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Pyruvate affects inflammatory responses of macrophages during influenza A virus infection.

ABSTRACT: Pyruvate is the end product of glycolysis and transported into the mitochondria for use in the tricarboxylic acid (TCA) cycle. It is also a common additive in cell culture media. We discovered that inclusion of sodium pyruvate in culture media during infection of mouse bone marrow derived macrophages with influenza A virus impaired cytokine production (IL-6, IL-1?, and TNF-?). Sodium pyruvate did not inhibit viral RNA replication. Instead, the addition of sodium pyruvate alters cellular metabolism and diminished mitochondrial reactive oxygen species (ROS) production and lowered immune signaling. Overall, sodium pyruvate affects the immune response produced by macrophages but does not inhibit virus replication.

SUBMITTER: Abusalamah H

PROVIDER: S-EPMC7345311 | biostudies-literature | 2020 Jul

REPOSITORIES: biostudies-literature

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Pyruvate affects inflammatory responses of macrophages during influenza A virus infection.

Abusalamah Hazar H Reel Jessica M JM Lupfer Christopher R CR

Virus research 20200704

Pyruvate is the end product of glycolysis and transported into the mitochondria for use in the tricarboxylic acid (TCA) cycle. It is also a common additive in cell culture media. We discovered that inclusion of sodium pyruvate in culture media during infection of mouse bone marrow derived macrophages with influenza A virus impaired cytokine production (IL-6, IL-1β, and TNF-α). Sodium pyruvate did not inhibit viral RNA replication. Instead, the addition of sodium pyruvate alters cellular metaboli ...[more]

PMID: 32634445

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Project description:Influenza A viruses cause infections in the human respiratory tract and give rise to annual seasonal outbreaks, as well as more rarely dreaded pandemics. Influenza A viruses become quickly resistant to the virus-directed antiviral treatments, which are the current main treatment options. A promising alternative approach is to target host cell factors that are exploited by influenza viruses. To this end, we characterized the phosphoproteome of influenza A virus infected primary human macrophages to elucidate the intracellular signaling pathways and critical host factors activated upon influenza infection. We identified 1675 phosphoproteins, 4004 phosphopeptides and 4146 nonredundant phosphosites. The phosphorylation of 1113 proteins (66%) was regulated upon infection, highlighting the importance of such global phosphoproteomic profiling in primary cells. Notably, 285 of the identified phosphorylation sites have not been previously described in publicly available phosphorylation databases, despite many published large-scale phosphoproteome studies using human and mouse cell lines. Systematic bioinformatics analysis of the phosphoproteome data indicated that the phosphorylation of proteins involved in the ubiquitin/proteasome pathway (such as TRIM22 and TRIM25) and antiviral responses (such as MAVS) changed in infected macrophages. Proteins known to play roles in small GTPase-, mitogen-activated protein kinase-, and cyclin-dependent kinase- signaling were also regulated by phosphorylation upon infection. In particular, the influenza infection had a major influence on the phosphorylation profiles of a large number of cyclin-dependent kinase substrates. Functional studies using cyclin-dependent kinase inhibitors showed that the cyclin-dependent kinase activity is required for efficient viral replication and for activation of the host antiviral responses. In addition, we show that cyclin-dependent kinase inhibitors protect IAV-infected mice from death. In conclusion, we provide the first comprehensive phosphoproteome characterization of influenza A virus infection in primary human macrophages, and provide evidence that cyclin-dependent kinases represent potential therapeutic targets for more effective treatment of influenza infections.

