Nickel, cobalt and chromium-induced cytotoxicity and intracellular accumulation in human hacat keratinocytes.
Nickel, cobalt and chromium can induce allergic contact dermatitis (ACD) and may provoke irritant reactions in the skin. This study aimed at investigating cytotoxicity and cell viability along with intracellular metal accumulation in HaCaT human keratinocytes exposed to soluble forms of nickel, cobalt or chromium. The EC50 (24 h) values as detected by MTT test were 30 microM for sodium chromate (Na2CrO4), 475 microM for cobalt chloride (CoCl2) and 600 microM for nickel chloride (NiCl2). Chromium chloride (CrCl3) was not toxic up to 1 mM. No clear effects were observed after 4 h, but 24-h treatments with 1 mM CoCl2 or 10 microM Na2CrO(4) were found to almost completely abolish the ability of the cells to form colonies, whilst 1 mM NiCl2 reduced cellular survival to only 70% of control cultures. Intracellular accumulation of metals was evaluated by the use of radioisotopes at the EC50 value and at 1/10-1/5 of this concentration. Accumulation of Na2(51)CrO4 was linear with increasing dose. This was not the case for 63NiCl2 and 58CoCl2. All the metals were accumulated preferentially in the cytosols; 96% or more for 63NiCl2, approximately 90% for 58CoCl2 and 60-70% for Na2(51)CrO4. Finally, it was observed that HaCaT human keratinocytes can concentrate the metals present in the media up to 3.9 and 12.5 times for NiCl2 and CoCl2, respectively, and up to 167 for Na2CrO4. These striking metal intracellular accumulation patterns, which have not been earlier described in keratinocytes, highlight the relevance of searching for specific biomarkers of early cellular toxic effects, such as cytosolic proteins that bind the metals.
Environmental Impact Unit, TP 460, Environment Institute, DG Joint Research Center, I-21020 VA, Ispra, Italy.
M
M
Ermolli
C
C
Menné
G
G
Pozzi
M A
MA
Serra
L A
LA
Clerici
D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002857:Chromium / Q000378:metabolism* / Q000633:toxicity*; D003035:Cobalt / Q000378:metabolism* / Q000633:toxicity*; D003114:Colony-Forming Units Assay; D006801:Humans; D007202:Indicators and Reagents; D015603:Keratinocytes / Q000187:drug effects* / Q000378:metabolism*; D009532:Nickel / Q000378:metabolism* / Q000633:toxicity*; D009580:Nitroblue Tetrazolium; D013347:Subcellular Fractions / Q000187:drug effects / Q000378:metabolism
D007202:Indicators and Reagents; D002857:Chromium; D009580:Nitroblue Tetrazolium; D003035:Cobalt; D009532:Nickel