be analyzed at the Genome Center at Washington University (GCWU) using next generation 454 high throughput sequencing to detect and sequence all viral sequences present. We anticipate detecting and sequencing a broad range of viruses, including previously unrecognized agents. A variety of techniques will be used to investigate the significance of viruses detected. Virus-specific PCR assays will be used to determine the frequency and extent of viruses detected by sequencing, using the full range of samples collected. Host response to the detected viruses will be investigated using serologic analysis, cytokine profiling, and microarrays to characterize host gene expression. These studies will take advantage of follow-up samples to compare the acute response with the response in the convalescent period. This study will draw upon the expertise and technological assets of one of the world's most powerful sequencing centers to provide the research community with a comprehensive sequence data base of the viruses that are present in children, which can be used to improve our understanding of the causes of febrile illnesses in young children, many of which are currently undiagnosed.</p>

Project description:Background: We report the detailed development of biomarkers to predict the clinical outcome under dengue infection. Transcriptional signatures from purified peripheral blood mononuclear cells were derived from whole-genome gene-expression microarray data and validated by quantitative PCR and tested in independent samples. Methodology/Principal Findings: The study was performed on patients of a well-characterized dengue cohort from Recife, Brazil. The samples analyzed were collected prospectively from acute febrile dengue patients who evolved with different degrees of disease severity, classic dengue fever or dengue hemorrhagic fever (DHF) and compared with similar samples from other non-dengue febrile illnesses. The DHF samples were collected 2-3 days before the presentation of the plasma leakage symptoms. Differentially-expressed genes were selected by univariate statistical tests as well as multivariate classification techniques. The results showed that at early stages of dengue infection, the genes involved in effector mechanisms of innate immune response presented a weaker activation on patients who later developed hemorrhagic fever, whereas the genes involved in apoptosis were expressed in higher levels. Conclusions/Significance: Some of the gene expression signatures displayed estimated accuracy rates of more than 95%, indicating that expression profiling with these signatures may provide a useful means of DHF prognosis at early stages of infection The samples correspond to blood samples from 26 patients from a cohort in Brazil, divided into three classes, DHF, DF, ND, as follows: 18 were confirmed dengue 3, genotype III cases, among which 10 were diagnosed as dengue hemorrhagic fever (DHF) and 8 as classical dengue fever (DF), and 8 control samples (ND) from febrile patients confirmed to be not infected with dengue. None of the DHF patients presented vasculopathy signs and symptoms at the time the samples used in the functional genomic characterization were collected. At the time of collection the patients referred approximately 5 days of disease and the absence of fever was reported two to three days after enrollment. The samples from the DF, DHF and ND patients were matched to avoid significant differences regarding patient age, gender, dengue infection history and days of symptoms among the groups.

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Fever-range temperature alters continual efferocytosis mediated by mouse pro-inflammatory macrophages

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