glucose complete media  (Thermo Fisher)

Journal: Nature Communications

Article Title: Glucose-6-phosphate dehydrogenase maintains redox homeostasis and biosynthesis in LKB1 -deficient KRAS -driven lung cancer

doi: 10.1038/s41467-024-50157-8

Figure Lengend Snippet: a Scheme illustrating KL tumor-derived cell lines (TDCLs) generation (Created with BioRender.com released under a Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International license). b Western blot of G6pd WT and G6pd KO KL TDCLs. Uncropped Western blot image is shown in Source Data file. c Pool size of NADPH and NADP + , and NADPH/NADP + ratio of KL TDCLs in nutrient rich conditions (complete RPMI medium). n = 4 replicates for each clone. d Pool size of GSH and GSSG, and GSH/GSSG ratio of KL TDCLs in nutrient rich conditions. n = 4 replicates for each clone. e Basal ROS level of KL TDCLs in nutrient rich conditions. n = 3 replicates for each clone. f Proliferation rate of KL TDCLs in nutrient rich conditions measured by Incucyte for 4 days (left) with statistical analysis at day 4 (right). n = 3 replicates for each clone. g Relative proliferation rate of KL TDCLs treated with G6PDi-1 for 48 hours. n = 6 replicates for each clone at different G6PDi-1 concentrations, except 2489-2 at 40 μmol/L G6PDi-1 with n = 5 replicates. h Relative proliferation rate of KL TDCLs treated with H 2 O 2 for 24 hours. For each clone, n = 6 replicates at 0 μmol/L H 2 O 2 , n = 3 replicates at 20, 40, 80 μmol/L H 2 O 2 . i ROS levels of KL TDCLs treated with 20 μmol/L H 2 O 2 for 24 hours. For each clone, n = 3 replicates at 0 and 20 μmol/L H 2 O 2 . j Growth curve of KL allograft tumors from mice treated with or without high-dose Vitamin C (Vit C). n = 10 allograft tumors for each group. k Gross pathology of allograft tumors from ( j ). Scale bar = 1 cm. l Graph of allograft tumor weight from ( k ). m Representative IHC images and quantification of NRF2 ( n = 15 images for each quantification), NQO1 ( n = 10 images for each quantification), and 8-oxo-dG ( n = 15 images for each quantification) of allograft tumors from ( k ). Scale bar = 100 μm. Data are presented as mean ± SEM, significance was calculated by two-tailed unpaired t -test (NADPH and NADP + in c , GSSG and GSH/GSSG in d ), two-tailed unpaired t -test with Welch’s correction (NADPH/NADP + in c , GSH in d ), one-way ANOVA followed by Bonferroni’s multiple comparisons test ( e , f , g , h , i , m ), or one-way ANOVA followed by t -test ( l ). Source data are provided as a Source Data file.

Article Snippet: G6pd WT ;KL and G6pd KO ;KL TDCLs were cultured in customized complete RPMI medium (RPMI medium without glucose, serine, and glycine (Teknova, Cata#R9660), supplemented with 2 g/L glucose (Sigma, Cata#G8270-1KG), 10 mg/L glycine (Sigma, Cata#50046-50 G) and 30 mg/L serine (Sigma, Cata#S4311-25G)) with 10% fetal FBS, 1% Penicillin-Streptomycin, 0.075% sodium bicarbonate, at 37°C with 5% CO 2 .

Techniques: Derivative Assay, Western Blot, Two Tailed Test

Journal: Nature Communications

Article Title: Glucose-6-phosphate dehydrogenase maintains redox homeostasis and biosynthesis in LKB1 -deficient KRAS -driven lung cancer

doi: 10.1038/s41467-024-50157-8

Figure Lengend Snippet: a Normalized 13 C labeling fraction from glucose to serine and glycine of KL lung tumors. b Pool size of serine and glycine of KL lung tumors. c Serine consumption of G6pd WT ;KL and G6pd KO ;KL TDCLs in nutrient rich conditions. d 2 H labeling fraction of serine in G6pd WT ;KL and G6pd KO ;KL TDCLs after 4 hours [2,3,3- 2 H]-serine labeling in nutrient rich conditions. n = 6 replicates for each clone. e Scheme of hydrogen contribution from [2,3,3- 2 H]-serine to NADPH (Created with BioRender.com released under a Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International license). f Scheme of in vivo [2,3,3- 2 H]-serine tracing (Created with BioRender.com released under a Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International license). g 2 H labeling fraction of serine in G6pd WT ;KL ( n = 8 mice) and G6pd KO ;KL ( n = 7 mice) lung tumors at 12 weeks post-tumor induction. h NADPH active-H labeling from [2,3,3- 2 H]-serine in G6pd WT ;KL ( n = 8 mice) and G6pd KO ;KL ( n = 7 mice) lung tumors at 12 weeks post-tumor induction. i NADPH/NADP + ratio of G6pd WT ;KL and G6pd KO ;KL TDCLs cultured with RPMI medium with or without serine and glycine for 24 hours. RPMI denotes cells cultured in complete RPMI medium, RPMI w/o GS denotes cells cultured in complete RPMI medium without serine and glycine. n = 3 replicates for each clone under different conditions. j Relative GSH/GSSG ratio of same samples from ( i ). k ROS levels of G6pd WT ;KL and G6pd KO ;KL TDCLs cultured with RPMI medium with or without serine and glycine for 48 hours. n = 6 replicates for each clone under different conditions, except 2489-2 with n = 4 replicates. l Relative proliferation rate of G6pd WT ;KL and G6pd KO ;KL TDCLs cultured with RPMI medium with or without serine and glycine for 48 hours. n = 3 replicates for each clone under different conditions. m Model of G6PD-mediated KL lung tumor growth (Created with BioRender.com released under a Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International license). Data are presented as mean ± SEM, significance was calculated by two-tailed unpaired t -test ( a , b , d , g , h ), or two-way ANOVA followed by Bonferroni’s multiple comparisons test ( i , j , k , l ). Source data are provided as a Source Data file.

Article Snippet: G6pd WT ;KL and G6pd KO ;KL TDCLs were cultured in customized complete RPMI medium (RPMI medium without glucose, serine, and glycine (Teknova, Cata#R9660), supplemented with 2 g/L glucose (Sigma, Cata#G8270-1KG), 10 mg/L glycine (Sigma, Cata#50046-50 G) and 30 mg/L serine (Sigma, Cata#S4311-25G)) with 10% fetal FBS, 1% Penicillin-Streptomycin, 0.075% sodium bicarbonate, at 37°C with 5% CO 2 .

Techniques: Labeling, In Vivo, Cell Culture, Two Tailed Test