Cell biological and biochemical study on the mechanism and oxidative regulation of photoaging

由于与光老化过程相关的生化变化尚未完全了解，累积的光氧化细胞损伤在皮肤光老化过程中起重要作用的理论仍然存在争议，在本研究中，我们已经证明，反复UVB照射后，SOD活性增加，并逐渐恢复到对照水平，但不受UVA照射的影响。据报道，急性光化损伤后SOD活性降低，可能是由于反复暴露于UVB而引起的，可能是对慢性光氧化应激的反应，然而，持续的累积应激可能会超过该系统的能力，从而导致光老化症状。我们还采用高效液相色谱法研究了无毛小鼠皮肤中GAG的主要二糖单元在慢性和重复紫外线照射后的变化。我们发现主要二糖单元的总量 ...更多信息 36周时UVA照射组增加。关于重复暴露36周后双糖组成的变化，对照组、UVA照射组和UVB照射组的DELTADi-HA（HA）减少，DELTADi-4S（DS）增加。这些结果首次证实了GAG在总量和组成上的精确改变，证实了先前的组织化学发现。我们还开发了三维培养系统补充L-抗坏血酸2-磷酸（Asc 2-P）使用真皮成纤维细胞。Asc 2-P的加入使成纤维细胞在体外形成真皮样三维结构，而不需要任何塑料培养皿的预处理。与单层培养系统相比，细胞外基质包围的多层成纤维细胞易于观察，并且主要二糖单位和羟脯氨酸的量显著增加，并且，当与单层培养系统相比时，其组成显示出更类似于真皮。使用这种培养系统，它应该是可能的调查从光老化皮肤获得的真皮成纤维细胞产生的胶原蛋白的生化变化。少

Since the biochemical changes associated with the photoaging process are not fully understood, the theory that accumulated photo-oxidative cellular injury contributes significantly to the photoaging process in skin still remains open to debate.In the present study, we have demonstrated that SOD activity was increased by repeated UVB irradiation and gradually returned to the control levels but unaffected by UVA irradiation. The SOD activity, which has been reported to decrease after acute actinic injury, is induced by repeated exposures to UVB presumably in response to the chronic photo-oxidative stress, however, continual cumulative stress may overwhelm the capacity of this system with resultant photoaging symptoms. We have also investigated the alteration of main disaccharide units of GAGs from hairless mice skin after chronic and repeated UV radiation using high performance liquid chromatography by our newly developed method. We found that the total amount of main disaccharide units … More increased by UVA irradiation at 36th week. Concerning the changes in the disaccharide composition after 36 weeks'repeated exposure, DELTADi-HA (HA) decreased and DELTADi-4S (DS) increased in the order of control, UVA- and UVB- irradiated group. These results, for the first time, demonstrate the precise alterations of GAGs both in total amount and composition confirming the previous histochemical findings. We have also developed the three-dimensional culture system supplemented with L-ascorbic acid 2-phosphate (Asc 2-P) using dermal fibroblasts. The addition of Asc 2-P rendered fibroblasts to the organization of the dermis-like three-dimensional structure in vitro without any pretreatments of plastic dish. Multilayred fibroblasts surrounded by extracellular matrices were readily observed and the amounts of both main disaccharide units and hydroxyproline are significantly increased and, furthermore, the composition revealed more similar to that of dermis when compared with monolayr culture system. Using this culture system, it should be possible to investigate the biochemical changes of collagen produced by dermal fibroblasts obtained from photoaged skin. Less