Title Page
Contents
ABSTRACT 8
Ⅰ. Introduction 9
Ⅱ. Material and Methods 10
2.1. Reagents 10
2.2. Cell culture 10
2.3. Transfection of HPV16 E6E7 10
2.4. Cell proliferation assay 11
2.5. RNA isolation and reverse-transcription PCR (RT-PCR) 11
2.6. Immunoblotting assay 13
2.7. Immunofluorescence staining 13
2.8. RGB profiling of immunofluorescence images 14
2.9. Evaluation of 3D spheroid construction ability 14
2.10. Statistical analysis 14
Ⅲ. Results 15
3.1. Detection of HPV16 E6/E7 in HDPCs 15
3.2. The effect of E6/E7 on proliferation and morphology in HDPCs 17
3.3. The effect of E6 on expression of p53 and cell cycle related factors 18
3.4. The effect of E7 on expression levels of pRb and transcription factor E2F 21
3.5. Responsiveness to Wnt/β-catenin signaling and hair follicle induction in E6/E7 immortalized HDPCs 24
Ⅳ. Discussion 27
Ⅴ. Conclusion 29
References 30
Abstract (in Korean) 34
Table 1. Primer sequences and product sizes used in RT-PCR. 12
Figure 1. Quantification of E6 and E7 expression in HDPCs. (A) Expression levels of E6, E7, and GAPDH were determined. (B) The relative band intensities... 16
Figure 2. Comparison of cell growth and morphology between primary and immortalized HDPCs. (A) Cell growth curves of primary and E6/E7 immortalized... 17
Figure 3. Expression levels of p53 and cell cycle related factors in HDPCs. (A-B) The TP53 gene and p53 protein expression level was determined. The relative... 20
Figure 4. Expression levels of pRb protein and E2F genes in HDPCs. (A) pRb was labeled with FITC (green) in cells, and nuclei were labeled with DAPI (blue). Scale... 23
Figure 5. Responses to Wnt/β-catenin signaling and hair follicle formation capability in HDPCs. (A) β-catenin was labeled with Cy3 (red) in cells, and... 26