In vitro cytotoxicity assessment of two heavy metal salts in a fish cell line (RF)

Abstract

Labeo rohita (rohu) is one of most important of Indian major carps, which is highly ranked among the important freshwater aquaculture species in the world. Heavy metals are major risk factors for aquatic health, which are biomagnified through the food chain. The present study was carried out to investigate the effect of different divalent salts of two heavy metals, such as zinc (ZnCl₂, ZnSO₄ and ZnNO₃) and cadmium (CdCl₂ and CdSO₄), in an established fish cell line, RF developed from fin tissue of L. rohita. The RF cell line was used for assessment of heavy metal cytotoxicity through various endpoint assays, including maximum tolerated dose (MTD) determination, 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) assay, neutral red (NR) uptake assay, and Coomassie brilliant blue (CBB) assay. Results revealed that these heavy metal salts were cytotoxic to the RF cell line at varied concentrations. MTD values were found to be 1.563, 3.125, 6.25, 12.5 and 25 mg/L for CdCl₂, CdSO₄, ZnCl₂, Zn(NO₃)₂ and ZnSO₄, respectively. The half-maximal inhibitory concentration values calculated by MTT, NR and CB assay were 53.83 ± 7.02, 58.03 ± 9.12 and 79.20 ± 15.27 for ZnSO₄, 26.44 ± 7.01, 36.60 ± 7.82 and 155.6 ± 14.75 for Zn(NO₃)₂, 20.26 ± 17.95, 16.94 ± 7.05 and 87.54 ± 7.58 for ZnCl₂, 5.166 ± 0.57, 15 ± 1 and 41.80 ± 8.38 for CdSO₄ and 4.966 ± 0.56, 9.56 ± 1.73 and 20.93 ± 4.47 for CdCl₂. This study establishes the RF cell line as an in vitro tool for assessment and monitoring of heavy metal concentration in the aquatic environment.

• Cadmium salts
• CB assay
• in vitro model
• MTT assay
• NR assay
• zinc salts

Acknowledgements

The authors thank the Director, NBFGR, Lucknow, for his support and encouragement for the research program.