| Background and ObjectThe adoptive immunotherapy of immunocytes transfusion in vivo is the 4~th therapeutic tool following operation, chemo, radiotherapy. CIK cells are killed cells and tumor - nonspecific killer immunocytes induced by varied cytokines. the lethal effect is non - MHC - restricted , which has some good quality such as easy to proliferate in vitro,little side effect and so on. so CIK cells are considered having satisfactory clinical application prospect. However, CIK cells used in clinical research now have no enough amplification amount and ideal kill activity. And the killer mechanism are still not clear. Therefore, lucubrating the killer mechanism of CIK cells is important for further raising killer acitivity and efficiency of CIK cells and improve clinical therapeutic effect. The laryngeal cancer cell line Hep - 2 cells infected with a constructed recombinant adenovirus carrying human wild - type p53 and immunity - related genes - GM - CSF and B7 - 1 ( named as BB - 102) can control the generating and developing of tumor in different aspects. Wild - type P53 gene is one of the most well - studied tumor suppressor genes. GM - CSF play an important role in maturing angtigen presenting cells (APC). The interaction of the B7 - 1 molecule with the T cell CD28 receptor can induce secretion of some cytokines such as IL - 2 and prevent immune tolerance of tumor - special antigen, which is a critical factor of determining the first signal whether generates T cells functional or unresponsive activation, we presume the co - culturing tumor vaccine and CIK cells might induce anti - tumor - specific immunity. Our research aim at improving CIK cells cyto-toxicity and CIK cells clinical therapeutic effect by inducing CIK cells'specificimmunity by the Hep -2 tumor vaccine with BB - 102, which might serve as an alternative strategy for clinical adoptive immunotherapy.MethodThe laryngeal cancer cell line Hep — 2 were transfected or non - transfected with BB - 102, Mitiomycin was used to make Hep -2 with high expression of B7. 1 into tumor vaccine and co - culturing with CIK cells, the growth rate of CIK cells was determined by viable count. Phenotypes of CIK cells were identified by flow cytometric analysis. Cytotoxicity assay was performed by testing the activity of lactate dehydrogenase ( LDH) in culture supematants using CytoTox 96 Non - Radioactive Cytotoxicity Assay Kit.ResultAfter co - cultured 21 d, the growth rate of CIK cells stimulated by three genes - vaccine was significantly proliferated compared with that of CIK cells ( p <0.01,) , whereas Hep -2 tumor vaccine - CIK cells had a similar growth rate compared with CIK cells. The percentage of CD3 + CD56 + cells and CD3 + CDS + cells of CIK cells stimulated by BB - 102 - vaccine was significantly higher than that of CIK cells ( p < 0. 01) , whereas percentage of CD3 + CD56 + cells and CD3 +CD8 + cells of CIK cells stimulated by Hep -2 tumor vaccine wase similar with that of CIK cells. In Cytotoxicity assay, when the effector cells and target cells were co - cultured at ratio of 40:1, different CIK cells showed high cytolyt-ic activity in different time. When Hep - 2 cells were used as target cells , CIK cells stimulated by BB —102 — vaccine show significantly higher cytolytic activity compared with that of CIK cells ( p <0. 01 ,two — tailed t - test) , whereas CIK cells stimulated by Hep - 2 tumor vaccine had a similar cytolytic activity with CIK cells. When K562 cells (the target cells of natural killer cells) were used as target cells, each group all showed relatively similar cytolytic activity(p >0. 05).ConclusionCo — culturing Hep -2 tumor vaccine with BB -102 and CIK cells may increase the growth rate of CIK cells, the percentage of CD3 + CD56 + cells and CD3 + CD8 + cells of CIK cells and the cytolytic activity of the CIK cells, the tumor vaccine might increase tumor - specific CTL activity, and may provide a rationale for clinical adoptive immunotherapy. |
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