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. Author manuscript; available in PMC: 2022 Aug 12.
Published in final edited form as: Cell Signal. 2020 Apr 23;72:109651. doi: 10.1016/j.cellsig.2020.109651

Figure 3.

Figure 3.

Inhibition of c-Src increases OXPHOS expression in the Hep3B cell line. A) Hep3B cells were treated with 0 – 5 μM Src family kinase (SFK) inhibitor PP2 for 72 h before collection. Changes in the expression of OXPHOS complex subunits, c-Src, pSFK, Tyr phosphorylation (pTyr), EF-Tu, and TFAM were measured by Western blot analyses with respect to GAPDH antibody probing and Ponceau S staining of the membranes as equal protein loading controls. The phosphorylation of SFK members (pSFK) was detected by the pTyr antibody, indicated by an arrow, as well as the pSFK antibody that detects phosphorylation at Tyr416. The quantitative analyses of the expression of OXPHOS subunits are represented as mean ± SD of at least three experiments for each group (bar graph, right panel). Significant differences were observed in the expression of COII (P = 0.0091) and NDUFB8 (P = 0.024) at 5 μM PP2. B) Cell proliferation of PP2 treated Hep3B cells were measured with the Trypan blue exclusion assays after 72 h treatment. Statistical difference was shown between the control and 2.5 μM (P = 0.001) and 5 μM (P = 0.0001) PP2. Complex I and III and complex IV activities were measured by determining the rates of cytochrome c reduction and oxidation, respectively, at 550 nm. Significant differences were observed in the complex I and III activity at 2.5 μM (P = 0.0122) and 5 μM (P = 0.0006) PP2 and complex IV activity at 2.5 μM (P = 0.0002) and 5 μM (P = 0.0001) PP2. C) Western blot analyses were used to identify changes in Hep3B cells transfected with with c-Src siRNA (c-Src) relative to control siRNA (cont). The quantitative analyses of OXPHOS expression are represented as the mean ± SD of at least three experiments for each group (bar graph, right panel). Significant differences were observed in the expression of COII (P = 0.0397) and NDUFB8 (P = 0.0022) between the control and c-Src siRNA transfected Hep3B cells. Data represents the mean ± SD of at least three independent experiments. The results are presented as a percentage of the control cells (cont = 100%). Statistical difference was observed between the control and treatment groups via unpaired Student’s t-test (2-tailed), *P < 0.05.