Induction Of Dendritic Cells-Cytokine-Induced Killer Cells In Vitro

Objective:To investigate the effect of interleukin15(IL-15) and bone marrow-derived dendritic cells (DC) on proliferation, phenotype and cytotoxicity against Hepal-6mouse hepatic carcinoma cells of cytokine-induced killer (CIK) cells.Methods:In vitro culture of bone marrow derived-DC was performed and then CD80, CD86, MHCII and CDllc expression were analyzed with flow cytometry (FCM). CIK and DC-CIK cells were culture either with culture medium containing IL-15or without IL-15, and then expression of CD3+、CD3+CD8+、CD3+NK1.1+cells were analyzed with FCM. In order to observed the effect of IL-15and DC on the proliferation ability of CIK cells, co-culture of CIK and DC were divided as follows: CIK, CIK+IL-15, CIK+DC, and CIK+IL-15+DC, and at the same time phenotype of these cells were analyzed with FCM. Both CIK cells and CIK cells simulated with IL-15were co-cultured with DC with a effector-target ratio of5:1. Cell proliferation ratio was analyzed with cell count, and phenotypes were analyzed with FCM. Cytotoxicity of CIK cells were then determined through detection of lactate dehydrogenase (LDH). All data were then analyzed with SPSS.Result:In vitro results showed that (5.5±0.5)×106dendritic cells (DC) were successfully culture and isolated from each mouse bone marrow, and flow cytometry results showed that isolated DC expressed CD80(95.0±1.2%), CD86(88.5±1.4%), MHC-Ⅱ (89.2±1.2%) and CD11c (83.3±1.7%).(4.5±0.5)×106CIK cells were successfully culture and isolated from each mouse spleen, and FCM results on day14showed that expression of CD3+,CD3+CD8+,and CD3+NK1.1+cells within the isolated CIK cells were (35.0±2.7%).(18.8±2.4%). and (17.1±2.5%) of respectively. Both DC stimulation and IL-15can increased proliferation of CIK cells, and expression of CD3+,CD3+CD8+,and CD34NK1.1+cells after stimulation were also significantly increased (P<0.05). The CIK cells number were (2.1±0.1)×10’and (2.4±0.2)×10’at day10and15after cultured, which showed significant increased when compared to the control group (0.5X10’).CIK or DC-CIK cells stimulated with IL-15showed significant increased in cytotoxicity when effector-target ration range from2.5:1to20:1. Furthermore, cytotoxicity of DC-CIK stimulated with IL-15was (58.75±4.34)%with effector-target ratio of20:1.Conclusion:DC and CIK cells were successfully cultured and isolated from mouse bone marrow and spleen under the stimulations of different cytokines. Both IL-15and DC increased the proliferation and surface antigens expression in CIK cells, and these effects were further increased when IL-15and DC were applied together. CIK cells stimulated with DC showed increased anti-tumor effects, and IL-15can further increased the antitumor effect of DC-CIK.