Colistin-induced autophagy and apoptosis involves the JNK-Bcl2-Bax signaling pathway and JNK-p53-ROS positive feedback loop in PC-12 cells

Chem Biol Interact. 2017 Nov 1:277:62-73. doi: 10.1016/j.cbi.2017.08.011. Epub 2017 Aug 24.

Abstract

Our recent study demonstrated neurotoxicity of colistin-induced autophagy and apoptosis in PC-12 cells, and that autophagy reached peak level at 12 h. In this study, we scrutinized the role of JNK in colistin-induced neurotoxicity and demonstrated the relationship among JNK, p53 and ROS in colistin treated PC-12 cells. Colistin-induced autophagy and apoptosis by JNK inhibition/activation were examined by western blotting, electron microscopy, and immunofluorescence/fluorescence microscopy. The results indicated that colistin induced JNK activation reached peak level at 12 h, while the highest levels of p-Bcl2/Bcl2 were observed at 12 h and Bax/Bcl2 significantly increased in a time-dependent manner. In PC-12 cells, inhibition of JNK by SP600125 (JNK inhibitor) resulted in significantly lower levels of autophagy upon colistin treatment, depending on the expression levels of Beclin1, LC3-II, p62 degradation and reduction in the number of autophagic vacuoles. In contrast, anisomycin pretreatment PC-12 cells led to upregulated autophagy. Especially, the highest levels of Beclin1 and p-Bcl2/Bcl2 were observed at 6 h, and Bax/Bcl2, cleaved-caspase3 and cleaved-PARP significantly increased in a time-dependent manner. The results revealed that JNK activation mediated autophagy and apoptosis related to Beclin1-Bcl2 and Bax-Bcl2 complex in colistin-treated PC-12 cells. Silencing of p53 by siRNA before colistin treatment substantially reduced ROS production and transactivated JNK in PC-12 cells. Moreover, activation of JNK increased ROS generation in PC-12 cells. In conclusion, colistin-induced autophagy and apoptosis is correlated to JNK-Bcl2-Bax signaling pathway, and an interaction effect found between intracellular ROS level and JNK-p53 signaling pathway in apoptosis.

Keywords: Apoptosis; Autophagy; JNK; PC-12 cells; ROS.

MeSH terms

  • Animals
  • Anti-Bacterial Agents / toxicity*
  • Apoptosis / drug effects*
  • Autophagy / drug effects*
  • Colistin / toxicity*
  • JNK Mitogen-Activated Protein Kinases / metabolism
  • MAP Kinase Signaling System / drug effects
  • Neurons / cytology
  • Neurons / drug effects*
  • Neurons / metabolism
  • PC12 Cells
  • Proto-Oncogene Proteins c-bcl-2 / metabolism
  • Rats
  • Reactive Oxygen Species / metabolism
  • Signal Transduction / drug effects*
  • Tumor Suppressor Protein p53 / metabolism
  • bcl-2-Associated X Protein / metabolism

Substances

  • Anti-Bacterial Agents
  • Proto-Oncogene Proteins c-bcl-2
  • Reactive Oxygen Species
  • Tumor Suppressor Protein p53
  • bcl-2-Associated X Protein
  • JNK Mitogen-Activated Protein Kinases
  • Colistin