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探讨5-氟尿嘧啶(5-fluorouracil,5-FU)对人骨髓基质细胞(human bone marrow stromal cell,h BMSC)的损伤作用机制;当归多糖(angelica sinensis polysaccharides,ASP)对5-氟尿嘧啶损伤h BMSC的保护作用。采用CCK-8法测定人骨髓基质细胞株HS-5对不同浓度5-FU(0μg/mL,12.5μg/mL,25μg/mL,50μg/mL和100μg/mL)的敏感性。流式细胞术分析细胞周期;β-半乳糖苷酶染色检测衰老细胞;DCFH-DA荧光染色流式检测胞内活性氧(reactive oxygen species,ROS)水平;酶学法检测谷胱甘肽过氧化物酶(glutathione peroxidase,GSH-Px)、超氧化物歧化酶(superoxide dismutase,SOD)含量;流式细胞术分析γH2AX表达水平;ELISA检测8-羟基脱氧鸟苷(8-hydroxydeoxyguanosine,8-OHd G)的含量;5-FU 12.5~100μg/mL抑制HS-5增殖,抑制作用具有浓度依赖性和时间依赖性。5-FU组较与对照组相比细胞周期发生G1期阻滞,β-半乳糖苷酶染色阳性率升高;胞内ROS含量显著升高;细胞抗氧化能力降低;γH2AX和8-OHd G表达水平增高。ASP治疗组较5-FU组细胞相比细胞周期阻滞减少;衰老细胞比率降低;胞内ROS含量显著降低;细胞抗氧化能力升高;DNA损伤指标表达水平降低。5-FU可通过增强氧化应激诱发DNA损伤致骨髓基质细胞衰老;ASP对骨髓基质细胞损伤有保护作用,其机制可能与ASP降低损伤后骨髓基质细胞氧化应激减轻DNA损伤从而延缓骨髓基质细胞衰老有关。
To investigate the mechanism of 5-fluorouracil (5-FU) injury on human bone marrow stromal cells (hBMSCs); angelica sinensis polysaccharides (ASP) Protective effects. The sensitivity of human bone marrow stromal cell line HS-5 to different concentrations of 5-FU (0μg / mL, 12.5μg / mL, 25μg / mL, 50μg / mL and 100μg / mL) was determined by CCK- The cell cycle was analyzed by flow cytometry, the senescent cells were detected by β-galactosidase staining, the reactive oxygen species (ROS) level was detected by flow cytometry with DCFH-DA staining, the glutathione peroxidation Glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) were detected by flow cytometry. The expression of γH2AX was analyzed by flow cytometry. The levels of 8-hydroxydeoxyguanosine (8-OHdG) ). The inhibitory effect of 12.5-100μg / mL 5-FU on proliferation of HS-5 was concentration-dependent and time-dependent. Compared with the control group, the 5-FU group showed G1 arrest, the positive rate of β-galactosidase staining increased, the intracellular ROS content increased significantly, and the anti-oxidative ability of cells decreased. ΓH2AX and 8-OHd G The expression level increased. Compared with cells in 5-FU group, ASP treatment reduced the cell cycle arrest, the percentage of senescent cells decreased, the content of intracellular ROS decreased significantly, the cell antioxidant capacity increased and the expression level of DNA damage index decreased. 5-FU can induce bone marrow stromal cell senescence by inducing oxidative stress; ASP can protect bone marrow stromal cells, which may be related to the fact that ASP can reduce the oxidative stress of bone marrow stromal cells and reduce DNA damage, Related to aging.