Effects of ginsenoside Rg2 on the ultraviolet B-induced DNA damage responses in HaCaT cells
Effects of ginsenoside Rg2 on the ultraviolet B-induced DNA damage responses in HaCaT cells
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DOI：
10.1007/s00210-010-0522-9
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发表时间：
2010-07-01
影响因子：
3.6
通讯作者：
Park, Jong Kun
中科院分区：
文献类型：
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作者：
Ha, Se Eun;Shin, Dae Hyun;Park, Jong Kun
我们先前的研究证明了mRg2对UVB损伤的修复作用，mRg2是一种含有60%Rg2的人参皂苷混合物，在NIH3T3细胞中具有促进修复作用。本研究从基因表达水平研究了纯化的人参皂苷Rg2对中波紫外线(UVB)照射的HaCaT细胞修复和凋亡的影响。当细胞暴露于UVB并在正常培养液中孵育24小时后，细胞存活率下降到未处理对照组的50%左右。然而，当Rg2被孵育后，UVB诱导的细胞毒性以浓度和时间依赖的方式被显著阻止。Rg2孵育后对UVB暴露所致的细胞凋亡核碎裂也有明显的保护作用。基因芯片分析表明，Rg2单独作用后的基因包括与P53信号通路有关的基因，如GADD45α、GADD45β和细胞通讯基因。RT-PCR分析显示，与未处理的对照组相比，Rg2单独处理组P53和GADD45的转录和蛋白水平略有上调，约为1.5倍。与正常对照组相比，UVB照射后Rg2作用24 h后，细胞中P53和GADD45的mRNA表达水平下降，且呈浓度依赖性。而经5 mU视黄醇孵育后的UVB暴露细胞的mRNA水平与正常培养液中孵育后的基本相同。时程实验显示，50mU的Rg2作用于UVB暴露的细胞6h和9h后，P53和GADD45的mRNA表达水平分别上调，然后逐渐下降，直至24h，Western印迹分析还表明，Rg2作用后UVB暴露的细胞中P53、磷酸化P53、GADD45和ATM的表达均下降。时程分析还表明，这些表达的降低是由于P53和GADD45蛋白较早上调所致。UVB照射后，Rg2与细胞孵育24 h后，残留的环丁烷嘧啶二聚体水平随Rg2浓度和时间的增加而降低。所有这些结果表明，Rg2通过促进DNA修复来保护细胞免受UVB诱导的遗传毒性，这可能与调节参与P53信号通路的蛋白质水平有关。
Our previous study demonstrated the increase in the repair of UVB damage by mRg2, a mixture of ginsenosides containing 60% Rg2 in NIH3T3 cells. In the present study, the effects of purified Rg2 on the repair and apoptosis in ultraviolet B (UVB)-exposed HaCaT cells were investigated on gene expression levels. When cells were exposed to UVB and post-incubated in normal medium for 24 h, the cell viability decreased to about 50% of that in nontreated control. When Rg2 was post-incubated, however, the UVB-induced cytotoxicity was significantly prevented in an Rg2 concentration- and time-dependent manner. The apoptotic nuclear fragmentation resulting from UVB exposure was also significantly protected by the Rg2 post-incubation. Microarray analysis showed that the genes stimulated by the Rg2-alone treatment include those involved in p53 signaling pathway such as GADD45 alpha, GADD45 beta, and cell communication genes. RT-PCR analysis showed that the Rg2-alone treatment slightly upregulated the p53 and GADD45 transcript and protein levels by about 1.5-fold as compared with the nontreated control. The mRNA levels of p53 and GADD45 in cells exposed to UVB and post-incubated with Rg2 for 24 h decreased in an Rg2 concentration-dependent manner as compared with that post-incubated in normal medium. However, the mRNA level of the UVB-exposed cells post-incubated with 5 mu M retinol was essentially the same as that post-incubated in normal medium. Time course experiment showed that the mRNA levels of p53 and GADD45 in UVB-exposed cells were upregulated by post-incubation with 50 mu M Rg2 until 6 and 9 h, respectively, and then gradually decreased until 24 h. By Western blot analysis, it was also revealed that the Rg2 post-incubation decreases the expression of p53, phospho-p53, GADD45, and ATM in UVB-exposed cells. Time course analysis also indicated that these decreased expressions were due to the earlier upregulation of p53 and GADD45 proteins. When UVB-exposed cells were post-incubated with Rg2 for 24 h after UVB exposure, the level of remaining cyclobutane pyrimidine dimers decreased in both Rg2 concentration- and time-dependent manner. All these results suggest that Rg2 protects cells against UVB-induced genotoxicity by increasing DNA repair, in possible association with modulation of protein levels involved in p53 signaling pathway.