4. Conclusion
In this study, we systematically evaluated 12 combinations of quenching and extraction methods for acquiring quantitative intracellular metabolome with both 2D and 3D Hela carcinoma models based on the isotope dilution mass spectrometry method. Our results showed that liquid nitrogen as quenching agent and 50% acetonitrile as extracting agent was proven to be the most optimal method to acquire intracellular metabolome for both 2D Hela and 3D MTSs. Based on this, a quantitative metabolomics study showed that DOX exposure gave rise to pronounced metabolite changes between 2D and 3D Hela carcinoma models. Strikingly, our data suggested that compared to 2D cells the increased intracellular glutamine level in 3D cells can benefit replenishing the TCA cycle when the glycolysis was limited after dosing with DOX.
In summary, this study provides a well-established quenching and extraction protocol for quantitative metabolome profiling of Hela carcinoma cell under 2D and 3D cell culture conditions. Based on this, quantitative time-resolved metabolite data can serve to the generation of hypotheses on metabolic reprogramming to reveal its important role in tumor development and treatment.