LB042 - LPS INDUCES TLR4 AND CYTOKINES IN A 3D PORCINE ENTEROID SYSTEM TO DRIVE MULTISYSTEM INJURY

LB042

LPS INDUCES TLR4 AND CYTOKINES IN A 3D PORCINE ENTEROID SYSTEM TO DRIVE MULTISYSTEM INJURY

C. Manithody1, A. Bagwe1, A. Jain1,*, K. Kurashima1, M. Swiderska-Syn1, U. Ezekiel1, S. Mehta1

1Saint Louis University, St. Louis, United States

 

Rationale: The intestinal epithelium plays a critical role in gut homeostasis, serving as a physical barrier and modulating immune responses to microbial stimuli. This homeostasis is disrupted during Total Parenteral Nutrition (TPN), which bypasses enteral nutrient delivery. Lipopolysaccharide (LPS), a key component of Gram-negative bacterial membranes, is a potent activator of pro-inflammatory pathways. We hypothesized that porcine-derived 3D intestinal enteroids could serve as a robust model to study epithelial immune activation and barrier function in response to LPS exposure.

Methods: Enteroids were generated using Matrigel-based 3D cultures from neonatal Yorkshire pig small intestine and expanded over 7 days. Enteroids were treated with LPS at increasing concentrations (1, 2, and 5 µg/mL). Gene and protein markers for epithelial integrity and immune activation were measured. Serum cytokines were also measured in animals on enteral nutrition (EN control) or TPN as historical references.

Results: Exposure to 5 µg/mL LPS resulted in a marked reduction in Zonula Occludens-1 (ZO1), indicating compromised epithelial barrier integrity. TLR4 expression was significantly increased in both 2 and 5 µg/mL LPS groups, with maximal expression at 2 µg/mL. IL-8 and IL-10 were similarly elevated in these higher-dose groups. Serum cytokine profiling revealed elevated IFN-γ (p=0.009), IL-8 (p=0.011), and LPS (p<0.0001) in TPN versus EN-fed animals. No significant differences in serum IL-10 were observed.

Conclusion: Our porcine enteroid model effectively recapitulates intestinal epithelial responses to LPS, including innate immune activation (via TLR4), cytokine production, and barrier disruption. These data support the utility of this model in studying epithelial-driven mechanisms of inflammation relevant to TPN-associated intestinal dysfunction.  

Disclosure of Interest: None declared