| Skin photoaging is induced by ultraviolet (UV) irradiation, which results from the comitant effects of UVA and UVB. The mechanism of skin photoaging had been concentrated in the study of UVA in the past years. While the effects of UVB had been little mentioned. In recent years, with the improvement of living standards and increase of outdoor activities, people have attached more importance to the protection of skin photoaging. Studies of the photoaging preventive effects of botanical antioxidants have been gained considerable attention. Trans-Zeatin is a plant hormone,purified from zea mays, which possesses a variety of biological actions, such as anti-oxidant properties, anti-apoptosis activity and anti-inflammatory effects. Trans-Zeatin also has gerontomodulatory, youth preserving and antiaging effects on serially passaged adult skin fibroblasts undergoing aging in vitro. Although many studies have revealed the functions of trans-Zeatin, its effects on skin photoaging has not yet been previously reported. In this study (including three parts), we have attemped to investigate the effect and action mechanisms of trans-Zeatin on matrix metalloproteinase-1 (MMP-1), matrix metalloproteinase-3 (MMP-3) and type I procollagen expression in UVB- irradiated human skin fibroblasts (HSFs).Objective: In partⅠ, we explored the effects of UVB on MMP-1, MMP-3 expression and the protective effects of trans-Zeatin against UVB-induced MMP-1, MMP-3 expression in human skin fibroblasts (HSFs) in vitro. In partⅡ, we investigated the signaling pathways of UVB-induced MMP-1 expression and trans-Zeatin against UVB-induced MMP-1 expression in HSFs. In partⅢ, we examined the effects of trans-Zeatin on type I procollagen synthesis in UVB-irradiated HSFs and revealed the possible mechanism of its action. Methods: The cell viability of trans-Zeatin on HSFs and UVB-exposed HSFs was detected by MTT assay. Amounts of MMP-1, MMP-3 and type I procollagen in the culture supernatants were quantitated by enzyme-linked immunosorbent assay kits. The expression of type I procollagen mRNA was detected by RT-PCR. The change of ERK1/2, JNK, p38 MAPK and c-Jun phosphorylation, and MMP-1, TGFβ, Smad3 as well as Smad7 expression in cells were determined by Western blot.Results:PartⅠ: We found that trans-Zeatin did not affect cell viability at the concentration lower than 150μM. While the concentration was over 600μM, trans-Zeatin affected the cell viability and had cytotoxic effect on HSFs. Trans-Zeatin (20-80μM) dose-dependently enhanced the cell viability in HSFs irradiated by UVB. The results also showed that UVB irradiation significantly enhanced MMP-1 and MMP-3 expression. Pretreatment with trans-Zeatin markedly inhibited UVB-induced MMP-1 expression in a dose-dependent manner versus the only UVB-irriadiated group. However, pretreatment with trans-Zeatin almost had no effect on UVB-induced MMP-3 expression. Moreover, trans-Zeatin itself did not change the expression of MMP-1 and MMP-3 in HSFs.PartⅡ: The results showed that ERK1/2, JNK, p38 MAPK and c-Jun phosphorylations were significantly enhanced by UVB irradiation, compared with the control group. Pretreatment with trans-Zeatin (80μM) markedly inhibited UVB-induced ERK1/2 phosphorylation. Trans-Zeatin(20-80μM) dose-dependently inhibited UVB-induced JNK and c-Jun phosphorylations. Trans-Zeatin also inhibited UVB-induced p38 MAPK phosphorylation in a dose-dependent manner at the concentration of 40μM and 80μM. As expected, PD98059, a specific ERK1/2 inhibitor , SP600125, a specific JNK inhibitor and SB203580, a specific p38 MAPK inhibitor effectively inhibited UVB-induced phosphorylations of ERK1/2, JNK and p38 MAPK respectively. Moreover, PD98059(20μM), SP600125(10μM), SB203580(10μM)and trans-Zeatin(80μM) significantly suppressed UVB-induced MMP-1 secretion, which is consistent with the above results.PartⅢ: In this study, the results indicated that mRNA expression and protein secretion of type I procollagen as well as synthesis of TGFβand Smad3 were significantly reduced by UVB irradiation, while synthesis of Smad7 was markedly enhanced by UVB irradiation, compared with the control group. Trans-Zeatin(40μM-80μM) dose-dependently increased mRNA expression and protein secretion of type I procollagen in UVB-irradiated HSFs. However, trans-Zeatin itself did not change type I procollagen mRNA expression and protein secretion. Moreover, Trans-Zeatin also increased synthesis of TGFβand Smad3, and reduced synthesis of Smad7 in HSFs irradiated by UVB.Conclusion: Our study demonstrates that UVB irradiation could significantly enhance MMP-1 and MMP-3 expression , and suppress type I procollagen expression and secretion in HSFs. UVB-induced MMP-1 expression is mediated by MAPK(ERK1/2、JNK、p38)/AP-1(c-Jun)signaling pathways. Trans-Zeatin could inhibit UVB-induced MMP-1 expression, which may be mediated by the inhibition of MAPK(ERK1/2、JNK、p38)/AP-1(c-Jun) signaling pathways in HSFs. Trans-Zeatin also could promote type I procollagen expression and synthesis, which may be mediated by stimulation of the TGFβ/Smad signaling in UVB-exposed HSFs. Therefore, we suggest that trans-Zeatin might be considered as a potential agent for the management of skin photoaging. |
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