Megakaryocytic Differentiation of K562 Cells Induced by PMA Reduced the Activity of Respiratory Chain Complex IV
Figure 5
PMA induced K562 cells differentiation reduced the activity of respiratory chain complex IV.
(A) K562 cells were incubated with 5 nM PMA and harvested at 12 h, 24 h and 72 h respectively. Isolated mitochondria from these cells were solubilised with 1% dodecyl-maltoside (DM) before subjecting the samples to hrCN-PAGE. After electrophoresis, gels were incubated with in-gel catalytic activity assay buffers. Each lane was loaded with 100 µg protein. (B) K562 cells were incubated with 5 nM PMA and harvested at 12 h, 24 h and 72 h respectively. The activity of the complex IV activity was detected by spectrophotometric measurement method. Data were shown as mean ± SD of three independent experiments, *p≤0.05, **p≤0.001. (C) K562 cells were incubated with 5 nM PMA for the indicated time. Mitochondrial extracts were prepared and subjected to SDS-PAGE followed by immunoblotting with antibodies indicated. (D) Mitochondrial extracts were prepared and subjected to Blue Native gel subsequently processed by immunoblotting to analyze the levels of the four subunits (COX1, COX3, COX5A and COX6B1) of complex IV. The arrows represented the complex of interest. Blots were representative of three separate experiments.