Evaluation of Impacts of Cellular Metabolism on the Migration of Ovarian Cancer Cells by Two in Vitro Assays: A Method Comparison Study
AbstractBackground: Alteration of metabolic pathways in cancer cells can intensely modulate their migration as an important step in invasion and metastasis. Ketogenic diet showed some contradictory results in cancer patients. In this study the impact of metabolic reprogramming of A2780CP as a model of ovarian cancer stem-like cells on cell migration by two in vitro methods: wound healing and soft agar colony-forming assays. Materials and Methods: short term and long term metabolic reprogramming were done by restriction of glucose to 250mg/L with or without enrichment with beta-hydroxybutyrate (5 milimolar) for 48 hours and 30 days, respectively. Wound healing assay was done and the wound ratio was calculated for 24 and 48 hours. Soft agar colony formation assay was also done in treated and control cells. For method comparison, ten biological replicates were analyzed in triplicate. Results: Migration of A2780CP ovarian cancer stem-like cells were significantly alleviated by long term glucose restriction but no significant changes were observed in short term study. Beta-hydroxybutyrate enrichment did not produce significant impacts on glucose restriction in short or long term studies. Conclusion: The results of colony formation in soft agar and wound or scratch healing assay were in good correlation and convergence which could be used interchangeably in the investigation of metabolic reprogramming in cancer cells. [GMJ.2020;9:e1831]
Pijuan J, Barcelo C, Moreno DF, Maiques O, Siso P, Marti RM et al. In vitro Cell Migration, Invasion, and Adhesion Assays: From Cell Imaging to Data Analysis. Front Cell Dev Biol. 2019;7:107.
Cappiello F, Casciaro B, Mangoni ML. A Novel In Vitro Wound Healing Assay to Evaluate Cell Migration. J Vis Exp. 2018(133).
Grada A, Otero-Vinas M, Prieto-Castrillo F, Obagi Z, Falanga V. Research Techniques Made Simple: Analysis of Collective Cell Migration Using the Wound Healing Assay. J Invest Dermatol. 2017;137(2):e11-e6.
Martinotti S, Ranzato E. Scratch Wound Healing Assay. Methods Mol Biol. 2019.
Borowicz S, Van Scoyk M, Avasarala S, Karuppusamy Rathinam MK, Tauler J, Bikkavilli RK et al. The soft agar colony formation assay. J Vis Exp. 2014(92):e51998.
Horibata S, Vo TV, Subramanian V, Thompson PR, Coonrod SA. Utilization of the Soft Agar Colony Formation Assay to Identify Inhibitors of Tumorigenicity in Breast Cancer Cells. J Vis Exp. 2015(99):e52727.
Finley LWS. Metabolic signal curbs cancer-cell migration. Nature. 2019;571(7763):39-40.
Magee BA, Potezny N, Rofe AM, Conyers RA. The inhibition of malignant cell growth by ketone bodies. Australian Journal of Experimental Biology and Medical Science. 1979;57(5):529-39.
Seyfried TN, Flores RE, Poff AM, D'Agostino DP. Cancer as a metabolic disease: implications for novel therapeutics. Carcinogenesis. 2013;35(3):515-27.
Woolf EC, Curley KL, Liu Q, Turner GH, Charlton JA, Preul MC et al. The ketogenic diet alters the hypoxic response and affects expression of proteins associated with angiogenesis, invasive potential and vascular permeability in a mouse glioma model. PLoS One. 2015;10(6):e0130357.
Tseng P-L, Chen C-W, Hu K-H, Cheng H-C, Lin Y-H, Tsai W-H et al. The decrease of glycolytic enzyme hexokinase 1 accelerates tumor malignancy via deregulating energy metabolism but sensitizes cancer cells to 2-deoxyglucose inhibition. Oncotarget. 2018;9(27):18949-69.
O'Flanagan CH, Smith LA, McDonell SB, Hursting SD. When less may be more: calorie restriction and response to cancer therapy. BMC medicine. 2017;15(1):106.
Chung H-Y, Park YK. Rationale, feasibility and acceptability of ketogenic diet for cancer treatment. Journal of cancer prevention. 2017;22(3):127.
Karakuş F, Eyol E, Yılmaz K, Ünüvar S. Inhibition of cell proliferation, migration and colony formation of LS174T Cells by carbonic anhydrase inhibitor. Afr Health Sci. 2018;18(4):1303-10.
Gupta R, Yang Q, Dogra SK, Wajapeyee N. Serine hydroxymethyl transferase 1 stimulates pro-oncogenic cytokine expression through sialic acid to promote ovarian cancer tumor growth and progression. Oncogene. 2017;36(28):4014-24.
Saha S, Ghosh M, Dutta SK. The dual-hit metabolic modulator LDCA synergistically potentiates doxorubicin to selectively combat cancer-associated hallmarks. RSC advances. 2017;7(84):53322-33.
Authors who publish with this journal agree to the following terms:
- Authors retain copyright and grant the journal right of first publication with the work simultaneously licensed under a Creative Commons Attribution 4.0 International License that allows others to share the work with an acknowledgment of the work's authorship and initial publication in this journal.
- Authors are able to enter into separate, additional contractual arrangements for the non-exclusive distribution of the journal's published version of the work (e.g., post it to an institutional repository or publish it in a book), with an acknowledgment of its initial publication in this journal.
- Authors are permitted and encouraged to post their work online (e.g., in institutional repositories or on their website) prior to and during the submission process, as it can lead to productive exchanges, as well as earlier and greater citation of published work (See The Effect of Open Access).