Study Of Anti-mouse Hepatocellular Carcinoma On TIL Stimula By DC In Vitro

Objective: Dendritic cells (DC) were obtained from mouse marrow and stimulated with lysates mouse hepatocellular carcinoma cells (H22) antigen in vitro. To investigate the killing activity of tumor-infiltrating lymphocytes (TIL) stimulated by dendritic cells on H22 in vitro . Methods: Bone marrow cell suspensions were prepared with aseptic technique, and then eiythrocytes, T and B lymphocytes, granulocytes and monocyte macrophages were removed separately by treating successively with red cell lysing buffer, rat anti-mouse CD4 ,anti- mouse CD8a, anti-mouse CD45R monoclonal antibodies and rabbit complement buffer, and by semi-adhesion of DC to culture plates. The cells were cultured with granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin-4 (IL-4). A large number of DC were obtained and stimulated with lysates H22 antigen in vitro. In vitro cytoxicity assays experiment, the effector cells of experiment group were TIL stimulated by DC(A group), the effector cells of control groups were TIL not stimulated by DC (B group), splenocytes lymphocytes stimulated by DC(C group), and splenocytes not stimulated by DC(D group). Then their killing activities on H22 were assayed in vitro. Result: (1) (2-3) 106 DC were obtained from bone marrow of onemouse, and the purity of DC was more than 60%. (2) ( 3-8) 105TIL can get from per 1g tumor tissue.The average number is 6 105 . It can be proliferated 30-90 times , the average number is about 60 times after culturing 11 days. (3) TIL stimulated by DC had very high killing activity on H22 cells and the killing rate was (71.52 3.88)%, which was much higher than those of TIL not stimulated by DC and splenocytes lymphocytes stimulated by DC as well as splenocytes lymphocytes not stimulated by DC, the killing rates of the last three ones were (50.95 3.02)%, (49.46 3.7)% and (9.77 1.39)% respectively. Conclusion: (1) A large number of high purity cells which have DC characteristic of morphology and function, were generated by GM-CSF and IL-4 from mouse bone marrow. (2) TIL can be obtained from mouse bearing tumor ,and it can be proliferated efficiently. (3) DC can induce more efficient and specific killing activity of TIL against H22 In vitro.