Fig. 6.

Both GP and AC chondrocytes exhibit enhanced glutamine metabolism in the absence of glucose availability. a A Seahorse assay was used to assess the oxygen consumption rates (OCRs) of control and Glut KO primary GP and AC chondrocytes. The data are the mean ± SD. N = 8. *P < 0.05 relative to the controls. b Glutamine consumption by control and Glut1 KO primary GP and AC chondrocytes for 24 h. The data were normalized to the genomic DNA content and are expressed as the mean ± SD. N = 5. *P < 0.05. c Western blot analyses of glutaminase (Gls), Glut1, pS6 and S6 protein levels in control and Glut KO primary GP and AC chondrocytes. N = 3. d Immunofluorescence staining for Gls in tibia and knee sections of control and Glut1 LOF mice at 2 months. N = 6. Scale bar, 100 μm. e Graphical depiction of glutamine metabolism tracing with [U-¹³C] glutamine. The black filled circles denote ¹³C, and the black open circles indicate ¹²C. Gln glutamine, Glu glutamate, α-KG α-ketoglutarate, OAA oxaloacetate. Glutamine is converted to citrate through glutamine oxidation, as indicated by the red dashed line. The glutaminolysis pathway contains glutamine oxidation in the TCA cycle and the malate-aspartate shuttle. Glutamine can also be converted via reductive carboxylation, as denoted by the green dashed line. f–h Fractions of citrate m + 4 and m + 5 (f), aspartate m + 4 (g), and proline m + 5 (h) after culture of control and Glut KO GP and AC chondrocytes for 24 h with [U-¹³C] glutamine. The data are the mean ± SD. N = 3. *P < 0.05 compared to the controls