Yin L, Morita A, Tsuji T
Department of Dermatology, Nagoya City University Medical School, Japan.
Arch Dermatol Res. 2000 Apr;292(4):188-94. doi: 10.1007/s004030050476.
Epidemiologic studies have indicated the association between tobacco smoking and skin aging, but the exact mechanism of tobacco smoke-induced premature skin aging is currently unknown. In this study, we investigated the alterations of collagen, matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) in human fibroblasts treated with tobacco smoke extract. Human fibroblasts were exposed to different concentrations of water-soluble extract from tobacco smoke. Human fibroblasts irradiated with ultraviolet A1 (UVA1) were used as positive controls because the mechanism of UVA1-mediated MMP expression has been well characterized. The expression of MMP and TIMP was analyzed semiquantitatively following reverse transcriptase-polymerase chain reaction. Production of type I and type III collagens was detected by Western blotting and biosynthesis of new collagen was assessed by 3H-proline incorporation. Upon treatment with tobacco smoke extract or UVA1 irradiation, the expression of MMP-1 and MMP-3 mRNA was significantly increased in a dose-dependent manner. Maximum induction was observed with 25 microl/ml tobacco smoke extract. In contrast, the expression of TIMP-1 and TIMP-3 mRNA remained unchanged. Western blotting of the supernatant revealed that type I and type III collagens were decreased as compared with untreated controls. Collagen biosynthesis was significantly reduced by 40.1% following treatment with 25 microl/ml tobacco smoke extract. Sodium azide, L-ascorbic acid and Trolox (a water-soluble vitamin E) prevented both the UVA1- and the tobacco-induced alteration of MMP-1. These observations suggest that the imbalance of connective tissue matrix components might contribute to the molecular basis for premature skin aging in smokers. They also suggest that reactive oxygen species including singlet oxygen mediate this process.
流行病学研究表明吸烟与皮肤老化之间存在关联，但目前尚不清楚烟草烟雾导致皮肤过早老化的确切机制。在本研究中，我们调查了用烟草烟雾提取物处理的人成纤维细胞中胶原蛋白、基质金属蛋白酶（MMPs）和金属蛋白酶组织抑制剂（TIMPs）的变化。将人成纤维细胞暴露于不同浓度的烟草烟雾水溶性提取物中。用紫外线A1（UVA1）照射的人成纤维细胞用作阳性对照，因为UVA1介导的MMP表达机制已得到充分表征。逆转录聚合酶链反应后，对MMP和TIMP的表达进行半定量分析。通过蛋白质印迹法检测I型和III型胶原蛋白的产生，并通过3H-脯氨酸掺入评估新胶原蛋白的生物合成。用烟草烟雾提取物处理或UVA1照射后，MMP-1和MMP-3 mRNA的表达以剂量依赖性方式显著增加。在25微升/毫升烟草烟雾提取物中观察到最大诱导作用。相比之下，TIMP-1和TIMP-3 mRNA的表达保持不变。上清液的蛋白质印迹显示，与未处理的对照相比，I型和III型胶原蛋白减少。用25微升/毫升烟草烟雾提取物处理后，胶原蛋白生物合成显著降低了40.1%。叠氮化钠、L-抗坏血酸和生育酚（一种水溶性维生素E）可防止UVA1和烟草诱导的MMP-1改变。这些观察结果表明，结缔组织基质成分的失衡可能是吸烟者皮肤过早老化的分子基础。它们还表明，包括单线态氧在内的活性氧介导了这一过程。
