Tributyrin Supplementation Protects Immune Responses and Vasculature and Reduces Oxidative Stress in the Proximal Colon of Mice Exposed to Chronic-Binge Ethanol Feeding
Effects of tributyrin on presence of immune cells in proximal colon following chronic-binge ethanol exposure. Mice were fed an ethanol-containing (5% v/v) liquid diet or pair-fed a diet with maltose-dextrin isocalorically substituted for ethanol for 10 days. Diets were supplemented with glycerol or tributyrin (5 mM). Mice were then treated with a single 5 g/kg gavage of ethanol the next day containing glycerol or tributyrin (2.5 mM). At 9 h post-gavage, the proximal colon was collected and used to prepare RNA or embedded in optimal cutting temperature compound (OCT) for histology. (a) CD45 (green), (b) CD68 (green), (f) C3b/iC3b/C3c, and (g) NIMP-R14 were visualized by immunohistochemistry in sections of proximal colon frozen in OCT. Images were acquired using a 10x or 20x objective. A selected area was cropped and enlarged. (c–e) Expression of CD68, Ly6c, and ELANE mRNA was detected in the proximal colon using quantitative real-time reverse transcription polymerase chain reaction. (f) In addition to pair-fed and ethanol-treated mice, proximal colon frozen in OCT from age- and gender-matched C57BL/6 and C3−/− mice on a C57BL/6 background were stained for expression of C3b/iC3b/C3c. C3b-positive staining was visualized in wild-type mice similarly to that of pair-fed mice, and, as expected, C3−/− were negative for positive C3b staining. Images are representative of at least replicate images captured per mouse in four to six mice per treatment group. Data are error of the mean (SEM). Values with different alphabetical superscripts were significantly different from each other; and .