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A Study of Making DC Tumor Vacine by high Intensity Focused Ultrasound and Observing Its Antitumor and Promoting Tumor Cells Apoptosis Effect

Author: YangXue
Tutor: YeXin
School: Taishan Medical College
Course: Geriatrics
Keywords: Dendritic cells High Intensity Focused Ultrasound Dose Tumor antigen Tumor vaccine Therapeutic effect Apoptosis
CLC: R73-36
Type: Master's thesis
Year: 2007
Downloads: 12
Quote: 0
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Purpose: (1) the establishment of in vitro amplified mouse dendritic cells (dendritic cell DC); ② The application of high intensity focused ultrasound (high intensity focused ultrasound HIFU) irradiated tumor cells for tumor antigen and sensitized DC to prepare DC tumors vaccines; observed of DC vaccine prepared by this method on the role of active treatment of tumor cells in mice have been vaccinated. The ③ observed apoptosis of tumor cells, and explore its therapeutic mechanism, which seek new avenues for research in clinical oncology treatment, and also to explore new ideas for the development of tumor vaccines, and to open up new fields of application for HIFU. Method: ① Application 10ng/ml of white interleukin -4 (interleukin, IL-4) and 10ng/ml granulocyte - macrophage colony stimulating factor (granulocyte-macrophage colony stimulating factor, 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, CD86, H-2Kd and I-Ad expression; ② application of different doses of HIFU irradiation in vitro culture CT-26 tumor cells by trypan blue staining, MTT assay the number of viable cells, tumor cell viability was observed, and the morphological changes of the cells was observed, to find the cause of the work of preparation of tumor antigen dose. HIFU irradiation in vitro culture ③ with dose (1000W/cm2 × 30s) CT-26 tumor cells, tumor antigens were prepared and sensitized DC DC vaccine preparation. ④ DC Vaccine Therapeutic Action Research: the establishment of the HIFU irradiation group (A), the repeated freezing and thawing group (B), a simple DC group (C), as well as the negative control group (D). Normal BALB / c mice were inoculated subcutaneously Activity of CT-26 colon cancer cells (5 × 105 / PCS), after 7 days, respectively, by subcutaneous injection HIFU irradiated DC vaccine prepared by repeated freezing and thawing the DC vaccine prepared, pure DC cells, as well as the same amount of serum-free culture medium treatment of mice. A week later, the same dose strengthening treatment. 7 days after the part of the rats were sacrificed to remove the tumor measuring tumor weight, the mice were cured tumors the NW and tumor volume remaining for tumor growth was observed in mice and survival time and survival. Whether the differences were compared among groups. ⑤ DC tumor vaccine to promote tumor cell apoptosis observed: stripping tumors in mice were made into single cell suspensions and tissue sections using Annexin Ⅴ -FITC/PI measured by TUNEL method for apoptosis of tumor cells, comparing whether the differences. Results: ① mouse bone marrow cells cultured with IL-4 and GM-CSF for 3 days, visible morphological changes, irregular cell morphology and growth in clusters. 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) application of the HIFU irradiation mice of different doses of CT-26 colon cancer cells, with the increase of the the HIFU irradiation dose of tumor cell viability rapidly reduced, and gradually increasing the tumor cell debris. Ultrasonic dose 1000W/cm2 × 30 seconds, the cells were all dead, and the loss of cell morphology and all be torn to pieces. Therefore, the choice 1000W/cm2 × 30 seconds as a preparation of tumor cell lysis solution (tumor antigen) dose. ③ HIFU irradiation group, the group repeated freeze-thaw cycles, respectively, with a simple DC group, negative control group, tumor weight, volume, mice NW. Differences were significant (P lt; 0.01 or P lt; 0.05); with HIFU by irradiation of significance group, repeated freezing and thawing group with a single DC group, negative control group of mice survival time and survival rate, the difference was statistically significant (P lt; 0.01 or P lt; 0.05); with HIFU by irradiation group and repeated freezing and thawing group tumor weight, volume difference was also statistically significant (P lt; 0.01 or P lt; 0.05) with HIFU by irradiation group and repeated freezing and thawing group mice NW, mice survival time and survival differences nor significant (P gt; 0.05). ④ HIFU irradiation group, repeated freezing and thawing group with a single DC group, negative control group differences in tumor cell apoptosis also significantly significance (P lt; 0.01 or P lt; 0.05). HIFU irradiation group and repeated freezing and thawing group of tumor cell apoptosis difference was also statistically significant (P lt; 0.05). DC vaccines prepared HIFU irradiation and repeated freezing and thawing prepared DC vaccines have resistance to tumor growth and promote tumor cell apoptosis and treatment of cancer, and the former as the latter. Conclusion: ① application of 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. ② HIFU tumor cells can be inactivated tumor cells and can crusher, 1000W/cm2 × 30s can be used as a preparation of tumor cell antigen dose. This method to obtain tumor antigens can be effectively sensitized DC, preparation of DC vaccines. ③ HIFU irradiated tumor vaccine prepared a therapeutic effect on mouse tumor and can promote apoptosis of tumor cells and tumor vaccine group prepared better than freezing and thawing.

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