AMP-activated protein kinase (AMPK) is a well-studied molecule for the control of cellular metabolism. In addition, AMPK can affect autophagy to sustain homeostasis. Adult hippocampal neural stem (HCN) cells undergo autophagic cell death (ACD) without apoptotic features following insulin withdrawal, therefore, insulin-deprived HCN cells are regarded as a genuine model for studying autophagy and cell death-related signaling mechanisms. In this study, we investigated the role of AMPK on autophagy regulation in HCN cells following insulin-withdrawal. Genetic and pharmacological activation of AMPK promoted ACD, while inhibition of AMPK yielded the opposite outcomes. These data suggest that AMPK can be a positive regulator of ACD in HCN cells following insulin withdrawal. ⓒ 2016 DGIST
Table Of Contents
1. Introduction 1 -- 2. Materials and methods 5 -- 2.1 Cell culture 5 -- 2.2 Antibodies and reagents 5 -- 2.3 Cell death assay 5 -- 2.4 GFP-LC3 puncta assay 6 -- 2.5 Western blot 6 -- 2.6 Transfection 6 -- 2.7 Sh-RNA Lentiviral system 7 -- 2.8 Statistical analysis 7 -- 3. Results 8 -- 3.1 AMPK activator, A769662 increased cell death following insulin withdrawal. 8 -- 3.2 Ectopic expression of AMPK constructs modulated cell death. 8 -- 3.3 Silencing of AMPK reduced ACD in insulin-deprived HCN cells. 9 -- 3.4 Cell death mode induced by genetic and pharmacological activation of AMPK was ACD, not apoptosis. 9 -- 3.5 Suppression of autophagy related gene, Atg7 prevented ACD. 10 -- 4. Discussion 11 -- 5. Figure legends 13 -- 6. Figures 16 -- Reference 24 -- Abstract in Korean 26--