Fh15 Modulates Human Neutrophil Activation and Reduces NETosis 2335416

Abstract

Abstract Introduction Neutrophils are essential innate immune effectors, however, excessive activation can exacerbate inflammatory disease. A key pathogenic mechanism is neutrophil extracellular traps (NETs) formation, in which neutrophils release chromatin structures with antimicrobial proteins. While NETs can aid antimicrobial defense, dysregulated NETosis amplifies inflammation, promotes endothelial/epithelial damage, and has been linked to severe outcomes in conditions such as sepsis, autoimmune disease, and acute respiratory distress syndrome. Fh15, a recombinant Fasciola hepatica fatty acid-binding protein, is a parasite-derived immunomodulator with demonstrated anti-inflammatory activity in septic shock and ulcerative colitis. Fh15 has been shown to reduce neutrophils in inflamed colonic tissue and to decrease systemic myeloperoxidase, supporting an impact on neutrophil-associated inflammation. Here, we investigate whether Fh15 directly limits NETs and modulates neutrophil activation under pro-inflammatory stimulation. Methods Primary human neutrophils were stimulated with lipopolysaccharide (LPS) and treated with Fh15 at 5, 10, or 25 µg/mL. NETs formation was quantified using 6-hour live-cell imaging. Neutrophil activation was assessed by flow cytometry based on surface expression of CD11b, CD62L, and CD16. Results Fh15 reduced NETosis at 10 (p < 0.001) and 25 (p < 0.0001) µg/mL. Flow cytometry analysis showed that LPS reduced CD11b+CD16+ neutrophils (p < 0.0001), consistent with inflammation-associated shedding or downregulation of surface CD16. In contrast, Fh15 treatment increased the percentage of CD11b+CD16+ cells compared with LPS alone (p < 0.001). Conclusion Together, the data suggests that Fh15 helps preserve surface CD16 under pro-inflammatory stimulation. In conclusion, Fh15 dampens LPS-driven neutrophil effector responses, particularly NETs formation, and may modulate activation-associated phenotypes, supporting its potential as an immunomodulatory strategy to limit neutrophil-mediated tissue damage. Funding Source Funding was provided by the RCMI-CRRGMG (U54MD007579) and PR-INBRE DRPP (2P20GM103475-02). Topic Categories Innate Immune Responses and Host Defense: Cellular Mechanisms (INC)