Priority Research Paper Volume 12, Issue 11 pp 10041—10058

Figure 1. Age-related increase in Ldh expression is associated with elevated LDH protein levels, enzyme activity, and lactate concentration. (A) Profile of Ldh mRNA expression in the heads of young (5-days-old) and old (55-days-old) w¹¹¹⁸ flies measured by qRT-PCR. Compared to young, old flies have increased Ldh mRNA levels at each time point with a peak at ZT12. Values are averages of 4 biorepeats reported as a percentage of expression relative to young at ZT0 set to 100%. (B) Representative western blot of LDH protein levels in young and old fly heads with tubulin as a loading control. (C) Graph showing LDH enzyme activity for both the pyruvate to lactate and lactate to pyruvate reactions in the heads of young (grey bars) and old (blue bars) flies. Enzymatic activity of LDH is higher in the heads of old flies compared to young for the pyruvate to lactate reaction (*p<0.05 by unpaired one-tailed t-test with Welch’s correction). (D) Lactate levels are significantly higher in the heads of old flies (blue bars) at both time points compared to young. In old flies, there is also a significant difference in lactate levels between ZT0 and ZT12 (n=4; p<0.001 by unpaired t-test with Welch’s correction). (E) Pyruvate levels do not significantly change between young (grey bars) and old (blue bars) (n=4). (F) Average lactate/pyruvate ratios increase in old flies (blue bars) (n=4). Error bars in A, C–F indicate standard error of the mean (SEM). Significance between age groups and time of day in each graph determined by 2-Way ANOVA with Bonferroni's correction (**p<0.01, ***p<0.001, and ****p<0.0001).