OBJECTIVE: To investigate the effect of connexin43 (Cx43) protein on autophagy in cisplatin (DDP)-resistant testicular cancer I-10 cells. METHODS: The expression of Cx43 proteins in testicular cancer I-10 cells and I-10/DDP cells were detected with Western blotting. I-10/DDP cells were transfected with a full- length mouse Cx43 vector (mCx43) via Lipofectamine2000, the empty vector or Lipofectamine2000 (blank control group), and the changes in the expressions of LC3 and p62 proteins were determined with Western blotting. mCherry-GFP-LC3B transfection and transmission electron microscopy were used to analyze the changes in autophagy of the cells with Cx43 overexpression. RESULTS: Cx43 was significantly decreased in I-10/DDP cells compared with I-10 cells (P < 0.01). Transfection of the I-10/DDP cells with mCx43 vector resulted in significantly increased Cx43 expression in the cells (P < 0.01) and caused significantly decreased expression of LC3-Ⅱ (P < 0.01) and increased expression of p62 (P < 0.05) as compared with the negative control cells. Both transmission electron microscopy and mCherry-GFP-LC3B transfection showed that the number of autophagosomes was obviously reduced in mCx43-transfected cells as compared with the negative control cells. CONCLUSIONS: Cx43 inhibits autophagy in cisplatin-resistant testicular cancer I-10 /DDP cells.
OBJECTIVE: To investigate the effect of connexin43 (Cx43) protein on autophagy in cisplatin (DDP)-resistant testicular cancer I-10 cells. METHODS: The expression of Cx43 proteins in testicular cancer I-10 cells and I-10/DDP cells were detected with Western blotting. I-10/DDP cells were transfected with a full- length mouse Cx43 vector (mCx43) via Lipofectamine2000, the empty vector or Lipofectamine2000 (blank control group), and the changes in the expressions of LC3 and p62 proteins were determined with Western blotting. mCherry-GFP-LC3B transfection and transmission electron microscopy were used to analyze the changes in autophagy of the cells with Cx43 overexpression. RESULTS: Cx43 was significantly decreased in I-10/DDP cells compared with I-10 cells (P < 0.01). Transfection of the I-10/DDP cells with mCx43 vector resulted in significantly increased Cx43 expression in the cells (P < 0.01) and caused significantly decreased expression of LC3-Ⅱ (P < 0.01) and increased expression of p62 (P < 0.05) as compared with the negative control cells. Both transmission electron microscopy and mCherry-GFP-LC3B transfection showed that the number of autophagosomes was obviously reduced in mCx43-transfected cells as compared with the negative control cells. CONCLUSIONS: Cx43 inhibits autophagy in cisplatin-resistant testicular cancer I-10 /DDP cells.
Entities:
Keywords:
autophagy; cisplatinum; connexin43; gap junction; testis
Authors: Eloy Bejarano; Andrea Yuste; Bindi Patel; Randy F Stout; David C Spray; Ana Maria Cuervo Journal: Nat Cell Biol Date: 2014-04-06 Impact factor: 28.824