Fig. 4.

SH-SY5Y cells exhibit decreased mitochondrial activity and increased levels of ROS with age. (A) Quantification of fluorescent intensity in H₂-DCMFA labelled cultures 2 weeks and 4 weeks post-differentiation. The means and standard errors of the means of 8 separate experiments established in duplicate and with at least five fields of view imaged per coverslip are shown. *** represents statistical significance (P < 0.001). (B) Images of 2 (Bi) and 4 (Bii) week cultures stained with the fluorescent dye H₂-DCFDA to demonstrate ROS levels. Images were taken at 20 x magnification and scale bar represents 25 µm. (C) Bar chart of relative levels of mitochondrial redox capacity in cultures 2 weeks and 4 weeks post-differentiation. The means and standard errors of the means of 6 separate experiments are shown. *** represents statistical significance (P < 0.001). (D) Images of cultures immunocytochemically labelled to determine HNE expression cultures 2 weeks (Di) and 4 weeks (Dii) post-differentiation and negative control image (Diii). Images were taken at 40 x magnification and scale bar represents 25 µm. (E) Representative image of an oxyblot to detect protein oxidation in cultures 2 weeks and 4 weeks post-differentiation and image of a corresponding western blot of these samples demonstrating α-tubulin expression as a loading control. (F) Quantification of the amount of oxidatively modified protein in cultures 2 weeks and 4 post-differentiation. The means and standard errors of the means of 11 separate experiments are shown. ** represents statistical significance (P < 0.01).