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MARC
논문명/저자명
Functional studies of culture-derived megakaryocytes and platelets from K562 cells by (R)-NALPCE = (R)-NALPCE 에 의해 분화된 K562 세포의 거핵구적인 기능 연구 및 유전자 연구 / 조현진
발행사항
서울 : 이화여자대학교 대학원, 2006.2
청구기호
TD 572.86 ㅈ523f
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xiii, 102 p. ; 26 cm
자료실
전자자료
제어번호
KDMT1200632961
주기사항
학위논문(박사) -- 이화여자대학교 대학원, 분자생명과학, 2006.2
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Title page=0,1,4

Abstract=i,5,4

Contents=v,9,3

List of Figures=viii,12,3

List of Tables=xi,15,1

Abbreviations=xii,16,2

1. INTRODUCTION=1,18,1

1.1. Background=1,18,4

1.2. Megakaryocytopoiesis (Megakaryocyte cell biology)=4,21,5

1.3. Differentiation system asing OP9 stromal cells=8,25,2

1.4. Integrin αIIbβ₃(Glyeoprotein IIb/IIIa) signaling(이미지참조)=9,26,3

1.5. Model of platelet production=11,28,5

1.6. Transcriptional regulation of megakaryocyte differentiation=15,32,4

2. MATERIALS＆METHODS=19,36,1

2.1. Reagents=19,36,2

2.2. Cell culture and differentiation of megakaryocytes=20,37,2

2.3. Cytotoxicity and growth assay=21,38,1

2.4. Flow cytometric analysis and cell sorting=21,38,2

2.5. Fibrinogen binding assay=22,39,1

2.6. Transmission electron microscopy=23,40,1

2.7. Fluorescence image=23,40,2

2.8. Immunoblotting=24,41,1

2.9. Oligonucleotide array=24,41,1

2.10. Total RNA preparation and RT­PCR=24,41,2

2.11. Northern hybridization=25,42,2

2.12. si­RNA transfection=26,43,1

3. RESULTS=27,44,1

3.1. Functional analysis of differentiated K562 cells by (R)­NALPCE=27,44,1

3.1.1. Cytotoxicity and growth of (R)­NALPCE­treated K562 cells=27,44,2

3.1.2. Expression of hematopoietic cell surface markers in (R)­NALPCE­treated K562 cells=28,45,5

3.1.3. Comparison data of hematopoietic specific cell surface markers between (R)­NALPCE­and PMA­treated K562 cells=32,49,2

3.1.4. Morphological changes of (R)­NALPCE­treated K562 cells cocultured with OP9 stromal cells=33,50,6

3.1.5. Expression of hematopoietic cell surface markers in (R)­NALPCE­treated K562 cells cocultured with OP9 stromal cells=38,55,4

3.1.6. Fibrinogen binding of (R)­NALPCE­treated K562 cells cocultured with OP9 stromal cells=41,58,7

3.1.7. Fluorescence image of culture­derived megakaryoeytes by (R)­NALPCE=47,64,4

3.1.8. Transmission electron micrograph of culture­derived megakaryocytes and platelet­like cells by (R)­NALPCE=50,67,4

3.2. Study of gene expression by (R)­NALPCE during megakaryocytic differentiation=54,71,1

3.2.1. Sustained MAPK expression during megakaryoeytic differentiation induced with (R)­NALPCE=54,71,4

3.2.2. Data analysis of oligonucleotide array=57,74,5

3.2.3. Induction of IL­8 gene expression by (R)­NALPCE during megakaryoeytie differentiation=62,79,1

3.2.4. IL­8 expression inhibited by siIL­8=62,79,5

3.2.5. Modulation of cell surface markers in (R)­NALPCE­treated K562 cells by siIL­8=66,83,4

4. DISCUSSION=70,87,7

REFERENCES=77,94,17

[Abstract in korean]=94,111,4

Acknowledgement=98,115,3

Appendix=101,118,2

Fig.1. Structural formula for (R)­NALPCE (M.W. 550.71)=2,19,1

Fig.2. Growth factors involved in megakaryocyte growth and maturation=6,23,1

Fig.3. Glycoprotein IIB/IIIa (IntegrinαIIbβ₃) structure(이미지참조)=12,29,1

Fig.4. Model illustrating the sequence of events throughout thrombin­ induced signaling process=13,30,1

