NOX4 knockout inhibits cell proliferation.
(A) Proliferation of three NOX4 knockout cell lines was quantitated by MTT assay. HeLa control and knockout clones were seeded into 24 well plate and the MTT assay were performed in quadruplicate. The graph shows the average and the standard deviation (SD). Control vs knockout cells. *: P <0.005, **: P <0.00005. (B) NOX4 knockout HeLa cells were analyzed with flow cytometry. The percentage of cells at sub-G1 and S/G2/M was measured, and is shown in the graph. Experiment was repeated three times with similar observations, a representative data is shown. (C) NOX4 knockout may affect activation of Akt phosphorylation. NOX4 depleted cells showed significantly lower levels of phospho-Akt in comparison to parental normal cell. However, NOX4 knockout did not affect phosphorylation of S6 Ribosomal Protein. Cells were serum starved by incubation in DMEM containing 1% FBS for 10 hrs, then exposed to EGF (20ng/ml) for 20 min before collection of cells for analysis. Experiments were repeated three times with similar observations, and representative results are shown.