Fig. 3

Lactate conversion to pyruvate in AF cells. ¹³C-lactate tracing to pyruvate in rabbit AF cells in cell culture containing 4 mM 3-¹³C-lactate (a) and in AF tissue extract derived from the rabbit functional spine unit (FSU) culture 3 days post injection of 5 μl of 40 mM ¹³C-lactate into the NP region of the disc (b). Intracellular enrichment of ¹³C lactate or ¹³C pyruvate AF cells from AF cell culture or AF tissue of the ex vivo disc organ culture is reported as atomic percent excess (APE) of the total amount of lactate or pyruvate, e.g., 10% APE of pyruvate indicates 10% of total pyruvate contains ¹³C. Percent (%) APE shown (M + 1) indicates that one ¹³C carbon is present. c, Schematic of gene expression and assembly of lactate dehydrogenase (LDH), the primary enzyme that catalyzes the interconversion of lactate and pyruvate. LDH exists in five isozymes composed of a tetramer of M and H protein subunits encoded by the LDHA and LDHB genes, respectively. LDH5, composed of four M subunits, preferentially converts pyruvate to lactate while LDH1, consisting of four H subunits, preferentially converts lactate to pyruvate. Western blot of LDHM (d) and LDHH (e) in rabbit AF and NP tissue protein extract and their protein levels were quantified by normalizing to β-actin as loading control (graphs). Data are means ± SD of four independent experiments (four rabbits). * p < 0.05