Elucidating FH15-mediated modulation of TLR4/NF-KB in neutrophils to suppressinflammation in sepsis and ulcerative colitis - the increasing application of helminth antigen-derived therapy to prevent…

The National Institute of Allergy and Infectious Diseases awarded the University of Puerto Rico Medical Sciences Campus $149,000 on September 1, 2026, under the Allergy and Infectious Diseases Research program (CFDA 93.855) to investigate FH15-mediated modulation of TLR4/NF-κB signaling in neutrophils as a therapeutic approach for sepsis and ulcerative colitis.

The research tests whether FH15, a recombinant molecule derived from the Fasciola hepatica fatty acid binding protein family, suppresses inflammatory pathways in animal models and human neutrophils. Prior work funded under a companion SC1 grant demonstrated that a single intraperitoneal dose of FH15 administered to mice one hour before or after lethal lipopolysaccharide or live E. coli infusion significantly suppresses cytokine storm and inflammatory markers in sepsis models, and that three spaced doses improve disease activity in dextran sulfate sodium-induced ulcerative colitis models. The previous award generated one patent, approximately five publications, and supported three PhD and one master's degree graduates.

The central hypothesis posits that FH15 modulates TLR4/NF-κB signaling to reduce extracellular DNA concentrations and suppress netosis, a form of neutrophil cell death associated with inflammatory disease. The award supports this continuation research through August 31, 2030, with place of performance in Puerto Rico. This is a project grant, the standard assistance type for individual investigator-directed research under the program.

Elucidating FH15-mediated modulation of TLR4/NF-KB in neutrophils to suppressinflammation in sepsis and ulcerative colitis - the increasing application of helminth antigen-derived therapy to prevent inflammatory diseases represents a promising and innovative approach. Over the past decade, our research group has concentrated on FH15, a recombinant molecule from the fasciola hepatica fatty acid binding protein family, which exhibits potent antiinflammatory effects in animal models of sepsis and ulcerative colitis. We have demonstrated that a single intraperitoneal (I.P.) dose of FH15 administered to animals one hour before or after the administration of a lethal dose of lipopolysaccharide (LPS) or live E. Coli infusion significantly suppresses the cytokine storm and other inflammatory markers. Additionally, we found that a single I.P. Dose of FH15 given three times, two days apart, significantly improves the disease activity index (DAI) in a dextran sulfate sodium (DSS)-induced ulcerative colitis mouse model. These therapeutic effects correlated with reduced levels of intestinal inflammatory markers, diminished neutrophil infiltration, and lower levels of myeloperoxidase-a protein essential for neutrophil extracellular trap (net) formation. Furthermore, we demonstrated that FH15 can inhibit netosis in human neutrophils and acts as a TLR4 antagonist, a receptor involved in the immune response against sepsis and ulcerative colitis, as well as in neutrophil activation and net release. This application stems from a previous SC1 grant (1SC1AI155439-01), which resulted in one patent, approximately five publications, and the graduation of three PHD and one MS students. Our central hypothesis posits that FH15 modulates the TLR4/NF-ÊB signaling pathway to suppress inflammation in animal models of sepsis and ulcerative colitis, which is associated with decreased concentrations of free extracellular DNA (EDNA) and reduced netosis. In the aim-1 will demonstrate the relationship between the suppression of inflammatory markers induced by FH15 in the sepsis and ulcerative colitis models and the reduction of EDNA and nets in vivo. We will isolate free EDNA from plasma in sepsis and colitis mice treated with FH15 and characterize the specific protein-DNA components of this EDNA through immunofluorescence staining and confocal microscopy. The results will be correlated with levels of inflammatory markers for colitis and sepsis. In the aim-2 will identify the mechanisms underlying the inhibitory effects of FH15 on netosis. We will isolate neutrophils from mouse bone marrow and analyze activation markers such as CD11B, CD66B, CD62L, CD64, and fcri using flow cytometry. Additionally, we will assess the expression of TLR4, CD14, and proteins involved in the NF-ÊB pathway using qpcr and western blotting. Proximity ligation assays will determine whether FH15 directly interacts with CD14 on these cells. We will also employ proteomics and single-cell RNA sequencing to investigate changes in protein and gene expression in neutrophils and identify molecules involved in pro-inflammatory pathways that may be regulated by FH15. The outcome of this study will provide a more comprehensive understanding of how F. Hepatica molecules interact with the immune system and lay the groundwork for the development of a novel class of anti-inflammatory drugs.