Study on the culture and the function of dendritic cells of patients with laryngeal squamous cell carcinoma in vitro / 中国耳鼻咽喉头颈外科

ABSTRACT

OBJECTIVE To culture dendritic cells from the peripheral blood ofpatients with laryngeal squamous cell carcinoma. METHODS Peripheral blood was collected from patients under sterile conditions. The mononuclear cells were separated by centrifugation through a density gradient (Ficoll-Hypaque). The adhesion cells, collected from the mononuclear cells, were cultured with interleukin-4 (IL-4, 100ng),granulocyte/macrophage colony stimulating factor (GM-CSF, 100ng) and tumor necrosis factor ? (TNF-?, 100ng) in RPMI 1640 medium for 7 days. The cell morphology was observed under electronic microscope, and the characteristic surface marker CD80 of the cells was analyzed by flow cytometry. MTT colorimetry was employed to assess the proliferation of the mixed lymphocytes. The mixed lymphocytes were stimulated by DC loaded with laryngeal carcinoma antigen. RESULTS A large quantity of cells characteristic of dendritic cell was obtained. Cytometry analysis showed that 33.76 % of mononuclear cells expressed the characteristic marker CD80. After being cultured for 7 days, the expression level of CD80 was significantly increased (69.15 %). Few DC with laryngeal carcinoma antigen could play a role in stimulating lymphocyte proliferation, and the proliferation effect was optimized when the ratio between DC and T cells was 1:5. CONCLUSION A large number of DCs with high purityand sufficient treatment dose can be cultured in vitro from the peripheral blood of patients with laryngeal carcinoma. This technique may play an important role in experimental studies for clinical applications.