Figure 6. GLS2 mediates p53’s function in inhibiting migration, invasion and lung metastasis of HCC cells.
(A) Knockdown of endogenous WT p53 reduced GLS2 expression in Huh-1 and HepG2 cells as measured by western-blot (left) and Taqman real-time polymerase chain reaction assays (right), respectively. (B, C) Knockdown of p53 promoted the migration (B) and invasion (C) of Huh-1 and HepG2 cells measured by trans-well assays. (D, E) Simultaneous knockdown of GLS2 and p53 by shRNA vectors in Huh-1 and HepG2 cells did not display an addictive promoting effect on the migration (D) and invasion (E) of cells. In A–E, data represent mean ± SD (n=6). **p<0.001; Student’s t-test. (F, G) Simultaneous knockdown of GLS2 and p53 in Huh-1 and HepG2 cells did not display an addictive promoting effect on lung metastasis in vivo. Huh-1 and HepG2 cells with individual knockdown of GLS2 or p53, or simultaneous knockdown of GLS2 and p53 were used for assays. In F, lung metastasis was analyzed by in vivo bioluminescence imaging at 7 weeks after inoculation of cells. Upper panels: representative images of lung metastasis of Huh-1 cells analyzed by in vivo imaging. Lower panels: quantification of lung photon flux. In G, lung metastasis was analyzed by histological analysis at week 7. Left panels: hematoxylin and eosin staining of lung metastasis of Huh-1 cells. Scale bars: 200 μm. Right panels: The average number of tumors/lung. Data represent mean ± SD (n=10 mice/group). **p<0.001; Student’s t-test. GLS, glutaminase; HCC, hepatocellular carcinoma; shRNA, short hairpin RNA.
Figure 6—figure supplement 1. The viability and number of HCC cells with p53 knockdown cultured in serum-free medium for 36 hr.
