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Study of the Molecular Mechanism of UVB Irradiation on Dermal Fibroblasts in Photoaging

Author: SongXiuZu
Tutor: BiZhiGang
School: Nanjing Medical University
Course: Dermatology and Venereology
Keywords: UVB radiation Fibroblast Mechanism of Action Rational probe Photoaging mRNA expression Lipid peroxidation products Signal transduction pathway Expression EGCG
CLC: R751
Type: PhD thesis
Year: 2004
Downloads: 287
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Abstract


Photoaging was thought to be due to the quantitative and qualitative alterations of structural proteins of the dermis induced by ultraviolet irradiation. Histological analysis showed the degeneration of collagen fibers and changes in the ratio of different types of collagen were caused by MMP. It was indicated that dermal fibroblasts played an important role in photoaging. The mechanism of photoaging had been concentrated in the study of UVA in the past years, while the effects of UVB had been little mentioned. Photoaging was a result of long-term ultraviolet accumulation and UVB could also produce a marked effect although its quantity was less than UVA into dermis. In this study, we explored the molecular mechanism of UVB irradiation on fibroblast in photoaging.Objectives: Part I was to explore the mechanism of ultraviolet-induced fibroblast apoptosis. In part II we studied the mechanism of ultraviolet-induced TNF-a and IL-1B secretions. Part III was to explore the mechanism of ultraviolet-induced oxidation damage.In part IV, we studied the effects of UVB on AP-1, MMP-1 and TIMP-1 of fibroblasts in vitro. C-jun is a key point in the signal transduction of ultraviolet irradiation. So in part V the inhibition mechanism of antisense c-jun ODN on MMP expression induced by UVB was explored.Methods: In this study, the apoptosis rate of fibroblasts was detected by flow cytometer. The activity of antioxidases and the content of MDA in the culture medium were measured by biochemical assay. The TNF-a, IL-1B, MMP-1 and TIMP-1 protein were detected by ELISA. The mRNA expression levels of Fas and Bcl-2, TNF-a and IL-1B, MMP-1 and TIMP-1 were determined by RT-PCR. The transcription factor Jun protein was measured by western blot. The C-Jun and C-Fos protein were measured by Western blot method.Results: Part I: The apoptosis peak was observed in both fibroblast groups treated with UVA or UVB respectively. After treated with EGCG, the apoptosis rate was decreased obviously. The mRNA level of Fas was both elevated after UVA or UVB irradiation and was inhibited by EGCG treatment. The mRNA expression of Bcl-2 was decreased after UVA irradiation, but had no obvious difference between the UVB group and the control group. The EGCG treatment could inhibit the mRNA change of Bcl-2 induced by UVA irradiation.Part II: The secretions and the mRNA expression of TNF-a and IL-1B were obviously increased after UVB irradiation compared with the control group. After treated with tea polyphenol and aloin, the secretions were both decreased. The data showed tea polyphenol and aloin treatment could relieve the skin damage induced by UVB through decreasing the secretions of TNF-a and IL-1B and their mRNA expression.Part III: EGCG could reduce the MDA accumulation induced by UVB and increase the activity of antioxidases. The mRNA expression of MMP-1 was increased after UVB irradiation and could be down-regulated by EGCG. However, the expression of TIMP-1 had no significant difference after UVB irradiation. EGCG could protect human fibroblasts against UVB damages.Part IV: EGCG could decrease transcription activity of Jun protein induced by UVB. The protein level of MMP-1 was increased by UVB irradiation, while no significant changes were observed in TIMP-1. The ratio of MMP-1 to TIMP-1 showed statistically significant difference compared with control. EGCG could decrease the ratio of MMP-1 to TIMP-1 by inhibiting the UVB-induced MMP-1 expression. It suggested EGCG could protect human fibroblasts against UVB damages by down-regulating the transcription activity of Jun protein and the expression of MMP-1. The MMP-1 to TIMP-1 ratio might play a more significant role in photoaging rather than MMP-1 or TIMP-1 alone.Part V: Results showed C-Jun protein level of fibroblasts induced by UVB was increased compared with non-irradiated controls, while C-Fos protein expression had no significant changes. Also UVB irradiation could obviously increase the pro-MMP-1 and MMP-3 synthesis. After transfection with c-jun antisense ODN at varying concentra

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