Title Page
Contents
Abstract 11
Ⅰ. INTRODUCTION 14
1. Structure and function of the skin 14
2. Psoriasis 17
A. Clinicopathological characters 17
B. Immunopathologic characters 18
C. Pathogenesis 22
D. Therapeutic strategies 25
3. In vitro reconstructed human skin models for psoriasis 26
4. Profilin-1 28
Ⅱ. MATERIALS AND METHODS 30
1. Subjects 30
2. Evaluation of psoriatic lesions 32
3. Producing reconstructed skin equivalent 33
4. Cell culture and treatment 37
5. Generation of PFN-1 knockdown cells 37
6. Transient overexpression of IκBζ in HaCaT cells 37
7. Transfection of siRNA targeting IκBζ 38
8. Analysis of cytokine secretion 38
9. PFN-1 uptake assay 39
10. ELISA 39
11. RNA isolation and quantitative real-time PCR 39
12. Immunostaining 40
13. Histological analysis 41
14. Western blot analysis 42
15. Statistical analysis 43
Ⅲ. RESULTS 46
PART Ⅰ. Development of an in vitro IL-17A-induced psoriasis-like skin equivalent model by self-assembly tissue engineering method 46
1. Reconstructed human skin equivalent consist of epidermis, dermis, and dermal fibroblasts-derived extracellular matrix (ECM) 47
2. Establishment of in vitro psoriasis-like reconstructed human skin model 53
PART Ⅱ. Profilin-1 prevents psoriasis pathogenesis through IκBζ regulation 60
3. PFN-1 is highly expressed in psoriatic skin lesions 61
4. IL-17A and TNF-α elevates PFN-1 expression, but only TNF-α induces its secretion 72
5. PFN-1 reduces IL-17A-induced psoriasis-like epidermal skin inflammation 88
6. PFN-1 affects keratinocyte proliferation and homeostatis 105
7. PFN-1 disturbs psoriasis-like skin inflammation via regulation of IκBζ 116
8. Suppression of IκBζ-mediated gene expression occurs by PFN-1 131
Ⅳ. DISCUSSION 140
Ⅴ. CONCLUSION 145
REFERENCES 149
ABSTRACT IN KOREAN 158
Table 1. Patient characteristics 31
Table 2. Primer sequences for qRT-PCR 44
Table 3. A list of primary antibodies 45
Figure 1. Scheme of producing the oelf-ascembled reconcti acted skin equivalent 34
Figure 2. The process of producing reconstructed skin equivalent. 35
Figure 3. Biopsies from the reconstructed skin equivalent. 36
Figure 4. Histological analysis of reconstructed skin equivalent. 50
Figure 5. Transmission electron microscopy analysis of reconstructed skin equivalent. 52
Figure 6. The self-assembled reconstructed skin equivalents. 56
Figure 7. IL-17A-induced psoriasis-like skin equivalents express psoriasis-related proteins. 59
Figure 8. PFN-1 is highly expressed in psoriasis lesional skin. 65
Figure 9. Psoriasis-associated biomarkers are increased in the lesional skin of patients with psoriasis. 69
Figure 10. Secreted PFN-1 in serum of psoriasis patients positively correlated disease severity. 71
Figure 11. TNF-α and IL-17A increases PFN-1 expression in human keratinocytes. 77
Figure 12. PFN-1 expression in the human primary keratinocytes in a time-dependent manner. 79
Figure 13. IL-17A-induces psoriasis-associated biomarker expression in human keratinocytes. 81
Figure 14. PFN-1 expression in human primary dermal fibroblasts. 83
Figure 15. Knockdown of PFN-1 in HaCaT cells increases the expression of psoriasis-associated markers. 85
Figure 16. TNF-α induces PFN-1 secretion from human keratinocytes, but IL-17A does not. 87
Figure 17. PFN-1 enters the human keratinocytes by clathrin-mediated endocytosis. 93
Figure 18. PFN-1 reduces IL-17A-induced psoriasis-like skin inflammation in human keratinocytes. 96
Figure 19. Psoriasis-associated genes are induced in HaCaT cells by IL-17A stimulation. 98
Figure 20. The effect of rPFN-1 in suppressing the increased psoriasis-associated markers in PFN-1-depleted HaCaT cells. 100
Figure 21. TNF-α-conditioned medium reduces IL-17A-induced psoriasis-associated inflammatory gene expression. 102
Figure 22. Analysis of cytokine secretion from human keratinocytes. 104
Figure 23. PFN-1 reduces IL-17A-induced cell proliferation. 108
Figure 24. PFN-1 relates to MKI67 regulation in human keratinocytes. 110
Figure 25. PFN-1 alleviates aberrant epidemal differentiation. 113
Figure 26. IL-22, an important cytokine in psoriasis involved in epidermal hyperplasia, is not affected by PFN-1. 115
Figure 27. IκBζ is highly expressed in psoriatic lesions. 119
Figure 28. PFN-1 knockdown induces IκBζ expression. 121
Figure 29. PFN-1 restrains IL-17A-induced IκBζ expression in human keratinocytes. 123
Figure 30. PFN-1 suppresses IκBζ translocation to the nucleus. 125
Figure 31. Effect of PFN-1 on IκBζ-mediated gene expression and IκBζ nuclear translocation in IL-17A-stimulated human... 128
Figure 32. Comparison of the reduction rate by PFN-1 against the Il-17A-induced mRNA expression. 130
Figure 33. IκBζ overexpression induces psoriasis-associated marker expression, and it does not abolished by PFN-1 treatment. 133
Figure 34. IκBζ knockdown abrogates IL-17A-induced inflammatory gene expression in PFN-1-depleted HaCaT cells. 136
Figure 35. PFN-1 regulated IκBζ via phosphorylation of NF-κB p65. 139
Figure 36. A proposed mechanism for the regulation of psoriasis pathogenesis by PFN-1. 148