Double-filtered leukoreduction as a method for risk reduction of transfusion-associated graft-versus-host disease

Background Transfusion-associated graft-versus-host disease (TA-GvHD) is caused by leukocytes, specifically T cells within a transfused blood product. Currently, the prevention of transfusion-associated graft-versus-host disease is performed by irradiation of blood products. With a sufficient reduction of leukocytes, the risk for TA-GvHD can be decreased. With consistent advances in current state-of-the-art blood filters, we herein propose that double filtration can sufficiently reduce leukocytes to reduce the risk for TA-GvHD. Materials Thirty RBC concentrates were filtered with leukocyte filters, followed by storage at 1–6 o C for 72 hours, and then a second filtration was performed. Residual leukocytes in the double-filtered RBC units (n = 30) were assessed with flow cytometric methods, and an additional assay with isolated peripheral blood mononuclear cells (PBMCs) (n = 6) was done by both flow cytometric methods and an automated hematology analyzer. Quality of the RBCs after filtration was evaluated by hematological and biochemical tests. In vitro T cell expansion was performed using anti-CD3/CD28-coated Dynabeads or anti-CD3 (OKT3). In vivo experiment for GvHD was performed by using NOD.Cg-Prkdc scid Il2rg tm1Wjl /SzJ (NSG) mice. Results Double-filtered blood products showed residual leukocyte levels below detection limits, which calculated to be below 1200–2500 cells per blood unit. In vitro expansion rate of T cells showed that 6x10 3 and 1x10 3 cell-seeded specimens showed 60.8±10.6 fold and 10.2±9.7-fold expansion, respectively. Cell expansion was not sufficiently observed in wells planted with 1x10 2 or 10 cells. In vivo experiments showed that mice injected with 1x10 5 or more cells cause fatal GvHD. GvHD induced inflammation was observed in mice injected with 1x10 4 or more cells. No evidence of GvHD was found in mice injected with 10 3 cells. Conclusions Our study suggests that additional removal of contaminating lymphocytes by a second leukodepletion step may further reduce the risk for TA-GvHD.