Project description:BackgroundExpression of host microRNAs (miRNAs) changes markedly during influenza A virus (IAV) infection of natural and adaptive hosts, but their role in genetically determined host susceptibility to IAV infection has not been explored. We, therefore, compared pulmonary miRNA expression during IAV infection in two inbred mouse strains with differential susceptibility to IAV infection.ResultsmiRNA expression profiles were determined in lungs of the more susceptible strain DBA/2J and the less susceptible strain C57BL/6J within 120 h post infection (hpi) with IAV (H1N1) PR8. Even the miRNomes of uninfected lungs differed substantially between the two strains. After a period of relative quiescence, major miRNome reprogramming was detected in both strains by 48 hpi and increased through 120 hpi. Distinct groups of miRNAs regulated by IAV infection could be defined: (1) miRNAs (n = 39) whose expression correlated with hemagglutinin (HA) mRNA expression and represented the general response to IAV infection independent of host genetic background; (2) miRNAs (n = 20) whose expression correlated with HA mRNA expression but differed between the two strains; and (3) remarkably, miR-147-3p, miR-208b-3p, miR-3096a-5p, miR-3069b-3p, and the miR-467 family, whose abundance even in uninfected lungs differentiated nearly perfectly (area under the ROC curve > 0.99) between the two strains throughout the time course, suggesting a particularly strong association with the differential susceptibility of the two mouse strains. Expression of subsets of miRNAs correlated significantly with peripheral blood granulocyte and monocyte numbers, particularly in DBA/2J mice; miR-223-3p, miR-142-3p, and miR-20b-5p correlated most positively with these cell types in both mouse strains. Higher abundance of antiapoptotic (e.g., miR-467 family) and lower abundance of proapoptotic miRNAs (e.g., miR-34 family) and those regulating the PI3K-Akt pathway (e.g., miR-31-5p) were associated with the more susceptible DBA/2J strain.ConclusionSubstantial differences in pulmonary miRNA expression between the two differentially susceptible mouse strains were evident even before infection, but evolved further throughout infection and could in part be attributed to differences in peripheral blood leukocyte populations. Thus, pulmonary miRNA expression both before and during IAV infection is in part determined genetically and contributes to susceptibility to IAV infection in this murine host, and likely in humans.

Project description:Airway epithelial cells and macrophages differ markedly in their responses to influenza A virus (IAV) infection. To investigate transcriptional responses underlying these differences, purified subsets of type II airway epithelial cells (ATII) and alveolar macrophages (AM) recovered from the lungs of mock- or IAV-infected mice at 9 h postinfection were subjected to RNA sequencing. This time point was chosen to allow for characterization of cell types first infected with the virus inoculum, prior to multicycle virus replication and the infiltration of inflammatory cells into the airways. In the absence of infection, AM predominantly expressed genes related to immunity, whereas ATII expressed genes consistent with their physiological roles in the lung. Following IAV infection, AM almost exclusively activated cell-intrinsic antiviral pathways that were dependent on interferon (IFN) regulatory factor 3/7 (IRF3/7) and/or type I IFN signaling. In contrast, IAV-infected ATII activated a broader range of physiological responses, including cell-intrinsic antiviral pathways, which were both independent of and dependent on IRF3/7 and/or type I IFN. These data suggest that transcriptional profiles hardwired during development are a major determinant underlying the different responses of ATII and AM to IAV infection.IMPORTANCE Airway epithelial cells (AEC) and airway macrophages (AM) represent major targets of influenza A virus (IAV) infection in the lung, yet the two cell types respond very differently to IAV infection. We have used RNA sequencing to define the host transcriptional responses in each cell type under steady-state conditions as well as following IAV infection. To do this, different cell subsets isolated from the lungs of mock- and IAV-infected mice were subjected to RNA sequencing. Under steady-state conditions, AM and AEC express distinct transcriptional activities, consistent with distinct physiological roles in the airways. Not surprisingly, these cells also exhibited major differences in transcriptional responses following IAV infection. These studies shed light on how the different transcriptional architectures of airway cells from two different lineages drive transcriptional responses to IAV infection.

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Pyruvate affects inflammatory responses of macrophages during influenza A virus infection.

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Pyruvate affects inflammatory responses of macrophages during influenza A virus infection.

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