(A) Total sciatic nerve lysates (25 µg/lane) from the indicated ages and diet were analyzed with anti-heat shock factor 1 (HSF1) antibody. The same nerve lysates (20 µg/lane) were also probed with antibodies against HSP90, HSP70, HSP40, HSP27 and αB-crystallin. (B) Quantification of HSF1 and HSP90 protein levels normalized to GAPDH from three independent experiments (##p<0.01, ###p<0.001, Fisher’s PLSD, mean ± SEM), AU: arbitrary units. The effect of CR on these proteins was analyzed by comparing HSF1 or HSP90 protein values with age-matched AL counterparts (*p<0.05, **p<0.05, unpaired t-test, mean ± SEM). (C) Steady-state levels of LAMP1, Atg7, pS6 and S6 proteins in sciatic nerves from AL and CR rats were analyzed by Western blots. Blots were reprobed with anti-GAPDH antibody as protein loading control. Molecular mass at the left, in kDa. (D) Quantification of LAMP1 and Atg7 protein levels normalized to GAPDH from three independent experiments (#p<0.05, ###p<0.001, Fisher’s PLSD analysis, *p<0.05, unpaired t-test, mean ± SEM). (E) Blots of pS6 and S6 from three independent experiments were quantified and the values are represented as ratio of pS6/S6. The pS6/S6 ratio of 8-mo old AL sample was set as 1 (*p<0.05, unpaired t-test, mean ± SEM). A-D, n=3 rats per condition.