Extended Data Figure 6. Dietary restriction dramatically preserves neurofunctioning of Ercc1^Δ/− mice.

a, Quantification of TUNEL-positive cells in the outer nuclear layer of retinal sections of 16 week old AL (blue) or DR (red) Ercc1^Δ/− mice; n=4 animals/group. b, Analysis of the total number of motor neurons with an abnormal Golgi apparatus (indicative of cells with ill health; see thick arrows in representative image; neuron with normal Golgi is indicated by a thin arrow) in C6 cervical spinal cord sections from 16 week old DR and AL Ercc1^Δ/− mice. n=4 animals/group. TUNEL-positive cells (a) and neurons with abnormal Golgi morphology (b) were absent in both AL-¹⁷ and DR-treated young wt mice. c, Quantitative stereological analysis of the total number of non-neuronal cells (Dapi+/NeuN-; p=0.2744) in the neocortex of transverse brain sections of 16 weeks old AL and DR Ercc1^Δ/− mice. n≥3 animals/group. Error bars indicate mean ± SE. ***p<0.001. d, Representative images of neocortex stained for NeuN (neurons), P53 and Dapi (for staining DNA) used for quantitative stereological analysis of the total number of neurons (NeuN+) and non-neuronal cells (Dapi+/NeuN−) in 16 weeks old AL- (n=3) and DR-(n=4) treated Ercc1^Δ/− mice. Quantification of the number of P53-positive neurons is shown in Figure 3f. The analysis was performed using the optical dissector probe from StereoInvestigator on a Zeiss LSM700 laser scanning microscope. e, Representative image of cerebellum stained for yH2AX (green; double stranded DNA breaks) and Dapi (blue; for staining DNA) in 16 weeks old AL (n=3) and DR (n=4) treated Ercc1^Δ/− mice. The Purkinje (PkJ) neurons are present in a single layer (PL; purkinje layer) in between the molecular layer (ML) and granular layer (GL)⁴¹. Quantification of the number of yH2AX-positive PkJ-neurons is shown in Figure 3i. The analysis was performed using a Zeiss LSM700 laser scanning microscope.