Diabetic nephropathy (DN) is a common complication in patients with diabetes, and most of them need renal replacement therapy such as haemodialysis (HD). These patients have a high tendency to develop infections and exhibit anomalies in the immune system. The objective of this study was to assess the expression of activation-related markers on T cells, as well as to quantify inflammatory cytokines, before and after a single HD session in DN patients. The study involved DN patients under HD treatment who signed an informed consent form. Blood samples before and after one HD session were collected, to analyse the expression of CD25, CD69 and CD71 in T cells. We also quantified IL-12p70, IL-8, IL-10, IL-1β, TNF-α and IL-6 in serum samples using the cytometric bead array technique. After the HD session, there was an increase in the CD4/CD8 ratio due to significant alterations in both subsets. The relative percentage of CD25+ cells and CD8+ CD25+ increased significantly after the HD session, while the relative percentage of CD69 T cells decreased. There was a significant decrease in the CD25 mean fluorescence intensity values for CD4+ T, as well as in the case of CD71 in T cells after the HD session. Regarding cytokine synthesis, we found a significant increase in IL-10 and IL-6 and a decrease in IL-8 after HD session. This study showed that a HD session in DN patients affects the T cell activation status in the two major subpopulations and differentially modulates the production of inflammatory cytokines.