90 Methods to measure gut barrier function in ruminants.

Abstract Gut tissues are principally responsible for nutrient absorption, barrier function, and immune sensing and are thus key contributing factors to the productivity of cattle. Stressors such as weaning, transportation, changes in diet, and environmental factors can lead to microbial dysbiosis and inflammation, resulting in compromised gut barrier integrity and increased permeability. The intricate interplay among commensal microbes, pathogens, and host defenses significantly impacts barrier function, with distinct differences observed between the rumen and the post-rumen due to their unique anatomical and physiological characteristics. While ruminal health has received much attention, it is increasingly evident that the post-ruminal gastrointestinal tract (GIT) with its robust immune anatomy substantially contributes to the pathophysiology of microbial dysbiosis. Pathological factors, such as inflammation increase enterocyte cell loss leading to altered villus/crypt morphology. For this reason, villus height and crypt depth measures are routinely used to gauge GIT inflammation and/or effectiveness of nutritional regimens. Molecules infiltrate the ruminal and post-ruminal epithelium by two different routes: 1) paracellular – between two neighboring cells – and 2) transcellular – across cells. The passage of substances through the paracellular route is minimal and primarily limited to solutes, water, and ions. Paracellular permeability increases during inflammation; however, the majority of large antigens, including microbial products, traverse the GIT epithelium through transcellular endocytosis mechanisms used by the intestinal immune system. M-cells sample the digestive contents for antigens and present them to the underlying lymphoid tissue for phagocytosis. This process appears to be overwhelmed when stress and tissue damage are significant. Paracellular permeability is typically assessed through experimental methods involving the oral administration of non- or poorly metabolized, water-soluble marker compounds, such as Cr- or Co-EDTA, with measurements often taken from urine samples because Cr and Co are more concentrated in urine than blood. Diet or intake alterations that impact rate of passage may change the timing of peak urine Co and Cr concentrations and necessitate changes in delivery methods or data interpretation. Additionally, increased appearance of Co and Cr in urine may not necessarily be indicative of GIT inflammation but rather an improvement in absorptive capacity of the GIT. Acute phase proteins can serve as reliable indicators of infection in cattle, yet their lack of disease specificity poses challenges when pinpointing a particular ailment. Previous research has established that the primary source of microbial lipopolysaccharide (LPS) entering into the bloodstream is from the gut and that LPS binding protein (LBP) is positively correlated with reductions in jejunum villus height to crypt depth ratio. The complex effects of diet and stress on gut integrity necessitate diverse analytical methods. The physiology and rationale behind permeability assays, morphological measurements, and biomarker analyses for assessing gut barrier function in ruminants will be discussed.