Functional characterization of copper transporters zCtr1, zAtox1, zAtp7a and zAtp7b in zebrafish liver cell line ZFL

Abstract

Copper (Cu) is an essential element for all organisms, serving as an enzyme cofactor to maintain cellular activity and vitality.

Oxidative stress and apoptotic effects of copper and cadmium in the zebrafish liver cell line ZFL

“…ZFL is an adherent hepatocyte cell line [American Type Culture Collection (ATCC), CRL-2643TM] isolated from zebrafish ( Danio rerio ). The standard ZFL culture medium contained 50 % L-15 medium (Gibco, Waltham, Massachusetts, USA,11415064), 35 % Dulbecco’s Modified Eagle Medium (Gibco, Waltham, Massachusetts, USA,12100046) and 15 % Hans F12 (Gibco, Waltham, Massachusetts, USA, 21700075) supplemented with 0.15-g/L sodium bicarbonate, 15 mM HEPES (Gibco, Waltham, Massachusetts, USA,11344041), 10 % foetal bovine serum (Gibco, Waltham, Massachusetts, USA,10270106) and 1% antibiotic-antimycotic (Gibco, Waltham, Massachusetts, USA, 15240062) and was maintained at 28 °C as recommended by ATCC [ 26 , [42] , [43] , [44] ].…”

“…We used alamarBlue assay (Thermo Fisher Scientific Inc., Invitrogen Corporation, Carlsbad, CA92008, USA) to determine the mortality of cell and we followed the method as reported previously [ 26 ]. The cells were exposed to the chemicals (CuCl 2 , CdCl 2 , NAC or VE) at different concentrations for 24 h. After exposure, the alamarBlue medium was added and incubated for 2 h follow to measure fluorescence signal.…”

“…Generally, cDNA was generated by total RNA after exposure, and all the procedure was highly similar to method as reported previously [ 26 , 44 ]. In brief, all qPCR amplifications and detections were performed using Takara Premix ExTaq.…”

“…As Cu is an essential ion, a comprehensive biological system exists to balance Cu ion concentration. Notably, four proteins, namely CTR1, ATOX1, ATP7A and ATP7B, are the main regulators of Cu homeostasis [ 26 ]. CTR1 is a one way channel that imports Cu + into the cell [ 8 , 27 ].…”

“…We here used a zebrafish liver cell line, ZFL, to study combined Cu and Cd cytotoxicity. The zebrafish liver cell line ZFL has been established as a model cell line for in vitro assay and cell imaging [ 26 , [42] , [43] , [44] ]. We examined the responses of zebrafish liver cell line (ZFL) to oxidative stress and apoptotic effects created by the administrations of Cu 2+ and Cd 2+ .…”