Background: Despite several attempts, the etiopathogenesis of anorexia nervosa (AN) is still unknown. However, the activation of the immune response in neuropsychiatric diseases, including AN, is increasingly evident. We aimed to explore immune response parameters in patients with AN and identify the link between the presence of specific autoantibodies for hypothalamic antigens and the inflammatory response. The relationship between inflammatory markers and the duration of the disease has been also investigated. Methods: Twenty-two patients with AN were included, and none were under psychopharmacological treatment or suffering from autoimmune conditions. Serum concentrations of interleukin (IL)-6, IL-1β, tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-β, and IL-21 were determined by ELISA kits. In addition, autoantibodies against hypothalamic antigens are quantitatively evaluated. Results: IL-6, IL-1 β, TNF-α, and TGF-β are significantly increased in patients with AN. A positive correlation with body mass index and with the amount of autoantibody specific for hypothalamic antigens exists. Notably, a progressive reduction of cytokines correlates with the progression of AN. In addition, IL-21 is increased in the blood of patients with AN and negatively correlates with autoantibody concentrations. Conclusions: This study shows that the increased pro-inflammatory phenotype in patients affected by AN correlates with the concentration of autoantibody specific for hypothalamic antigens. Of interest, the pro-inflammatory state seems to be reduced with duration of AN. In addition, IL-21 could work as a stimulant of the immune response, thus possibly increasing the autoreactivity.