Fig.5. The production mechanism of platelets from mature megakaryocytes=16,33,1

Fig.6. The cytotoxicity of (R)­NALPCE for K562 cells=29,46,1

Fig.7. The cytotoxicity of (R)­NALPCE for MEG­01 cells=30,47,1

Fig.8. The cell growth curve of (R)­NALPCE for K562 cells=31,48,1

Fig.9. Expression of hematopoietie cell surface markers in (R)­ NALPCE­treated K562 cells=34,51,1

Fig.10. Time­dependent CD 61 expression in (R)­NALPCE­treated K562 cells=35,52,1

Fig.11. Concentration­dependent CD 61 expression in (R)­NALPCE­treated K562 cells=36,53,1

Fig.12. Comparison data of hematopoietic specific cell surface markers between (R)­NALPCE­and PMA­treated K562 cells=37,54,1

Fig.13. Morphological changes of K562 cells cocultured with OP9 cells=39,56,1

Fig.14. Morphological differences between (R)­NALPCE­and PMA­treated K562 cells cocultured with OP9 stromal cells=40,57,1

Fig.15. Expression of hematopoietice cell surface markers in (R)­NALPCE­treated K562 cells cocultured with or without OP9 stromal cells=42,59,1

Fig.16. Concentration­dependent CD 41 expression in (R)­NALPCE­treated K562 cells with or without OP9 stromal cells=43,60,1

Fig.17. Time­dependent CD 41 expression in (R)­NALPCE­treated K562 cells with or without OP9 stromal cells=44,61,1

Fig.18. Time­dependent CD 42b (GP Ibα) expression in (R)­NALPCE­treated K562 cells with OP9 stromal cells=45,62,1

Fig.19. Fibrinogen binding to αIIbβ₃­expressing (R)­NALPCE­treated K562 cells cocultured with OP9 stromal cells(이미지참조)=48,65,1

Fig.20. Fibrinogen binding of CD 41­positive K562 cells differentiated by (R)­NALPCE in coculture system=49,66,1

Fig.21. Fluorescence image of (R)­NALPCE­treated K562 cells cocultured with OP9 stromal cells activated by a combination of agonist mixture=51,68,1

Fig.22. Transmission electron microscopy of megakaryocytes derived from (R)­NALPCE­treated K562 cells=52,69,1

Fig.23. Transmission electron microscopy of platelets derived from (R)­NALPCE­treated K562 cells=53,70,1

Fig.24. ERK1/2 activation in (R)­NALPCE­treated K562 cells=55,72,1

Fig.25. ERK1/2 activation in (R)­NALPCE­treated K562 cells with various inhibitors=56,73,1

Fig.26. Fold induction genes regulated by (R)­NALPCE and LY294002+(R)­NALPCE=61,78,1

Fig.27. The expression of IL­8 by (R)­NALPCE during megakaryocyte differentiation=63,80,1

Fig.28. Northern blot analysis of IL­8 RNA expression in (R)­ NALPCE­treated K562 cells=64,81,1

Fig.29. The expression of IL­8 by (R)­NALPCE­ and LY294002­ treated before (R)­NALPCE stimulation=65,82,1

Fig.30. IL­8 expression inhibited by siIL­8=68,85,1

Fig.31. Comparison data of megakaryocyte, neutrophil specific cell surface marker with or without siIL­8 in 73 μM (R)­NALPCE­treated K562 cells with or without OP9 stromal cells=69,86,1

Table1. Regulation of indivisual steps in megakaryocyte (MK) differentiation and thrombopoiesis=7,24,1

Table2. Up­regulated genes showing more than 2­fold changes by (R)­NALPCE=58,75,3

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