A Stduy of High Intensity Focuse Ultrasound in Loading Tumour Antigen of Dendritic Cells and Lead T Cell to Secrete Interferon-Gamma in Vitro
|School||Taishan Medical College|
|Keywords||Dendritic cells High Intensity Focused Ultrasound Dose Tumor antigen Load Cytokines IL-12 IFN-γ Tumor vaccine|
Objective: To establish vitro amplification of mouse dendritic cells (dendritic cell, DC); explore irradiated tumor cells were prepared using high intensity focused ultrasound (high intensity focused ultrasound, HIFU) tumor antigen pulsed DC and preparation of DC tumors the feasibility of the vaccine, for HIFU to open up new fields of application. Methods: ① Application 1ng/ml IL-4 and lOng / ml GM-CSF induced by cultured mouse bone marrow cells, light microscopy and electron microscopy morphological changes in DC during development; flow cytometry DC surface CD80 and CD86 H-2Kd \u0026 the expression of I-Ad; ② fixed irradiation time, the application of different power the HIFU irradiation CT26 cells strains after trypan blue staining, MTT assay determination of the number of live tumor cells, while using the trypan blue observe the morphological changes of the tumor cells, the analysis of the relationship between the HIFU dose and survival of tumor cells, and to understand the dose - response relationship; (3) fixed irradiation time and ultrasonic intensity irradiated CT26 cell suspension prepared tumor cell antigen and sensitized DC pulsed DC 48 hours after incubation with T cells reactive to extract the supernatant, the supernatant IL-12, IFN-γ The content is detected by ELISA, and compared with the freeze-thaw group, simple the CT26, control group. Results: ① cultured mouse bone marrow cells in combination with IL-4 and GM-CSF 3 days after the visible change in cell morphology and cell irregular. 6 to 8 days of culture, the cell surface appear more burr-like protrusions, elongated, typical DC characteristics. Flow cytometry DC surface molecules CD80, CD86, H-2Kd and I-Ad expression; ② decreased rapidly with the the HIFU irradiation dose increase tumor cell survival, tumor cell debris gradually increased. Ultrasound dose 1000W/cm2 × 30 seconds, cell survival, and tumor cells completely lost cell morphology, all being torn to pieces; ③ is detected by ELISA HIFU, freezing and thawing, simplex CT26 three groups of IL-12, the supernatant, IFN-γ was significantly higher than that of the control group (p lt; 0.05), HIFU, freeze-thaw two groups supernatant IL-12, IFN-γ content was significantly higher than that of CT26 group (p lt; 0.05) But H1FU, between the freeze-thaw group there was no statistically significant significance (p gt; 0.05). Conclusion: IL-4 DC cultured mouse bone marrow cells and GM-CSF, a large number of amplification mature DC culture in line with its own characteristics; HIFU tumor cells inactivated tumor cells can crusher of HIFU prepared tumor antigen sensitized in vitro DC, and and enables DC maturation secrete large amounts of IL-12, while able to induce T cells to secrete large amounts of IFN-γ.