Figure 2. Hyperoxic exposure increases the PPP in cultured lung ECs.

(A–E) MFLM-91U cells (A and E) and primary LMVECs (B–D) were exposed to hyperoxia for 24 hours and then cultured in normoxia for 24 hours (refers to O₂) unless specifically mentioned. (A) [3-¹³C]lactate was measured by the NMR when cells were incubated with [1,2-¹³C]glucose (20 mM) for 24 hours during air recovery phase. n = 4 per group. (B) Western blot was performed to determine levels of PGD and G6PD proteins. O₂ w/o rec refers hyperoxic exposure for 24 hours without air recovery, while O₂ refers hyperoxic exposure for 24 hours, followed by air recovery for 24 hours. n = 6 in air, n = 7 in hyperoxia without air recovery, and n = 3 in hyperoxia with air recovery. (C) NADPH levels were measured using a commercially available kit. n = 6 in air and n = 9 in hyperoxia. (D) Levels of ribulose 5-phosphate, reduced and reduced/oxidized glutathione were determined through metabolomics analysis. n = 4 in air and n = 6 in hyperoxia. (E) Ratio of [3-¹³C]lactate to [2,3-¹³C]lactate was calculated based on results from Figure 1D and A. n = 4 per group. Data are expressed as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001 versus air using 1-tailed t test (A, C, D, and E) or ANOVA followed by Tukey-Kramer test (B).