CAJ | 학술논문

目的：研究连续长波紫外线（UVA）照射对人皮肤成纤维细胞胞吞和胞内降解细胞外弹性蛋白能力的影响，探讨光老化皮肤弹性蛋白堆积的机制。方法将人皮肤成纤维细胞分为空白对照组和连续 UVA照射组，连续 UVA 照射组予连续7 d 每天9.9 J/cm2 UVA 照射以构建人皮肤成纤维细胞慢性光损伤模型。用流式细胞仪和衰老相关β半乳糖苷酶染色法分别检测细胞周期和细胞老化程度以验证模型。用荧光示踪技术和共轭淬灭技术，比较两组细胞对培养基中弹性蛋白的胞吞情况和对胞吞进入细胞的弹性蛋白的降解情况差异。结果与空白对照组相比，连续 UVA 照射组 G1期细胞比例和衰老细胞比例明显增高，人皮肤成纤维细胞慢性光损伤模型构建成功。空白对照组人皮肤成纤维细胞在与弹性蛋白共孵育24、48、72 h 后对弹性蛋白的胞吞率依次为（10.0±1.4）%、（27.8±4.2）%、（39.9±4.1）%，连续 UVA 照射组人皮肤成纤维细胞在对应时间点的胞吞率依次为（9.9±1.6）%、（28.3±5.1）%、（42.0±5.7）%，在相同时间点两组细胞对弹性蛋白的胞吞率差异均无统计学意义（均 P >0.05）。空白对照组人皮肤成纤维细胞在与弹性蛋白共孵育24、48、72 h 后，胞内降解内吞入胞的弹性蛋白的细胞百分率依次为（7.7±0.9）%、（22.8±1.8）%、（33.9±3.1）%，而连续 UVA 照射组人皮肤成纤维细胞在对应时间点依次为（4.2±1.1）%、（16.5±2.4）%、（26.7±2.6）%，均较对照组显著降低（均 P <0.05）。结论连续7 d 每日9.9 J/cm2 UVA 照射对人皮肤成纤维细胞胞吞弹性蛋白的能力没有明显影响，却使细胞对胞吞的弹性蛋白的胞内降解能力下降。
목적：연구련속장파자외선（UVA）조사대인피부성섬유세포포탄화포내강해세포외탄성단백능력적영향，탐토광노화피부탄성단백퇴적적궤제。방법장인피부성섬유세포분위공백대조조화련속 UVA조사조，련속 UVA 조사조여련속7 d 매천9.9 J/cm2 UVA 조사이구건인피부성섬유세포만성광손상모형。용류식세포의화쇠로상관β반유당감매염색법분별검측세포주기화세포노화정도이험증모형。용형광시종기술화공액쉬멸기술，비교량조세포대배양기중탄성단백적포탄정황화대포탄진입세포적탄성단백적강해정황차이。결과여공백대조조상비，련속 UVA 조사조 G1기세포비례화쇠로세포비례명현증고，인피부성섬유세포만성광손상모형구건성공。공백대조조인피부성섬유세포재여탄성단백공부육24、48、72 h 후대탄성단백적포탄솔의차위（10.0±1.4）%、（27.8±4.2）%、（39.9±4.1）%，련속 UVA 조사조인피부성섬유세포재대응시간점적포탄솔의차위（9.9±1.6）%、（28.3±5.1）%、（42.0±5.7）%，재상동시간점량조세포대탄성단백적포탄솔차이균무통계학의의（균 P >0.05）。공백대조조인피부성섬유세포재여탄성단백공부육24、48、72 h 후，포내강해내탄입포적탄성단백적세포백분솔의차위（7.7±0.9）%、（22.8±1.8）%、（33.9±3.1）%，이련속 UVA 조사조인피부성섬유세포재대응시간점의차위（4.2±1.1）%、（16.5±2.4）%、（26.7±2.6）%，균교대조조현저강저（균 P <0.05）。결론련속7 d 매일9.9 J/cm2 UVA 조사대인피부성섬유세포포탄탄성단백적능력몰유명현영향，각사세포대포탄적탄성단백적포내강해능력하강。
Objective To investigate the effects of repetitive ultraviolet A(UVA) radiation on the endocytosis and intracellular degradation of extracellular elastin by human dermal fibroblasts(HDFs), and to explore the mechanism responsible for elastin accumulation in photoaging skin. Methods Cultured HDFs were divided into two groups:repetitive UVA radiation group treated with UVA radiation(9.9 J/cm2)once a day for seven consecutive days to establish a chronic photodamage model, and blank control group receiving no treatment. For the verification of the model, flow cytometry was performed to detect cell cycle, and senescence-associated β-galactosidase staining to determine the degree of cell senescence. Fluorescent tracer technique and conjugated polymer-based fluorescence quenching technique were conducted to observe the endocytosis and intracellular degradation of extracellular elastin in culture media by HDFs in these groups. Results Compared with the blank control group, the repetitive UVA radiation group showed increased proportions of G1-phase cells and senescent cells, which confirmed the successful establishment of chronic photodamage model in the repetitive UVA radiation group. After coculture with elastin for 24, 48 and 72 hours, the endocytosis rate of elastin was 10.0% ± 1.4%, 27.8% ± 4.2% and 39.9% ± 4.1% respectively in the blank control group, 9.9% ± 1.6%, 28.3% ± 5.1% and 42.0% ± 5.7% respectively in the repetitive UVA radiation group, with no significant difference between the two groups at the three time points (all P > 0.05). The percentage of cells showing intracellular degradation of extracellular elastin was significantly lower in the repetitive UVA radiation group than in the blank control group (4.2% ± 1.1% vs. 7.7% ± 0.9% at 24 hours, 16.5% ± 2.4% vs. 22.8% ± 1.8% at 48 hours, 26.7% ± 2.6% vs. 33.9% ± 3.1% at 72 hours, all P < 0.05). Conclusions UVA radiation at 9.9 J/cm2 for 7 consecutive days can decrease the intracellular degradation of extracellular elastin by HDFs, but has no obvious effect on endocytosis of elastin.