砷和铅对HepG2细胞毒性和氧化损伤的研究
Effects of arsenic and lead on cytotoxicity and oxidative stress in HepG2

为研究砷(Arsenic,As)和铅(Lead,Pb)单独以及联合作用对人肝癌细胞(Human liver carcinoma cells,HepG2)毒性作用和细胞氧化损伤的机制,采用不同质量浓度的As(2.65、3.15、3.65、4.15和4.65 μg/mL)和Pb(160、190、220、250和280 μg/mL)以及质量浓度比为1:55的As+Pb分别或联合处理HepG2细胞24、48和72 h,使用MTT法检测细胞活力;细胞培养24 h后测定细胞活性氧(ROS)、乳酸脱氢酶(LDH)、丙二醛(MDA)和还原性谷胱甘肽(GSH)的相对含量。MTT结果表明:As、Pb和As+Pb的24、48和72 h半数抑制浓度(IC50),As分别为4.20、4.03和3.74 μg/mL;Pb 分别为228.01、205.46和203.40 μg/mL;As+Pb 分别为3.10、3.17和3.18 μg/mL。3组处理分别作用HepG2细胞24、48和72 h表现出显著的生长抑制(P<0.05),且细胞毒性与时间和浓度呈依赖性。当As、Pb单独作用和As+Pb联合作用HepG2细胞诱导ROS、LDH和MDA水平显著升高(P<0.05),GSH的含量显著降低(P<0.05)。综上所述:As+Pb联合作用与单独作用表现出较强的细胞毒性,毒性大小顺序依次为:As+Pb > As > Pb,且As+Pb联合作用表现为拮抗效应。活性氧以及细胞氧化应激的产物可能是诱导重金属对细胞毒性的重要原因。
This study was aimed to evaluate the cytotoxic effects of single and combined of arsenic (As) and lead (Pb) and their joint action on human liver carcinoma cells (HepG2).The mechanism of oxidative damage on HepG2 was also investigated.The HepG2 cells were cultured by adding 2.65, 3.15, 3.65, 4.15, 4.65 μg/mL of As and 160, 190, 220, 250, 280 μg/ml of Pb, respectively;And the joint action of As+Pb was conducted at the concentration ratio of 1:55.The cell viability was assayed by MTT method after 24, 48 and 72 h of exposure.The quantitative analysis of lactate dehydrogenase (LDH), reactive oxygen species (ROS), glutathione (GSH) and malondialdehyde (MDA) was carried out to investigate oxidative damage.The results showed that:After 24, 48 and 72 h exposure of As, the IC50 values were 4.20, 4.03 and 3.74 g/mL, respectively;After 24, 48 and 72 h exposure of Pb, the IC50 values were 228.01, 205.46 and 203.40 μg/mL, respectively;After 24, 48 and 72 h exposure of the joint action of As+Pb, the IC50 values were 3.10、3.17、3.18 μg/mL, respectively;The HepG2 exposed in As, Pb and As+Pb with different concentration and time resulted in significant lower levels of cell viability and inhibition of cell growth (P<0.05);As and Pb as well as the joint action of As+Pb caused significant higher levels of LDH, ROS and MDA (P<0.05), but the GSH level showed significant lower level (P<0.05).In conclusion, the joint action of As+Pb displayed a stronger effect on cytotoxicity than that of As and Pb.The cytotoxicity order was As+Pb > As > Pb, indicating an antagonistic effect.The reactive oxygen and metabolites from oxidative stress may be an important factor for cytotoxicity.