迷迭香酸对X射线诱导微血管内皮细胞损伤的保护作用

Protective Effect of Rosmarinic Acid on Human Dermal Microvascular Endothelial Cell Injury Induced by X-ray Radiation

肿瘤放射治疗所致血管损伤是其严重不良反应根源之一,目前未有理想防治措施。本研究采用MTT比色法、Hoechst33258染色后形态学观察、Annexin V/PI双染后流式细胞仪分析等方法,考察了天然抗氧化剂迷迭香酸(Ros-marinic acid,RA)对X射线所致HMEC-1人真皮微血管内皮细胞增殖抑制以及凋亡的影响;另外采用相关试剂盒测定细胞内总抗氧化力(T-AOC)、还原型谷胱甘肽/氧化型谷胱甘肽比值(GSH/GSSG)水平,并以细胞内活性氧(iROS)荧光探针(DCFH-DA)检测了细胞内活性氧水平改变。结果表明,6.0Gy X射线照射可抑制HMEC-1细胞增殖达53.7%,凋亡率增至55.8%,细胞内T-AOC、GSH/GSSG水平均明显降低,iROS水平明显升高,各指标与正常对照组比较均有极显著性差异(P<0.05)。采用RA(5,10,20μg/mL)预处理细胞1h,可浓度依赖性地减轻X射线诱导的HMEC-1增殖抑制和凋亡程度(P<0.05),改善细胞内氧化胁迫状态。由此可知,RA对X射线照射所致HMEC-1细胞损伤具有明显保护作用,可能与其提高细胞内抗氧化能力、清除活性氧以改善氧化胁迫有关。

Microvascular injury is the major cause of the late normal tissue reaction after cancer radiotherapy.We investigated a natural antioxidant rosmarinic acid（RA） against the radiation injury induced by X-rays in human dermal microvascular endothelial cell line（HMEC-1） in vitro.The proliferation inhibition and apoptosis of HMEC-1 cells were measured by MTT colorimetry,Hoechst 33342 staining,and flow cytometric analysis after double staining with annexin-V and propidium（PI）.Furthermore,the intracellular total-anti-oxidizing-capability（T-AOC） and the ratio of reduced glutathione（GSH） to oxidized glutathione（GSSG）（GSH/GSSG） were assayed by commercial kits.And the intracellular reactive oxygen species（iROS） was detected by the fluorescent probe 2,7-dichlorodihydrofluorescein diacetate（DCFH-DA）.The results indicated that X-rays irradiation（6.0 Gy） induced significantly the proliferation inhibition and apoptosis in HMEC-1 cells（P0.01 vs control group）.X-rays irradiation decreased the T-AOC and GSH/GSSG levels（P0.01 vs control group） while elevated the iROS level（P0.01 vs control group） in HMEC-1 cells.Compared with X-rays irradiated group,the RA（5,10,20 μg/mL） pretreatment for 1 h obviously prevented HMEC-1 cells injuries from exposure to X-rays（P0.05 vs X-rays irradiated group）.Conclusion：RA possesses the protective ability in HMEC-1 cells against radiation injuries,which might be related to its modulation of intracellular oxidative stress status.