The use of a whole gut sac perfusion model to study the ex vivo inflammatory response of rainbow trout (Oncorhynchus mykiss) to lipopolysaccharide

• 2hr intestine static “tie-bag” perfusions withstood leaking • LPS treated middle intestine sections significantly upregulated IFN-I and TNF-α • LPS treated distal intestine sections significantly upregulated TNF-α and IL-1β • Both intestinal sections demonstrated minimal epithelial sloughing Ex vivo studies provide a valuable opportunity to study organ-level effects of a compound in an isolated system. As all vertebrate cells can mount an innate immune response, it is important to study the cumulative immune response of multiple cell types that remain in their natural conformation, proportions, and polarity; something that in vitro studies cannot fully provide. This study proposes a simplified static “tie-bag” perfusion model to study the short-term immunological stimulation of rainbow trout middle and distal intestine by Fluorescein isothiocyanate (FITC) tagged lipopolysaccharide (LPS), a component of the gram-negative bacteria cell wall. Middle intestinal segments perfused with 200µg/ml FITC-LPS demonstrated significant increases in the expression of type-I interferon 1 (IFN-1) and tumor necrosis factor alpha (TNF-α) transcripts, while distal intestinal perfusions demonstrated significant increases in TNF-α and interleukin 1β (IL-1β) transcript levels. While FITC-LPS distribution could not be visualized due to overlap with tissue autofluorescence, tissues appeared to only experience minor epithelial sloughing. Overall, this method can be used to evaluate organ-level immune stimulation of antigens while maintaining tissue composition that permits histological examination.