Characterized Helminth Defense Molecule from liver fluke Opisthorchis viverrini, investigating its structural features and dual immunomodulatory roles. The protein modulates host immune responses to facilitate chronic infection, interacting with the bile duct bacterial microbiome and cathelicidin-mediated defense.
Abstract
The liver fluke Opisthorchis viverrini causes chronic infections in humans and animals. This helminth is known to coexist with the bacterial microbiome in the host's bile duct, and their interaction potentially impacts the helminth's pathogenicity. While most infected individuals remain asymptomatic, the mechanism of immune modulation remains unclear. The immunomodulatory protein Helminth Defense Molecule (HDM) has been characterized in other liver fluke species; however, its functional role exhibits distinct variations. This study aims to uncover the structural properties and biological function of HDM from O.viverrini (OvHDM). Using bioinformatics tools, we predicted the protein structure, analyzed its physicochemical properties, and potential molecular interactions. Furthermore, the structural and molecular interactions were experimentally confirmed by Circular Dichroism (CD) Spectroscopy. For functional analysis, RAW 264.7 macrophage cells were treated with Escherichia coli LPS pre-incubated with synthetic OvHDM peptides, followed by TNF and IL-6 levels quantification using ELISA. The predicted structure of OvHDM predominantly consists of α-helices configuration, resembling the antimicrobial peptide LL-37. It exhibits a high distribution of positive amino acids, strong stability under physiological conditions, and potential functional sites in both the N- and C-terminal domains. OvHDM demonstrated stronger interactions with LPS compared to the FhHDM-1 P2 as a control. The CD spectroscopy showed secondary structure dynamics of HDMs synthetic peptides upon LPS addition. Finally, synthetic OvHDM peptides were shown to both reduced LPS-induced cytokine production in vitro, and independently induced IL-6 production. These findings highlight the dual immunomodulatory activity of OvHDM, suggesting its role in fine-tuning the host immune environment during infection.