Fig. 3.

Gln Induced Oxygen Consumption and Ammonia Production. (A) Seahorse XF Oxygen Consumption Response in Mito Stress Test. MCEC in complete media were preincubated for 45 min at 37 °C (in the absence of CO₂) in XF Base Medium containing 14 mM glucose ± 4 mM Gln, then placed into Seahorse analyzer. Basal OCR was followed by ATP related OCR (2 μM oligomycin), then complete uncoupling to give Maximum OCR (0.5 μM FCCP), followed by ETC blockade (0.5 μM Antimycin A and 0.5 μM Rotenone). (B) Summary of results from (A), n = 3, ±SEM, *p < 0.05 vs Glucose for same genotype; δ:p < 0.05 vs WT, for same treatment. (C & D) OCR trace and summary results of hSLC4A11 over-expressing PS120 fibroblasts. n = 3, *p < 0.05 vs Gluc, same genotype. δ: p < 0.05 vs EV; EV-Empty Vector. (E) Ammonia production in DMEM assay media (0.5% dialyzed-serum) with glucose or glucose + 0.5 mM glutamine in Slc4a11 WT and KO MCEC, n = 3 for each condition, *p < 0.05 less than WT for same condition. (F) Effect of CRISPR KD of SLC4A11 in HCEC on Ammonia production and (G) Basal OCR in Gln; n = 3, *p < 0.05.