|
|
|
| Acute toxicity determination of compounds by HepG2 cell neutral red staining |
| XU Jian*, TONG Xiao-ling, CEN Jiang-jie, JI Lei, LIN Shi-dao, ZHU Xin-qiang
|
| *School of Medicine,Zhejiang University,Hangzhou,Zhejiang 310058,China |
|
|
|
|
Abstract Objective To explore the feasibility of detecting compound toxicity in HepG2 cell neutral red staining method and to provide the basis for the establishment of a rapid screening method. Methods The cytotoxic IC50,IC45,IC40,IC35 and IC30 values of 30 kinds of compounds,including colchicine,cyclophosphamide pipion and so on,were detected by the neutral red staining method of human liver cancer HepG2 cells. The regression equation of the ICx value was used to predict the LD50 value of the rat via the linear regression method,and the LD50 was predicted by the regression equation with the highest correlation. The toxicity of the compound was graded by the actual value and predicted value of LD50,and the consistency of the two methods was compared. Results The correlation between lg（IC50）,lg（IC45）,lg（IC40）,lg （IC35）,lg（IC30）and lg（LD50）was statistically significant（P<0.01）,and correlation coefficient（r）was 0.779-0.815. The correlation between lg（IC50）and lg（LD50） is the highest,lg（LD50）=0.887×lg（IC50） + 0.414. The actual value of LD50 divided 30 compounds into 10 kinds of high toxicity,medium toxicity and low toxicity,and the predicted value of LD50 was divided into 12 types of high toxicity,10 medium toxicity and 8 low toxicity. There was no significant difference between the two methods（P>0.05）,and the concordant rate was 86.7%. Conclusion The cell neutral red staining method can be used as a method for rapid detection of acute toxicity.
|
|
Received: 02 January 2018
Revised: 09 February 2018
Published: 22 May 2018
|
|
|
|
|
|
[1] 杨海智,马海霞,杨信东. 国内关于半数致死量及类似生物效应指标测算方法研究进展[J]. 国外医药抗生素分册,2012,33(2):62-66.
[2] 施畅,马华智,王全军,等. 化学物(药物)毒性测试替代体系的建立及应用[J]. 中国比较医学杂志,2017,27(5):6-8.
[3] 曾丽海. 急性毒性体内及体外替代方法研究进展[J]. 中国公共卫生,2011,27(10):1331-1333.
[4] 许羚,胡,丁晓霜,等. 雷公藤红素体内与体外急性毒性试验结果的比较[J]. 环境与职业医学,2015,32(6):535-538.
[5] 中华人民共和国国家质量监督检验检疫总局.医疗器械生物学评价第5部分:体外细胞毒性试验:GB/T16886.5—2003[S]. 北京:中国标准出版社,2003.
[6] 秦丹丹,高南南,赵秀云,等. 相思子蛋白P2抗肝癌作用及对端粒酶活性影响[J]. 中国药理学通报,2011,27(12):1666-1671.
[7] FOTAKIS G,TIMBRELL J A.In vitro cytotoxicity assays:comparison of LDH,neutral red,MTT and protein assay in hepatoma cell lines following exposure to cadmium chloride[J]. Toxicol Lett,2006,160(2):171-177.
[8] 中华人民共和国农业部. 农药登记资料规定[Z].2007-12-08.
[9] Background Review Document (BRD):Validation of Neutral Red Uptake Test Methods .NIH,2006. Publication No.07-4518[EB/OL][2018-01-07]. http:// iccvam.niehs.nih.gov/ methods/ acutetox/inv_nru_brd.htm.
[10] 陈素珍,李瑾翡,曾秋敏. 8种烈性中药的体外急性毒性试验方法初探[J]. 医药前沿,2013,22(8):44-45.
[11] 李雪,牛勇,贾强,等. 四种细胞毒性试验测定柴油机尾气颗粒物提取物急性毒性的比较[J]. 卫生研究,2013,42(2):221-227.
[12] 袁海涛. 毒性测试替代方法在化学物心血管系统安全性评价中的应用[D]. 长春:吉林大学,2014.
[13] HOFFMANN S,KINSNER-OVASKAINEN A,PRIETO P,et al.Acute oral toxicity:variability,reliability,relevance and interspecies comparison of rodent LD50 data from literature surveyed for the ACuteTox project[J]. Regul Toxicol Pharmacol,2010,58(3):395-407.
[14] GUSTAFSSON H,RUNESSON J,LUNDQVIST J,et al.Neurofunctional endpoints assessed in human neuroblastoma SH-SY5Y cells for estimation of acute systemic toxicity[J]. Toxicol Appl Pharmacol,2010,245(2):191-202.
[15] KINSNER-OVASKAINEN A,PRIETO P,STANZEL S,et al.Selection of test methods to be included in a testing strategy to predict acute oral toxicity:an approach based on statistical analysis of data collected in phase 1 of the ACuteTox project[J]. Toxicol In Vitro,2013,27(4):1377-1394.
[16] CLOTHIER R,GÓMEZ-LECHÓN M J,KINSNER-OVASKAINEN A,et al. Comparative analysis of eight cytotoxicity assays evaluated within the ACuteTox Project[J]. Toxicol In Vitro,2013,27(4):1347-1356.
[17] BHHATARAI B,WILSON D M,BARTELS M J,et al.Acute toxicity prediction in multiple species by leveraging mechanistic toxcast mitochondrial inhibition data and simulation of oral bioavailability[J]. Toxicol Sci,2015,147(2):386-396. |
|
|
|
