高锶矿泉水的细胞毒性研究

Cytotoxicity study of strontium mineral water

目的比较不同浓度的高锶矿泉水对人肝细胞(LO2)、肾小管细胞(HK-2)和人脐静脉血管内皮细胞(HUVEC)增殖的影响,以了解高锶矿泉水的细胞毒性,探索对人体有益的锶矿泉水浓度及作用。方法将LO2、HK-2、HUVEC分为2、4、6、8 mg/L锶矿泉水组和对照组,先以4×104.mL-1的浓度接种于培养板中,18 h后,分别换成2、4、6、8 mg/L的锶矿泉水和双蒸水配制的DMEM高糖培养基继续培养。根据不同细胞的增殖周期,分别于1、2、3、4、5和7 d时,以CCK-8法观察细胞的增殖情况,用细胞相对增殖率按美国药典标准评价细胞毒性。用双抗体一步夹心法酶联免疫吸附试验检测培养3 d后的HUVEC培养基中血管内皮生长因子(VEGF)的分泌量。结果 2、4、6 mg/L锶矿泉水组中锶矿泉水对LO2、HK-2、HUVEC的毒性均为0～1级,8 mg/L锶矿泉水组中锶矿泉水在上述细胞增殖的早期或晚期对细胞的毒性为2～3级。LO2在4、6 mg/L锶矿泉水组中提前出现增殖峰,并较长时间地保持旺盛的增殖状态(P<0.05);HK-2在4、6、8 mg/L锶矿泉水组中生长周期延长(P<0.05),尤以4 mg/L锶矿泉水组的效果最突出(P<0.05)。HUVEC在2、4 mg/L锶矿泉水组中培养3 d后增殖速度加快(P<0.05),明显快于对照组和其他锶矿泉水组(P<0.05),而8 mg/L锶矿泉水组细胞增殖速度总体慢于对照组(P<0.05)。锶矿泉水组细胞分泌VEGF的总量均较对照组减少(P<0.05),但锶矿泉水组间差异无统计学意义(P>0.05)。结论高锶矿泉水在锶浓度不超过8 mg/L时对肝、肾和血管内皮细胞无明显毒性,对细胞增殖有益,其中锶浓度为4 mg/L的效果最突出,以延长其增殖周期为主。VEGF可能并不是高锶矿泉水促HUVEC增殖的主要作用。

Objective To investigate the effects of sttino of strontium mineral water（SMW）.Methods LO2,HK-2 and HUVEC cells were cultured in vitro with normal Dulbecco’s Modified Eagle’s Medium（DMEM） for 18 hours,and subsequently with conditioned medium containing SMW（2,4,6 and 8 mg/L） or double distilled water（DDW）.The cell proliferation was observed by cell counting kit-8（CCK-8） assay on the 1st,2nd,3rd,4th,5th and 7th days.The cytotoxicity was evaluated by cell relative growth rate according to the United States pharmacopoeia.Extracellular vein endothelial growth factors（VEGF） were assessed by enzyme-linked immunosorbent assay.Results The cytotoxicity grades of SMW of 2,4 and 6 mg/L were 0 ～ 1 on LO2,HK-2 or HUVEC cells,while they were 2 ～ 3 when SMW of 8 mg/L on cell at early or later period of proliferation.The proliferation peaks of LO2 cells came significantly earlier in SMW of 4 mg/L and 6 mg/L than in those in control group,and kept vigorous proliferation significantly longer（P 0.05）.Significantly prolonged growth cycle was observed in HK-2 cells with SMW of 4,6,or 8 mg/L（P 0.05）,which was significantly most prominent with SMW of 4 mg/L（P 0.05）.The proliferation activity of HUVEC cells in SMW of 2 and 4 mg/L was significantly increased（P 0.05） on the 3rd day;however,it was significantly inhibited with SMW of 8 mg/L（P 0.05）.The total VEGF in cells in SMW groups were significantly lower than control group（P 0.05）,though there was no significant difference among SMW groups（P 0.05）.Conclusion SMW has little cytotoxicity on LO2,HK-2 and HUVEC cells at the concentration lower 8 mg/L,even with beneficial effects on cell proliferation at certain concentration,which is prominent at 4 mg/L.VEGF may not be the major induced-proliferative factor on HUVEC.