title: AMP-activated protein kinase (AMPK)/Ulk1-dependent autophagic pathway contributes to C6 ceramide-induced cytotoxic effects in cultured colorectal cancer HT-29 cells.
abstract: Colorectal cancer is the second leading cause of cancer-related deaths. Drug resistance and/or off-target toxicity against normal cells limit the effectiveness of current chemotherapies for the treatment of colorectal cancer. In the current study, we studied the potential cytotoxic effects of short-chain and cell-permeable C6 ceramide in cultured colorectal cancer HT-29 cells and focused on the underlying mechanisms. We observed that C6 ceramide-induced HT-29 cell death and growth inhibition in a dose- and time-dependent manner. However, no significant apoptosis was observed in C6 ceramide-treated HT-29 cells. Our data support that autophagy contributed to C6 ceramide-induced cytotoxic effects, as autophagy inhibitors, 3-methyladenine (3-MA) and hydroxychloroquine, inhibited C6 ceramide's effect; however, autophagy activators, everolimus (RAD001) and temsirolimus, mimicked C6 ceramide effects and induced HT-29 cell death. Further, we indentified that AMP-activated protein kinase (AMPK)/Ulk1 signaling was required for autophagy induction by C6 ceramide, and AMPK silencing by a specific short hairpin RNA suppressed C6 ceramide-induced autophagy and cytotoxic effects. Reversely, forced activation of AMPK by its activator AICAR or by genetic manipulation caused autophagic death in HT-29 cells, which was inhibited by 3-MA. Our results suggest that autophagy, but not apoptosis, is a major contributor for C6 ceramide-induced cytotoxic effects in HT-29 cells, and activation of AMPK/Ulk1 is required for the process.
authors.affiliation: Department of General Surgery, The Ninth People's Hospital Affiliated to Shanghai Jiao-tong University School of Medicine, Shanghai, China.
authors.forename: Hai-zhong
authors.initials: HZ
authors.lastname: Huo
authors.forename: Bing
authors.initials: B
authors.lastname: Wang
authors.forename: Jian
authors.initials: J
authors.lastname: Qin
authors.forename: Shan-yu
authors.initials: SY
authors.lastname: Guo
authors.forename: Wen-yong
authors.initials: WY
authors.lastname: Liu
authors.forename: Yan
authors.initials: Y
authors.lastname: Gu
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000502:physiology*; D000620:Aminoimidazole Carboxamide / Q000031:analogs & derivatives / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis; D001343:Autophagy / Q000187:drug effects*; D000071189:Autophagy-Related Protein-1 Homolog; D002470:Cell Survival / Q000187:drug effects; D002518:Ceramides / Q000494:pharmacology*; D015179:Colorectal Neoplasms; D004789:Enzyme Activation / Q000187:drug effects; D000068338:Everolimus; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D012265:Ribonucleotides / Q000494:pharmacology; D015398:Signal Transduction; D020123:Sirolimus / Q000031:analogs & derivatives / Q000494:pharmacology
chemical_list: D000970:Antineoplastic Agents; D002518:Ceramides; D047908:Intracellular Signaling Peptides and Proteins; D012265:Ribonucleotides; D000620:Aminoimidazole Carboxamide; C401859:temsirolimus; D000068338:Everolimus; D000071189:Autophagy-Related Protein-1 Homolog; D017346:Protein Serine-Threonine Kinases; C484754:ULK1 protein, human; D055372:AMP-Activated Protein Kinases; C031143:AICA ribonucleotide; C504327:N-(alpha-hydroxyoctadecanoyl)phytosphingosine; D020123:Sirolimus