Lorena Guasconi

Involvement of a mitochondrial pathway and key role of hydrogen peroxide during eosinophil apoptosis induced by excretory–secretory products from Fasciola hepatica

Eosinophils (Eo) are typically associated with immune response to helminth. Previously, we demonstrated that excretory-secretory products (ESP) from Fasciola hepatica induce eosinophil apoptosis by a caspase-dependent mechanism. In this study, we observed that ESP caused mitochondrial-membrane depolarization of eosinophils leading to the release of cytochrome c. Also, ESP induced an increase in the reactive oxygen species (ROS) production, which preceded the mitochondrial injury. We found a significant rise in hydrogen peroxide, but not in the anion superoxide levels. Furthermore, catalase, but not superoxide dismutase, inhibited the mitochondrial depolarization as well as apoptosis. So, ESP induce in Eo an early increase in the ROS production, mainly hydrogen peroxide, which precedes mitochondrial injury and leads again to apoptosis. Finally, we demonstrated the participation of hydrogen peroxide in the peritoneal Eo apoptosis in vivo, both during the early stages of experimental fasciolosis in rats and after intraperitoneal ESP treatment.

C-type lectins on macrophages participate in the immunomodulatory response to Fasciola hepatica products

Fasciola hepatica releases excretory–secretory products (FhESP), and immunomodulatory properties have been described for the carbohydrates present in these parasite products. The interaction of FhESP with the innate immune cells, such as macrophages, is crucial in the early stage of infection. In this work we observed that peritoneal macrophages from naive BALB/c mice stimulated in vitro with FhESP presented: an increased arginase activity as well as Arginase I expression, and high levels of transforming growth factor‐β and interleukin‐10. A similar macrophage population was also observed in the peritoneum of infected mice. A partial inhibition of the immunomodulatory effects described above was observed when macrophages were pre‐incubated with Mannan, anti‐mannose receptor, Laminarin or anti‐Dectin‐1, and then stimulated with FhESP. In addition, we observed a partial inhibition of these effects in macrophages obtained from mice that were intraperitoneally injected with Mannan or Laminarin before being infected. Taken together, these results suggest the participation of at least two C‐type lectin receptors, mannose receptor and Dectin‐1, in the interaction of FhESP with macrophages, which allows this parasite to induce immunoregulatory effects on these important innate immune cells and may constitute a crucial event for extending its survival in the host.

Fasciola hepatica products induce apoptosis of peritoneal macrophages

Immunomodulatory properties have been described for Fasciola hepatica excretory-secretory products (FhESP), with their interaction with the innate immune cells being crucial during the early stages of infection. Previously, we demonstrated that FhESP induce eosinophil apoptosis. In this work, the ability of FhESP to induce apoptosis of peritoneal macrophages was evaluated. These parasite products were observed to induce apoptosis in peritoneal macrophages stimulated in vitro with FhESP, as well as in cells recovered from infected mice. The ability of FhESP to modify the viability of macrophages by apoptosis induction may constitute a crucial event for extending its survival in the host.

Fasciola hepatica excretory-secretory products induce CD4+T cell anergy via selective up-regulation of PD-L2 expression on macrophages in a Dectin-1 dependent way

Fasciola hepatica excretory-secretory products (FhESP) induce immunomodulatory effects on macrophages. Previously, we demonstrated that these effects are dependent on Dectin-1. Therefore, the aim of this study was to determine how this affects the CD4 T-cells immune response. We observed that FhESP induce an increased expression of PD-L2 in macrophages via Dectin-1. Furthermore, in co-cultures with CD4 T-cell we observed a suppressive effect on proliferative response, down-modulation of IFN-γ and up-modulation of IL-10 via Dectin-1 on macrophages. These results suggest that FhESP induce T-cell anergy via selective up-regulation of PD-L2 expression on macrophages in a Dectin-1 dependent way.

New Insights into the Modulation of Immune Response by Fasciola hepatica Excretory-Secretory Products

Fasciola hepatica is a trematode that affects human and domestic ruminant health, causing significant economic losses in cattle estimated at US

Immunization with crude antigens plus aluminium hydroxide protects cattle from Fasciola hepatica infection

2000 millon per year. Juvenile parasites migrating through the host tissues, as well as adults settle in the biliary ducts, are in contact with different cells from the immune system. Despite those interactions, the persistence of the parasite in the host for many years provides evidence of its ability to prevent or down- modulate the inflammatory response in the infection site. Different strategies have been developed by the parasite to prevent potential damage being induced by the immune response, thus allowing some parasites to reach the adult stage in a safe place such as the biliary ducts. In this review we discuss how excretory-secretory products (ESP) from F. hepatica can affect the functionality of pivotal immune cells, such as eosinophils and macrophages by inducing selective apoptosis pathways and alternative activation of macrophages. Furhermore, the modulatory effects of ESP on dendritic cell activation and lymphocyte proliferation is reviewed as a strategy to facilitate F. hepatica evasion of both innate and adaptive immunity.

Dectin-1 on macrophages modulates the immune response to Fasciola hepatica products through the ERK signaling pathway

The ability of total homogenate (TH) of Fasciola hepatica conjugated with aluminium hydroxide (alum) or Freunds complete adjuvant (FCA) to protect cattle against experimental fasciolosis was evaluated. Compared with the infected group, the immunized animals with alum-TH and FCA-TH presented a significant reduction in fluke burden (85.9% and 96.8%, respectively), a higher percentage of short-sized worms, a marked reduction in the released eggs in faeces (89% and 57%, respectively), as well as an increased production of specific antibodies before infection. The alum-TH immunized group also showed a significant increase in the antigen-specific proliferation of peripheral blood mononuclear cells (PBMC) as early as 4 weeks before infection. Although both immunized groups (alum-TH and FCA-TH) were able to develop an efficient protective immune response to metacercarial challenge, an earlier PBMC response, lower hepatic damage and less effect on weight gain were found in alum-immunized animals. Therefore, alum is a good candidate for future immunization against bovine fasciolosis.

Excretory-secretory products from Fasciola hepatica induce eosinophil apoptosis by a caspase-dependent mechanism

Fasciolosis is a zoonotic disease of increasing importance due to its worldwide distribution and elevated economic losses. Previously, we demonstrated that Fasciola hepatica excretory-secretory products (FhESP) induce immunomodulatory effects on peritoneal macrophages in a Dectin-1 dependent manner. In this study, we observed that peritoneal macrophages from naive BALB/c mice stimulated in vitro with FhESP presented increased expression levels of phosphorylated extracellular-signal-regulated kinase (ERK), and this effect was dependent on Syk, protein kinase C (PKC) and Dectin-1. In this sense, we observed increased levels of arginase activity, IL-10 and TGF-β in macrophages stimulated with FhESP, which were dependent on PKC and ERK. Furthermore, we observed that the increased arginase activity, as well as in TGF-β and IL-10 levels, was partially dependent on IL-10 receptor signaling in macrophages that were pre-incubated with anti-IL10R before being stimulated with FhESP. Taken together, these results suggest the participation of Dectin-1 and Syk in FhESP interaction with peritoneal macrophages and the possible role of ERK and IL-10 in downstream signaling pathways involved in the immunomodulatory effects induced by Fasciola hepatica products.