Immune regulatory effect of tacrolimus in patients with severe aplastic anemia and in murine model

Abstract

Objective: To investigate the immunomodulatory effect of tacrolimus in severe aplastic anemia (SAA). Methods: Patients diagnosed with SAA at the Department of Hematology, General Hospital of Tianjin Medical University from June 2015 to January 2018 were enrolled. CD8+T cells were sorted by immunomagnetic separation from peripheral blood of SAA patients. MTT method was used to detect the proliferation of CD8+T cells. The SAA mouse model was established by total body irradiation (TBI) and donor lymphocyte infusion (DLI). There were 10 normal controls without pretreatment, 10 rats in TBI group, 15 rats received TBI and DLI. The expression of perforin and granzyme in CD8+T cells and the ratio of CD4+/CD8+cells in peripheral circulation were measured by flow cytometry. The level of interferon-γ (IFN-γ) in medium supernatant of cultured CD8+T cells was tested with enzyme-linked immunosorbent assay (ELISA). SAA mouse model was established to study the recovery of hemogram and survival time after treatment. Results: A total of 16 SAA patients were enrolled, and there were 10 males and 6 females, with a median age of 35 (22-49) years. Tacrolimus inhibited the proliferation of CD8+T cells when IL-2 concentration was 20.0,200.0 and 2 000.0 U/ml (P<0.05). The expression of perforin in CD8+T cells of SAA patients treated with tacrolimus was significantly lower than that in blank control group and IL-2 group [(2.25±0.76)%, (6.70±0.82)% vs (9.10±1.90)%,all P<0.05]. The level of IFN-γ in CD8+T cells group after applying tacrolimus was significantly lower than that in the blank control group (P<0.05). After 10 days of administration, the peripheral blood hemoglobin (Hb), white blood cell (WBC) and platelet (PLT) counts of SAA mice in tacrolimus group were all higher than those in SAA group (all P<0.05). The expression of perforin in CD8+T cells in tacrolimus group was significantly lower than that in SAA group [(18. 39±6.65) vs (29. 99±9.83),P<0.05]. The median survival time of SAA group was 18.6 days, and the 90 day survival rate was 0. The median survival time of tacrolimus group was 44.6 days, and the 90 day survival rate was 80%. The survival time of SAA mice in tacrolimus group was significantly longer than that in SAA group (P<0.05). Conclusion: The immunomodulatory effect of tacrolimus in SAA is similar to CsA. It has an immunosupressive effect on CD8+T lymphocyte.