크로만 유도체 TAL-1036의 GM-CSF 활성 증강 효능 = Enhancement of GM-CSF activity by a chroman derivative, TAL-1036

다국어 초록 (Multilingual Abstract)

Granulocyte-macrophage colony stiomulating factor (GM-CSF) is a protein secreted by macrophages, T cells, mast cells, endothelial cells and fibroblasts. It is a cytokine that functions as a white blood cell growth factor. GM-CSF stimulates stem cells to produce granulocytes and monocytes. GM-CSF is currently used as a medication to stimulate the production of white blood cells following chemo or radio-therapy. It is also used to accelerate white blood cell recovery following autologous bone marrow transplantation. In an effort to search for and characterize compounds enhancing the efficacy of GM-CSF, several hundreds of compounds were screened for their ability to stimulate the growth of a GM-CSF-dependent cell line, NFS-60. Chroman-2-carboxylic Acid-N-(3, 4-dimethoxyphenyl)amide (TAL-1036) was found to enhance the growth of NFS-60 cells in the presence of low concentration of GM-CSF. Addition of TAL-1036 to cultures of NFS-60 cells increased the proliferation of the cells almost 100 percent compared to GM-CSF only control. However, TAL-1036 by itself did not induce increase of the proliferation of NFS-60. TAL-1036 did not affect the growth of other cell lines such as IL-2-dependent cell line (CTLL-2), IL-3-dependent cell line (Y-16), and RAW 264.7 cells. The effects of TAL-1036 on the generation of DCs from mouse BM cells were examined in vitro. Addition of TAL-1036 together with low concentration of GM-CSF to cultures of BM cells increased the generation of DCs almost 40% in number compared to the GM-CSF only control. To examine whether the DCs generated with TAL-1036 is functional, the MHC-restricted antigen presentation capability, phagocytic activity, expression level of cell surface molecule and secretion of cytokines were examined. DCs generated with TAL-1036 exhibited almost the same functional characteristics compared to the DCs generated in high concentration of GM-CSF. These results suggest that TAL-1036 enhances GM-CSF activity and may have potential to be developed as an immunostimulating drug that? can restore blood leukocyte counts in patients receiving chemo- or radio-therapy.

목차 (Table of Contents)

• I. 서 론 1
• II. 재료 및 방법 5
• 1. 세포 및 세포주 5
• 2. 세포의 배양 5
• 3. 크로만 화합물의 합성 6
• 4. NFS-60 세포의 GM-CSF 의존성 확인 및 GM-CSF 효능 증강 물질 탐색 7
• 5. [3H]-thymidine uptake assay를 이용한 세포 증식력 측정 7
• 6. TAL-1036이 cytokine 생산에 미치는 영향
• 6-1. TAL-1036에 의한 세포의 자극 8
• 6-2. NO assay 9
• 6-3. TNF-α assay 9
• 7. 골수 세포로부터 수지상세포 (BM-DC)의 생성 10
• 8. Microencapsulated OVA의 제조 11
• 9. BM-DC의 표면 분자 발현 분석 11
• 10. 탐식력 측정 12
• 11. 외부 항원의 MHC class Ⅰ분자를 통한 제시능 측정 13
• 12. 외부 항원의 MHC class Ⅱ 분자를 통한 제시능 측정 14
• 13. BM-DC의 cytokine 생산력 측정
• 13-1. BM-DC의 자극 15
• 13-2. TNF-α assay 15
• 13-3. IL-1β assay 16
• III. 결과 17
• 1. NFS-60 세포의 GM-CSF 농도 의존성 17
• 2. Chroman 유도체가 세포 성장에 미치는 영향 19
• 3. TAL-1036이 NFS-60 세포의 증식에 미치는 영향 22
• 4. GM-CSF 농도에 따른 TAL-1036의 효능 증강 25
• 5. TAL-1036이 CTLL-2, Y-16, RAW 264.7 세포의 증식에 미치는 영향 27
• 6. Cytokine 생산에 미치는 TAL-1036의 효능
• 6-1. Nitric oxide 생산에 미치는 영향 31
• 6-2. TNF-α 생산에 미치는 영향 31
• 7. TAL-1036의 첨가가 BM-DC의 생성에 미치는 영향 34
• 8. TAL-1036의 첨가에 의해 생성된 BM-DC의 특성 및 기능
• 8-1. 세포 표면 분자의 발현 37
• 8-2. 탐식력 39
• 8-3. 외부 항원 제시능 41
• 8-4. Cytokine 생산능 44
• Ⅳ. 결론 및 고찰 46
• Ⅴ. 참고 문헌 51