Figure 7.

Cells are more dependent on exogenous glycine and are more sensitive to folate deficiency after suppression of MTHFD2. A. U87 and U118 cells were transfected with miR-NC or miR-940 mimic, and the expression of MTHFD2, SHMT2 and MTHFD1L was measured by Western blotting. B. After 96 hours of transfection, intracellular serine and glycine levels were determined by GC-MS (three replicates). C. After 48 hours of transfection, U87 and U118 cells were seeded into 96-well plates with medium containing glycine (glycine+) or not containing glycine (glycine-). Cell proliferation was detected by CCK-8 assay. D. After 2 days of transfection, cells were cultured in folate-free 1640 medium until day 6. The culture medium was replaced every 2 days with medium containing formate (10 mM). Cell viability was not rescued by the addition of formate in the miR-940 mimic group from day 6 to day 12. Cell viability was assayed by trypan blue exclusion according to the equation cell viability (%) = number of unstained cells/total number of cells observed * 100. E. In folate-free culture medium (day 2 to day 6), cell viability decreased faster in the miR-940 mimic groups than in the negative control groups. All data are shown as the mean ± SD of three replicates. *P<0.05, **P<0.01, and ***P<0.001.