Title Page
Contents
ABSTRACT 9
국문초록 11
Ⅰ. INTRODUCTION 13
Ⅱ. MATERIALS AND METHODS 16
1. MATERIALS 16
2. METHODS 17
2.1. Preparation of Diesel Particulate Matter (DPM) 17
2.2. Preparation of ethanol extracts of Hibiscus cannabinus L. flower (HCFE) 17
2.3. Analysis of total anthocyanin content 20
2.4. HPLC analysis 21
2.5. DPPH radical scavenging assay 22
2.6. Cell culture 22
2.7. Cell Viability Assay 23
2.8. Measurement of Reactive Oxygen Species (ROS) Production 24
2.9. Real-Time Polymerase Chain Reaction (PCR) 25
2.10. Western blot analysis 27
2.11. Statistical Analysis 28
Ⅲ. RESULTS 29
1. Total anthocyanin content in HCFE 29
2. The caffeic acid contents in HCFE 31
3. DPPH radical scavenging activity in HCFE 33
4. The effect of DPM and HCFE on cell viability in HaCaT cells 34
5. Inhibition of ROS Production by HCFE in HaCaT Cells 36
6. HCFE inhibit inflammation in DPM-stimulated HaCaT cells 38
6.1. Inhibition of IL-1 β, IL-6, and IL-8 mRNA expression by HCFE in DPM-stimulated HaCaT cells 38
6.2. Inhibition of CCL17 and MDC mRNA Expression by HCFE in DPM-stimulated HaCaT cells 41
7. Effects of HCFE on NF-κB signaling pathways in DPM-stimulated HaCaT cells 43
Ⅳ. DISCUSSION 44
REFERENCES 47
Table 1. HPLC conditions 21
Table 2. Sequences of oligonucleotide primer for RT-PCR 26
Table 3. The contents of total anthocyanin in HCFE 30
Fig. 1. Hibiscus cannabinus L. flower. (A) original, and (B) dry flower 18
Fig. 2. Preparation ethanol extracts of HCF 19
Fig. 3. Chemical structure of anthocyanin 29
Fig. 4. Chemical structure of caffeic acid 31
Fig. 5. HPLC analysis of HCFE 32
Fig. 6. DPPH radical scavenging activity of HCFE. 33
Fig. 7. Cell viability of DPM and HCFE-treated in HaCaT cells. 35
Fig. 8. Inhibition of ROS production by HCFE in HaCaT Cells. 37
Fig. 9. Inhibition of IL-1ß, IL-6 and IL-8 mRNA expression by HCFE in DPM-stimulated HaCaT cells. 40
Fig. 10. Inhibition of CCL17 and MDC expression by HCFE in DPM-stimulated HaCaT cells. 42
Fig. 11. Effects of HCFE on NF-κB signaling pathways in DPM-stimulated HaCaT cells. 43