不同剂量长波紫外线照射对HaCaT细胞增殖活力和自噬体表达瞬时影响的研究

The transient effects of different doses of ultraviolet A on the proliferation and autophagosome expression in HaCaT cells

目的:观察人永生化角质形成细胞系HaCaT细胞接受不同剂量长波紫外线(UVA)照射后细胞增殖活力和自噬体表达水平的变化,并初步评估增殖活力损伤程度和自噬体表达水平之间的相关性。方法:将HaCaT细胞分为6组,分别为10 J/cm^2UVA对照组、10 J/cm^2 UVA照射组、25 J/cm^2 UVA对照组、25 J/cm^2 UVA照射组、50 J/cm^2 UVA对照组及50 J/cm^2 UVA照射组。照射结束后即刻进行噻唑蓝(MTT)实验或单丹(磺)酰戊二胺(MDC)染色,全波长酶标仪下读取各孔A值,倒置荧光显微镜下随机选取视野,计数每视野下自噬体表达阴性细胞和阳性细胞数目。结果:经不同剂量UVA照射后,HaCaT细胞的增殖活力(A值)下降,呈剂量相关性。其中10、25及50 J/cm^2 UVA照射组(A值分别为1.179±0.007、0.791±0.015、0.522±0.046)两两之间以及与各自对照组(1.370±0.007、1.254±0.012、1.177±0.009)间差异均有统计学意义(P均<0.01);10、25及50 J/cm^2 UVA照射后,HaCa T细胞MDC染色结果示自噬体表达阳性的细胞比率增加,且呈剂量相关性。50 J/cm^2 UVA照射组自噬体表达阳性率较其对照组显著上升(χ~2=11,P<0.01)。结论:10~50 J/cm^2 UVA照射后,HaCaT细胞瞬时增殖活力降低及自噬体表达增加,且呈剂量依赖性。

Objective： To assess the changes in proliferative activity and autophagosome expression in human keratinocyte cell line（HaCaT cells） after different doses of ultraviolet A（UVA） irradiation, and to determine the relationship between the extent of proliferation impairment and the level of autophagosome expression. Methods： HaCaT cells were divided into 3 irradiation groups, each of which was treated with respective dose（10, 25 and 50J/cm^2） of UVA. The respective non-irradiated groups were served as controls. After irradiation, MTT and MDC assay were used to measure cell proliferation and autophago- some expression levels, respectively. Results： UVA irradiation significantly induced a dose-dependant reduction in the prolifer- ative activity of HaCaT cells. Absorbance（A ） values, representing proliferative activity, were significantly lower in irradiated groups than their controls（1.179±0.007 vs. 1.370±0.007 for 10 J/cm^2; 0.791±0.015 vs. 1.254±0.012 for 25 J/cm^2 and 0.522±0.046 vs. 1.177±0.009 for 50 J/cm^2; Pvalue〈0.01 for all）. Moreover, UVA irradiation induced a dose-dependant increase in autophago- some-positive cells. The percentage of autophagosome-positive cells in 50 J/cm^2 UVA irradiated group was significantly higher than that in control group （χ^2=11, P〈0.01）. Conclusions： UVA irradiation（ 10-50 J/cm^2） can decrease the proliferation activity and increase autophagosome expression in a dose-dependent manner in HaCaT cells.