Fascioliasis is primarily caused by two flukes: Fasciola hepatica and Fasciola gigantica. The parasite infect domestic and wild animals, leading to significant economic and productive losses in the livestock industry, estimated at approximately US $3 billion annually (Spithill et al., 2012). This infectious disease has been listed by the World Health Organization (WHO) as one of the Neglected Tropical Diseases (NTDs), with around 17 million people infected and 0.18 billion at risk of fascioliasis (Cwiklinski et al., 2016). The pathogenesis of fascioliasis begins with the accidental ingestion of encysted metacercariae by definitive hosts (humans and grazing animals) (Hu et al., 2022). Upon reaching the small intestine, metacercariae excyst into newly excysted juveniles (NEJs), which then penetrate the duodenal wall. These NEJs migrate through parenchyma and ultimately establish themselves in the liver and gall bladder. Once settled, they engage in continued feeding, burrowing, and growth within the liver parenchyma. (Soliman, 2008; Young et al., 2011). The management of fascioliasis largely relies on anthelmintics practices, with triclabendazole being a particularly common choice. However, instances of resistance and/or treatment failures have been documented in various parts of the world (Fairweather, 2011; Hodgkinson et al., 2013). To address the challenges in controlling fascioliasis, which include issues like inaccurate diagnosis, drug resistance, and health risks to both humans and animals, there is an urgent need for innovative diagnostic tools and novel immunotherapeutic control strategies.

The immune response of hosts during natural infection by F. hepatica in ruminants has been well-characterized. Similar findings have been reported on F. gigantica infection (Kumar et al., 2013; Molina and Skerratt, 2005). Throughout its dissemination, localization, and development, F. gigantica encounters various host cell-types, eliciting both humoral and cellular immune responses, and this contributes to the development of an effective immune-modulatory capacity, enabling the parasite's long-term survival within its hosts. (Robinson et al., 2013). Similar to other helminth parasites, Fasciola spp., employ various immunological strategies through released of numerous antigens/molecules during parasite-host interface (Dalton et al., 2003; Zhu et al., 2019). The PBMCs comprise of diverse populations of immune cells, including T-lymphocytes, B-lymphocytes, NK cells, and monocytes), performing crucial part in immune responses. T-helper-1 and -2 (Th1 and Th2) represent major subsets of CD4+ T cells, categorized based on their secretion patterns and cytokines functions. Th1-cells produce interferon gamma (IFN-γ) and tumor necrosis factor alpha (TNF-α) cytokines, in addition to stimulation and activation of macrophages, and mediate delayed-type hypersensitivity and inflammatory responses. On the other hand, Th2-cells secrete interleukin IL-4 and IL-10 cytokines, triggering immediate-type hypersensitivity through IgE, eosinophils, and mast cells. Cytokines from individual T-cell subtypes mutually suppress the differentiation and effector functions of the opposing subset, resulting in the polarization of the immune response into Th1 or Th2 (Gironènes et al., 2007). Previous studies suggested that, at early stages of fascioliasis, Th cytokines induce a polarized Th2 response (Tliba et al., 2002).

Legumains and asparagine endopeptidases (AEPs) are a members of the C_13 peptidase family of proteins, which utilize the thiol group of a cysteine residue to hydrolyze the asparaginyl bond (Chen et al., 1997). Legumains have been identified from various vertebrates (humans, cattle, pigs, and mice) (Chen et al., 1997; DANDO et al., 1999; Yamane et al., 2002), and invertebrate species, including F. hepatica (Tkalcevic et al., 1995), F. gigantica (Adisakwattana et al., 2007), Schistosoma mansoni (Dalton et al., 1995), Trichomonas vaginalis (Rendón-Gandarilla et al., 2013), Haemonchus contortus (Oliver et al., 2006), Ticks (Alim et al., 2007) and Clonorchis sinensis (Ju et al., 2009). Some studies demonstrated that Legumains are associated with a range of biological activities, such as post-translational modification, antigen presentation to class II of the major histocompatibility complex (MHC), mobilization of storage proteins, digestion of host blood, and other beneficial physio-chemical functions of parasites (Alim et al., 2007; Dalton et al., 1995). Furthermore, the roles of Legumain in apoptosis and the regulation of the cell cycle have also been documented (Gawenda et al., 2007; Zornig et al., 2002). Previous transcriptional data has shown the extensive involvement of Legumains, along with cathepsin B and cathepsin L proteases, in the immune system. This encompasses roles in antigen presentation and processing, activation and migration of immune cells, as well as manipulation of the host immune response and evasion by the parasite (Zhang and Cwiklinski, 2019). The genomic characterization of Fasciola spp., has identified a number of cathepsin L and cathepsin B peptidase genes (Cwiklinski et al., 2019), and their activation has been shown to be regulated by Legumain genes encoding asparaginyl endopeptidase (Cwiklinski and Dalton, 2022). This suggests that Legumains could be a promising target for novel drug and vaccine-based therapeutic interventions. (Dalton et al., 2009).

Previous study has suggested a highly expression of Legumain in colon, prostate, and breast cancer (Murthy et al., 2005). In addition, Legumain has also been found in both intracellular and extracellular in tumor-associated endothelial cells as well as on macrophage cell surfaces (Liu et al., 2003). In a prior study, the proteomic profile of F. gigantica ESPs detected LGMN-1, an interacting protein, predicted to be involve in the host immune response during F. gigantica infection (Huang et al., 2019). However, the precise mechanism of how the Fg-LGMN-1 protein interacts with host immune cells remains unknown. The present study is dedicated to characterizing a recombinant protein from F. gigantica (Fg-LGMN-1), and exploring its immunomodulatory characteristics in vitro with the goat monocytes. This research aims to establish the foundation for understanding the cellular and biological mechanisms that regulate the immune responses during host-parasite interplay.