Browsing by Autor "Javiera Villar"

Filter results by typing the first few letters
Now showing 1 - 2 of 2
  • Thumbnail Image
    Item type: Item ,
    Hemocyanins Stimulate Innate Immunity by Inducing Different Temporal Patterns of Proinflammatory Cytokine Expression in Macrophages
    (American Association of Immunologists, 2016) Ta-Ying Zhong; Sergio Arancibia; Raimundo Born; Robert Tampé; Javiera Villar; Miguel Del Campo; Augusto Manubens; Marı́a Inés Becker
    Hemocyanins induce a potent Th1-dominant immune response with beneficial clinical outcomes when used as a carrier/adjuvant in vaccines and nonspecific immunostimulant in cancer. However, the mechanisms by which hemocyanins trigger innate immune responses, leading to beneficial adaptive immune responses, are unknown. This response is triggered by a proinflammatory signal from various components, of which macrophages are an essential part. To understand how these proteins influence macrophage response, we investigated the effects of mollusks hemocyanins with varying structural and immunological properties, including hemocyanins from Concholepas concholepas, Fissurella latimarginata, and Megathura crenulata (keyhole limpet hemocyanin), on cultures of peritoneal macrophages. Hemocyanins were phagocytosed and slowly processed. Analysis of this process showed differential gene expression along with protein levels of proinflammatory markers, including IL-1β, IL-6, IL-12p40, and TNF-α. An extended expression analysis of 84 cytokines during a 24-h period showed a robust proinflammatory response for F. latimarginata hemocyanin in comparison with keyhole limpet hemocyanin and C. concholepas hemocyanin, which was characterized by an increase in the transcript levels of M1 cytokines involved in leukocyte recruitment. These cytokine genes included chemokines (Cxcl1, Cxcl3, Cxcl5, Ccl2, and Ccl3), ILs (Il1b and Ifng), growth factors (Csf2 and Csf3), and TNF family members (Cd40lg). The protein levels of certain cytokines were increased. However, every hemocyanin maintains downregulated key M2 cytokine genes, including Il4 and Il5 Collectively, our data demonstrate that hemocyanins are able to trigger the release of proinflammatory factors with different patterns of cytokine expression, suggesting differential signaling pathways and transcriptional network mechanisms that lead to the activation of M1-polarized macrophages.
  • Thumbnail Image
    Item type: Item ,
    Mechanisms underlying the mollusk hemocyanin processing and presentation through MHC-dependent pathways in antigen presenting cells of mammals
    (American Association of Immunologists, 2022) Marı́a Inés Becker; Michelle L. Salazar; Diego A. Díaz-Dinamarca; Abel E. Vásquez; Javiera Villar; Alejandra Alvarado; Byron Castillo; Daniel Navarro; Fabián Salazar; Augusto Manubens
    Abstract Hemocyanins are oligomeric glycoproteins widely used as immunomodulators because they bias immunity towards a Th1 profile when inoculated in mammals. We have demonstrated that hemocyanins are internalized through receptor-mediated endocytosis, and TLR4 and C-type lectin receptors (MR, DC-SIGN, MGL) participate in the hemocyanin-mediated proinflammatory response in mouse and human dendritic cells (DCs). However, despite the massive use of hemocyanins, their intracellular processing route for MHC presentation to T lymphocytes has been scarcely studied. Therefore, we hypothesized that hemocyanins follow the MHC-II pathway as a classical T-cell-dependent antigen. Interestingly, our results analyzing the processing pathway of hemocyanins in mouse DCs showed that hemocyanins from Fissurella latimarginata (FLH) and Megathura crenulata (KLH) co-localized with Rab5+, Rab7+, and Lamp-1+ compartments. This observation strongly suggests that hemocyanins could be cross-presented by MHC-I molecules. Furthermore, DCs incubated with FLH showed an increase in the percentage of MHC-I+ cells versus the control cells. FLH-induced cytokine secretion decreased in J774.2 macrophages treated with pharmacological inhibitors of both MHC-II and MHC-I pathways, supporting our previous results on hemocyanin cross-presentation but also the MHC-II pathway. Furthermore, immunoblot confirmed different FLH proteolysis patterns in macrophages treated with MHC-I and MHC-II pathway inhibitors. Hence, we postulate that hemocyanins undergo both MHC-I and MHC-II dependent antigen presentation pathways in antigen-presenting cells. These findings offer molecular clues to underlying hemocyanin processing and presentation mechanisms. Supported by grants from FONDECYT N° 1151337 and N° 1201600 (MIB), ANID/Beca Doctorado Nacional N° 21210946 (MLS) and N° 21200880 (DDD).