Plant-derived betaine enhances barrier integrity and immune responses in canine intestinal epithelial cells and macrophages under endotoxin challenge

Interest in the utilization of novel plant-based extracts for supporting gut health and nutrition in companion animals is growing. The effect of betaine on canine intestinal barrier integrity and immune-related gene and protein expression by epithelial and macrophage-like cells was evaluated under conditions of inflammatory challenge from lipopolysaccharide (LPS). We evaluated: (1) the effect of betaine pre-treatment (0 to 1,000 µg/mL for 24 h) on the viability of canine intestinal epithelial MCA-B1 and macrophage-like DH82 cells; (2) epithelial barrier function of LPS-challenged MCA-B1 cells pre-treated with betaine, by measurement of 4-kDa fluorescein isothiocyanate (FITC-dextran [FD4] flux); (3) gene and protein expression of tight junction proteins and adhesion molecules using quantitative reverse transcription polymerase chain reaction (qRT-PCR) and proteomics; (4) anti- and pro-inflammatory and regulatory gene expression in LPS-challenged, betaine pre-treated, MCA-B1, DH82 and peripheral blood mononuclear cells and; (5) the phagocytic activity of DH82 cells pre-treated with betaine. No cytotoxicity of betaine against MCA-B1 or DH82 cells were observed. Betaine pre-treatment of MCA-B1 reduced FD4 permeability compared with untreated cells (P < 0.05), accompanied by increased (mRNA) expression of claudin-1 (P < 0.001) and E-cadherin (P < 0.05) and increased protein expression of junctional adhesion molecules (P < 0.05), vinculin (P < 0.05) and aquaporin-3 (AQP3; P < 0.01). Betaine pre-treatment of LPS-challenged cells also increased the expression of regulatory cytokines TGF-β1 (P < 0.01), TGF-β2 (P < 0.01), and IL-33 (P < 0.05) in MCA-B1 cells and reduced the expression of IL-6 (P < 0.05) and MyD88 (P < 0.01) in DH82 cells compared with untreated cells. In addition, betaine enhanced the phagocytic activity of DH82 cells (P < 0.01) compared with untreated cells. Confirmatory studies in vivo are needed but these results suggest that plant-derived betaine has potential as a nutritional supplement for companion animals to support gut health and immunity.