Figure 4.

Effects of 3-MA and starvation on GFAP accumulation. (A) U251 cells stably expressing vector control (V), wt GFAP(WT) and mt GFAP(mt) were incubated with medium with or without 3-MA or starved for 12 h. Total cell lysates were subjected to SDS–PAGE and western-blotting analysis with antibodies against GFAP and GAPDH. (B) U251 cells were subject to 3-MA and BfaA1 for 24 h, to starvation for 12 (starved12) or 24 (starved24) hours. Cell viability was represented by methylthiazoletetrazolium (MTT) activity. Each result represents a mean ± SD of MTT activity from three independent experiments carried out in triplicate. * By student's t-test, and compared with untreated controls, P < 0.05. Control: untransfected U251 cells. (C) Morphological changes of GFAP expressing U251 cells. Cells were incubated with 3-MA or starved, then fixed and processed for GFP fluorescence. More GFAP filamentous bundles occurred in wt and dot aggregates in mt after 3-MA treatment. Starved cells are more spread out, with fewer GFAP aggregates.