Fig. 2. A time-dependent increase of DNA fragmentation and caspase activity is detected in MN9D cells cultivated in 5 mM glucose but not in 17.5 mM glucose. MN9D cells cultivated in N2 medium containing either 5 mM or 17.5 mM glucose were exposed to 200 µM MPP+ for the indicated time periods. (A) MTT reduction assay was performed. Values were expressed percent survival over the untreated control (100%). Data represent the mean±SEM from 3 independent cultures in triplicate. No significant difference was measured between two groups. (B) Total DNA extracted from MN9D cells exposed to 200 µM MPP+ for 4 (lane 1, 5), 8 (lane 2, 6), 12 (lane 3, 7) and 18 hr (lane 4, 8) was separated on 1.2% agarose gel. Gels were stained with ethidium bromide and then photographed on a UV-transilluminator. (C) Immunoblot analysis for active forms of capsase-9 and caspase-3 was carried out as described above. (D, E) Cells cultivated either in (D) 5 mM glucose or (E) 17.5 mM glucose were exposed to 200 µM MPP+ for 24 hr in the presence or the absence of the indicated concentrations of zVAD or BAF. Viability was measured using MTT reduction assay and data represent the mean±SEM from 3 independent cultures in triplicate. **p<0.01; *p<0.05; n.s., not significant.
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