In endodontic infections, inflammatory mediators such as cytokines are released, recruited and retained until the infection is eradicated. Root canal therapy is performed to prevent the spread of infection. The aim of this study was to investigate the effects of root canal debridement (cleaning and shaping) on periapical inflammation by measuring the levels of inflammatory cytokines, Interleukin-8 (IL-8) and Interleukin-10 (IL-10). The study includes twenty patients with pulp necrosis and asymptomatic apical periodontitis. Periradicular sample was collected using paper points before and after root canal debridement. Cytokine levels were determined by Sandwich Enzyme-Linked Immunosorbent Assay (ELISA). Data were analysed using paired t-test (PASW Statistics 18) (P = 0.05). All samples showed the presence of IL-8 and IL-10 prior to root canal debridement. Significantly reduced levels (P < 0.05) of IL-8 and IL-10 were detected after root canal debridement. In conclusion, root canal debridement significantly decreased the levels of the tested pro- and anti-inflammatory cytokine in the periradicular interstitial fluid.