Updated project metadata. Dietary fats have been shown to affect gut microbiota composition and aging gene transcription of middle-aged rats at a normal dose, but little is known about such an effect on gut barrier. In colon, the main component of mucus layer is Muc2, produced by the goblet cells. This study investigated the changes in Muc2 expression, goblet cells proliferation, TLRs and inflammatory cytokines in the colon of middle-aged rats. Proteome technology was applied to explore the possible molecular mechanisms. The results indicated that intake of fish oil at a normal dose downregulated colonic Muc2 expression, and this negative effect of fish oil probably involved the suppression of mucin glycosylation process.