Autophagy has been reported to have a pivotal role in maintaining stemness, regulating immunomodulation and enhancing the survival of mesenchymal stem cells (MSCs). However, the effect of autophagy on MSC metabolism is largely unknown. Here, we report a workflow for examining the impact of autophagy on human placenta-derived MSC (hPMSC) metabolome profiling with chemical isotope labeling (CIL) LC-MS. Rapamycin or 3-methyladenine was successfully used to induce or inhibit autophagy, respectively. Then, 12C- and 13C-dansylation labeling LC-MS were used to profile the amine/phenol submetabolome. A total of 935 peak pairs were detected and 50 metabolites were positively identified using the dansylation metabolite standards library, and 669 metabolites were putatively identified based on an accurate mass match in metabolome databases. 12C/13C-p-dimethylaminophenacyl bromide labeling LC-MS was used to analyze the carboxylic acid submetabolome; 4736 peak pairs were detected, among which 33 metabolites were positively identified in the dimethylaminophenacyl metabolite standards library, and 3007 metabolites were putatively identified. PCA/OPLS-DA analysis combined with volcano plots and Venn diagrams was used to determine the significant metabolites. Metabolites pathway analysis demonstrated that hPMSCs appeared to generate more ornithine with the arginine and proline metabolism pathway and utilized more pantothenic acid to synthesize acetyl-CoA in the beta-alanine metabolism pathway when autophagy was activated. Meanwhile, acetyl-CoA conversion to fatty acids led to accumulation in the fatty acid biosynthesis pathway. In contrast, when autophagy was suppressed, a reduction in metabolites demonstrated weakened metabolic activity in these metabolic pathways. Our research provides a more comprehensive understanding of hPMSC metabolism associated with autophagy.
Keywords: Autophagy; Chemical isotope labeling; LC−MS; Mesenchymal stem cells; Metabolomics.
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