A healthy diet rich in fish, fruit, and vegetables, moderate in alcoholic beverages, and low in dairy products has been associated with lower circulating concentrations of biomarkers of endothelial dysfunction (ED) and low-grade inflammation (LGI). It is, however, unknown how consumption of these food groups affects ED and/or LGI over time. We measured diet by the computer-assisted crosscheck dietary history method at 36 ± 0.63 y of age (n = 301, women = 161). At 36 and 42 y of age, we measured von Willebrand factor, soluble intercellular adhesion molecule 1 (sICAM-1), soluble endothelial selectin, soluble vascular cell adhesion molecule 1 and soluble thrombomodulin (circulating biomarkers of ED); and C-reactive protein, serum amyloid A, IL-6, IL-8, TNFα, and sICAM-1 (circulating biomarkers of LGI). We investigated the associations between food groups and changes in combined biomarker Z-scores of ED and LGI [higher scores associated with greater risk of (incident) cardiovascular disease]. After adjustment for sex, energy intake, BMI, physical activity, alcohol consumption, smoking behavior, and other food groups, consumption of fish (per 100 g/wk), but none of the other food groups, was inversely associated with changes in ED [β (95%CI) = -0.06 (-0.10; -0.02); P = 0.003] and LGI [-0.05 (-0.09; -0.003); P = 0.036]. Additionally, EPA+DHA intake was inversely associated with changes in ED [β (95%CI) = -0.13 (-0.19; -0.07); P ≤ 0.001] and LGI [-0.09 (-0.16; -0.02); P = 0.013] and explained 83 and 40% of the association between fish and changes in ED and LGI. In conclusion, fish consumption, but not fruit, vegetable, alcoholic beverage, or dairy product consumption, was associated with decreased ED and LGI in healthy adults.