Changes in extracellular matrix rigidity induces metabolic reprogramming of U87 glioblastoma cells
Introduction
Glioblastoma (GBM) is a highly aggressive and invasive form of brain cancer, with 5-year survival rate less than 10%. Recent research has proposed important roles of mechanotransduction in the development of the aggressive phenotype of this cancer . It has been suggested that biochemical and biophysical cues of the tumor microenvironment, influence cellular survival, proliferation and invasiveness of GBM. The extracellular matrix (ECM) is a critical component of the microenvironment and stiffening of the ECM has been shown to enhance GBM cell migration and proliferation. However, the effect of ECM on metabolic reprogramming of GBM is not known. The purpose of this study was to investigate the effect of ECM stiffness on the metabolic profile of GBM cells.
Methods
In this project, we used U87MG cells and premanufactured collagen coated hydrogels of 100Pa, 4kPa, and 25kPa stiffness. Hydrogels were selected based on the stiffness of normal brain tissue (100Pa) and the tumor stiffness seen in the GBM ECM (4kPa and 25kPa). Effect of changing ECM stiffens on U87MG cells was evaluated by metabolomics, label-free two-photon imaging for optical REDOX ratio (NADH/FAD) providing spatial cellular assessment and heterogeneity as well as morphometry, global RNAseq, and cell migration assay. We measured stiffness-dependent metabolic reprogramming with increase of key metabolites and amino acids of glycolysis and glutamine pathways.
Results
Both, metabolomics data and two-photon imaging of the optical REDOX ratio, identified a decrease in FAD and an increase of NADH as a function of increased ECM stiffness. We also detected decrease in oxidative phosphorylation with an increase in ECM stiffness, suggesting transition from oxidative phosphorylation to glycolysis. Global RNAseq data identified over 100 significant changes in RNA expression between he 100Pa, 4kPa and 25kPa stiffnesses, and provides critical insight into the effect ECM rigidity has on gene expression relating to metabolism and cell proliferation. Finally, marked changes cell morphology and migration as a function of stiffness were detected.
Conclusion
Together, our study shows a metabolic reprogramming of U87MG cells increased glycolysis and glutaminolysis in response to increase ECM stiffness, suggesting a potential link between the biomechanical microenvironment with metabolic activity that could support the aggressive and resistant nature of GBM. Ongoing studies are investigating potential mechanosensors that induce metabolic switching observed in these GBM cells.