Oxidative stress in canine histiocytic sarcoma cells (DH82 cells) induced by a persistent canine distemper virus infection leads to impairment of the HIF-1α downstream pathway in vitro

Introduction Histiocytic sarcoma (HS) is a highly aggressive neoplasm with a limited response to therapies. A promising new approach might be oncolytic virotherapy. Angiogenesis is essential for tumor growth and metastasis. One of the key regulator factors for angiogenesis is hypoxia inducible factor-1α (HIF-1α). In addition, angiogenesis is regulated by reactive oxygen species (ROS). The aim of this study was to evaluate the influence of canine distemper virus (CDV) infection on HIF-1α expression in canine HS cells (DH82 cells). Materials and methods Non-infected and persistently CDV infected DH82 cells were investigated in vitro. The expression of oxidative stress and angiogenesis markers were evaluated on a molecular and protein level using microarray data, immunofluorescence microscopy, Western blot and flow cytometry. Results Persistently CDV-infected DH82 cells displayed an increased oxidative stress due to viral infection that leads to a dysregulated HIF-1α expression with a consecutive downregulation of the downstream angiogenetic pathway. Conclusion These results indicate that a persistent CDV-infection seems to affect the HIF-1α pathway resulting in a decrease in the downstream production of angiogenetic factors in vitro

Pragmatic guide to Subversion

Macrophages are a heterogeneous cell population playing a pivotal role in tissue homeostasis and inflammation, and their phenotype strongly depends on the micromilieu. Despite its increasing importance as a translational animal model for human diseases, there is a considerable gap of knowledge with respect to macrophage polarization in dogs. The present study comprehensively investigated the morphologic, phenotypic, and transcriptomic characteristics of unstimulated (M0), M1- (GM-CSF, LPS, IFNγ-stimulated) and M2- (M-CSF, IL-4-stimulated)-polarized canine blood-derived macrophages in vitro. Scanning electron microscopy revealed distinct morphologies of polarized macrophages with formation of multinucleated cells in M2-macrophages, while immunofluorescence employing literature-based prototype-antibodies against CD16, CD32, iNOS, MHC class II (M1-markers), CD163, CD206, and arginase-1 (M2-markers) demonstrated that only CD206 was able to discriminate M2-macrophages from both other phenotypes, highlighting this molecule as a promising marker for canine M2-macrophages. Global microarray analysis revealed profound changes in the transcriptome of polarized canine macrophages. Functional analysis pointed out that M1-polarization was associated with biological processes such as "respiratory burst", whereas M2-polarization was associated with processes such as "mitosis". Literature-based marker gene selection revealed only minor overlaps in the gene sets of the dog compared to prototype markers of murine and human macrophages. Biomarker selection using supervised clustering suggested latexin (LXN) and membrane-spanning 4-domains, subfamily A, member 2 (MS4A2) to be the most powerful predicting biomarkers for canine M1- and M2-macrophages, respectively. Immunofluorescence for both markers demonstrated expression of both proteins by macrophages in vitro but failed to reveal differences between canine M1 and M2-macrophages. The present study provides a solid basis for future studies upon the role of macrophage polarization in spontaneous diseases of the dog, a species that has emerging importance for translational research