螯合剂BPCBG对铀的促排效果和防护铀致人肾小管上皮细胞损伤的作用

Effect of the chelator BPCBG on the decorporation of uranium in vivo and uranium-induced damage of human renal tubular epithelial cells in vitro

从动物和细胞水平观察螯合剂N,N'-1,2-亚乙基双[N-(2,3-二羟基苯甲基)]甘氨酸(BPCBG)对铀的促排效果,以及对铀致人肾近曲小管上皮细胞(HK-2)损伤的保护作用。大鼠腹腔注射(ip)醋酸铀酰后立即肌内注射(im)不同剂量的BPCBG,采用ICP-MS方法测定24 h的尿铀排出量和肾、骨中铀蓄积量。人肾近曲小管上皮HK-2细胞染铀24 h后给予不同剂量的BPCBG作用24 h,采用ICP-MS方法检测细胞内铀含量。不同剂量醋酸铀酰染毒HK-2细胞后立即或延迟24 h给予BPCBG作用24或48 h,采用CCK-8试剂盒检测细胞存活率,采用CB微核法观察BPCBG对铀致染色体损伤的影响,采用DCFH-DA荧光探针法检测BPCBG对铀诱导细胞内氧自由基生成的影响。以上实验均以DTPA-CaNa3作对照。结果表明,BPCBG(60、120和600μmol.kg-1)使24 h尿铀排出量较铀中毒对照组明显增加约37%～61%,肾、骨中铀蓄积量降低至中毒对照组的41%～31%和86%～42%,促排效果随给药剂量增加而明显增强。HK-2细胞铀染毒后延迟24 h给予10～250μmol.L-1 BPCBG作用24 h,使细胞内铀含量较铀染毒组显著降低约55%～60%,并能明显提高铀染毒HK-2细胞的存活率,显著降低铀诱导的微核形成,有效抑制铀诱导的细胞内氧自由基的产生。DTPA-CaNa3虽能明显降低大鼠肾铀蓄积量和细胞内铀含量,但促排效果显著低于BPCBG,且对铀致细胞损伤无保护作用。以上动物和细胞实验均证明BPCBG是有效的铀促排剂,明显优于DTPA-CaNa3,并具有清除铀诱导细胞内氧自由基产生的作用,可以保护铀致肾细胞损伤,值得进一步研究。

This study is to assess the efficacy of BPCBG on the decorporation of uranium（Ⅵ） and protecting human renal proximal tubular epithelial cells（HK-2） against uranium-induced damage.BPCBG at different doses was injected intramuscularly to male SD rats immediately after a single intraperitoneal injection of UO2（CH3COO）2.Twenty-four hours later uranium contents in urine,kidneys and femurs were measured by ICP-MS.After HK-2 cells were exposed to UO2（CH3COO）2 immediately or for 24 h followed by BPCBG treatment at different doses for another 24 or 48 h,the uranium contents in HK-2 cells were measured by ICP-MS,the cell survival was assayed by cell counting kit-8 assay,formation of micronuclei was determined by the cytokinesis-block（CB） micronucleus assay and the production of intracellular reactive oxygen species（ROS） was detected by 2＇,7＇-dichlorofluorescin diacetate（DCFH-DA） oxidation.DTPA-CaNa3 was used as control.It was found that BPCBG at dosages of 60,120,and 600 μmol.kg？1 resulted in 37%-61% increase in 24 h-urinary uranium excretion,and significantly decreased the amount of uranium retention in kidney and bone to 41%？31% and 86%？42% of uranium-treated group,respectively.After HK-2 cells that had been pre-treated with UO2（CH3COO）2 for 24 h were treated with the chelators for another 24 h,55%？60% of the intracellular uranium was removed by 10？250 μmol.L-1 of BPCBG.Treatment of uranium-treated HK-2 cells with BPCBG significantly enhanced the cell survival,decreased the formation of micronuclei and inhibited the production of intracellular ROS.Although DTPA-CaNa3 markedly reduced the uranium retention in kidney of rats and HK-2 cells,its efficacy of uranium removal from body was significantly lower than that of BPCBG,and it could not protect uranium-induced cell damage.It can be concluded that BPCBG effectively decorporated the uranium from UO2（CH3COO）2-treated rats and HK-2 cells,which was better than DTPA-CaNa3.It could also scavenge the uranium-induced intracellular ROS and protect against the uranium-induced cell damage.BPCBG is worth further investigation.