Use of probiotics to modulate antimicrobial peptide signaling in the neonatal innate immune response to influenza virus

The National Institute of Allergy and Infectious Diseases awarded Drexel University $554,786 on September 17, 2025, under the Allergy and Infectious Diseases Research program (CFDA 93.855) to fund basic research on mechanisms by which the probiotic Lactobacillus rhamnosus (LGG) modulates antimicrobial peptide signaling and innate immune response to influenza virus in neonates. The project, titled "Use of Probiotics to Modulate Antimicrobial Peptide Signaling in the Neonatal Innate Immune Response to Influenza Virus," addresses the high mortality rate from influenza in infants under six months of age, a population currently ineligible for influenza vaccination. Research activities employ a neonatal murine infection model to determine how LGG-derived signals impact pulmonary epithelial cells and innate immune cells, including alveolar macrophages and neutrophils. Prior work demonstrated that intratracheal administration of LGG prior to infection improved neonatal survival; this award funds investigation of the cellular and molecular mechanisms underlying that protective effect. Work is performed in Philadelphia, Pennsylvania. The period of performance runs through August 31, 2027.

Abstract

The mortality rate from influenza viral (IV) infection is highest in infants less than age six months, currently an age group not eligible for the available influenza vaccine, but the mechanisms for this clinical observation are not well understood. Vulnerability to respiratory viruses during infancy is likely manifested by an immature lung and immune system. To understand age-specific differences in hematopoietic and epithelial cell function at the air-blood interface in the developing lung, an age-appropriate pre-clinical neonatal murine IV infection model is employed. Previously, we demonstrated that murine neonates are exceptionally susceptible to IV infection. Differences in the dominant airway microbial communities in these first few months of life have been linked to susceptibility to respiratory infections. Commensal-derived signals establish an activation threshold of the innate immune system required for optimal antiviral immunity. Therefore, modulating early airway microbial communities presents a potential therapeutic strategy to prevent or ameliorate respiratory tract infections. Recently, we demonstrated that intratracheal administration of lactobacillus rhamnosus (LGG), a probiotic, prior to IV infection improved neonatal survival. However, the specific mechanisms by which LGG acts, the cells on which IT exerts the most effect and the dominant pattern recognition receptors IT modulates remains to be determined. The goal of this project is to determine mechanisms by which these LGG-derived signals impact the function of 1. Pulmonary epithelial cells, the target of IV infection, and 2. Innate immune cells, such as alveolar macrophages (AMS), sentinel tissue-resident immune cells which drive immune cell recruitment, and neutrophils, which are pathologic in the developing lung during respiratory viral infection. We have identified an antimicrobial peptide as a potential signaling mediator between infected epithelial cells and AMS. Our recently published work demonstrates that murine neonates deficient in catheliciden-related antimicrobial peptide (cramp) are protected during IV infection. Importantly, LGG pretreatment decreases cramp production. Here, we seek to investigate the pathogenic role of cramp as a signaling mediator to orchestrate the initial response to IV. The overarching hypothesis of this proposal is that in IV-infected neonates, LGG reduces cramp production by epithelial cells, which diminishes cramp-mediated alveolar macrophage (am) activation through toll-like receptor 2 (TLR2) and subsequent neutrophil chemoattractant production. Neutrophil infiltration is a potent driver of mortality. To test our hypothesis, we will: (1) define IV tropism and the primary cell source of cramp in LGG treated and untreated IV-infected murine neonates and adults using single cell sequencing; (2) evaluate the age- specific role of LGG and cramp-dependent TLR2 signaling in key innate immune cell production of neutrophil chemoattractants; and (3) determine if cramp acts directly on neutrophils to increase oxidative stress. Our studies in an innovative neonatal pre-clinical animal IV model will bring forth new understanding of infant mucosal immunity to develop targeted therapeutics for the infant population.