Fig. 2.

HepaRG cell response to 4 and 16 hour Mn exposure. HepaRG cells were exposed to Mn for 4 and 16 hours in standard culture conditions followed by assessment of cell viability or ⁵⁴Mn uptake. a. HepaRG cells remain viable following exposure to Mn (5, 25, 50, 100, 1000μM) for 4 hours (open circles) and 16 hours (open squares). Cell viability was determined using MTT and normalized to control (untreated) cells. Shown are means ± SEM, n = 3. b. HepaRG cells accumulate Mn with increasing Mn concentration. Cells were exposed to 50, 100, and 500μM Mn for 4 or 16 hours. Cells were washed with cold PBS, removed from the wells by trypsinization, and digested for ICP-MS analysis. Mn levels increased in the cells with increasing dose at both the 4 and 16 hour incubation periods. Values are means ± SEM, n=3 biological samples. a, significant from 50μM; b, significant from 100μM. c. Shown is a diagram depicting the protocol used for the experimental results shown in panel C. HepaRG cells were pre-incubated with MnCl₂ for 4 or 16 hours prior to the ⁵⁴Mn uptake assay. HepaRG cells were then rinsed and incubated with 1μM ⁵⁴MnCl₂ for 30 minutes at 37°C. Cells were rinsed, removed from the plate, and cell ⁵⁴Mn content determined using a gamma counter. d. Values for the ⁵⁴Mn uptake experiment are shown as mean ± SEM, n=4 biological; a, significant from control; b, significant from 5μM, c, significant from 50μM. P < 0.05.