Possible Role for Toll-Like Receptors in Interaction of Fasciola hepatica Excretory-Secretory Products with Human Monocyte Cell Line.

This chapter presents a proteomic approach to purify and identify native excretory-secretory products (ESPs) in the range of >10-30 kDa proteins capable of interacting with toll-like receptors (TLRs). Here we present a protocol to fractionate the total ESPs using an ultrafiltration system to recover ESP proteins >10-30 kDa. The fraction of the proteins >10-30 kDa is purified by ion exchange chromatography (IEC) using a mono Q-column in a fast protein liquid chromatography system (FPLC) to separate its components based on charge.

Purification of Native Fasciola hepatica Fatty Acid-Binding Protein and Induction of Alternative Activation of Human Macrophages.

This chapter presents the different techniques to purify the native forms of Fasciola hepatica fatty acid-binding protein (Fh12) using size exclusion chromatography and isoelectric focusing (IEF). Also, it presents the procedure to study the immunological effect of the purified protein Fh12 using monocyte-derived macrophages (MDM) obtained from healthy human donors. For this purpose, I present the procedure to isolate and culture peripheral blood mononuclear cells (PBMCs) to generate alternatively activated macrophages (AAMΦ) by in vitro exposure to Fh12.

ESPs Could Indistinctly Activate or Block Multiple Toll-Like Receptors in a Human Monocyte Cell Line.

is a parasitic helminth that induces Th2/Treg responses in its mammalian host. Some reports have suggested that ESPs achieve these polarized immune responses by delaying the activation of dendritic cells and macrophages during the early stages of innate immunity, a process that is mediated by TLR4. The present study aimed to investigate whether TLRs other than TLR4 could also be targeted by ESPs.

Fasciola hepatica fatty acid binding protein inhibits TLR4 activation and suppresses the inflammatory cytokines induced by lipopolysaccharide in vitro and in vivo.

TLR4, the innate immunity receptor for bacterial endotoxins, plays a pivotal role in the induction of inflammatory responses. There is a need to develop molecules that block either activation through TLR4 or the downstream signaling pathways to inhibit the storm of inflammation typically elicited by bacterial LPS, which is a major cause of the high mortality associated with bacterial sepsis. We report in this article that a single i.

Fasciola hepatica fatty acid binding protein induces the alternative activation of human macrophages.

The liver fluke Fasciola hepatica is a highly evolved parasite that uses sophisticated mechanisms to evade the host immune response. The immunosuppressive capabilities of the parasite have been associated with antigens secreted through the parasite's tegument, called excretory-secretory products (ESPs). Proteomic studies have identified the fatty acid binding protein (FABP) as one of molecules present in the parasite ESPs.

Fasciola hepatica saposin-like protein-2-based ELISA for the serodiagnosis of chronic human fascioliasis.

An indirect enzyme-linked immunosorbent assay (ELISA) was developed and evaluated for its diagnostic ability to detect human IgG antibodies against Fasciola hepatica saposin-like protein-2. The assay was compared with an indirect ELISA with excretory-secretory products (FhES) from adult F. hepatica.