Figure 4: Recombinant PK2 protects dopaminergic N27 cells against MPP⁺-induced dopaminergic neuronal cell death.

(a) Representative western blot from one of the three experiments showing no changes in PKR2 expression during MPP⁺-induced cell death in N27 dopaminergic cells. (b) Calcium mobilization (flux) induced by rPK2 in N27 cells, measured as mean fluorescent intensity (MFI) of Fluo-4. Calcium-free HBSS was used on the control group (one-way ANOVA with Bonferroni post-test comparison and n=3). (c,d) Exogenously stimulated PK2 signalling attenuated caspase-3 activation induced by MPP⁺ at 8 h (c) and DNA fragmentation at 16 h after MPP⁺ treatment (d). N27 cells were treated with MPP⁺ (200 μM) or co-treated with PK2 (25 nM) and cells were collected for assays of caspase-3 activity at 8 h and DNA fragmentation at 16 h. The data were expressed as fluorescence units (FU) per mg of protein for caspase-3 activity and absorbance units (AU) per mg protein for DNA fragmentation (one-way ANOVA with Bonferroni post-test comparison and n=3). (e,f) Co-treatment with the specific PK2 receptor antagonist PKRA7 blocked PK2’s neuroprotection in MTS (e) (one-way ANOVA with Bonferroni post-test comparison and n=4–8) and caspase-3 activity (f) (one-way ANOVA with Bonferroni post-test comparison and n=3) assays. N27 cells were treated with 200 μM MPP⁺ for 24 h in the presence or absence of 25 nM PK2 and 2 μM PKRA7. Data represented by group mean±s.e.m. (*P<0.05, ^**P<0.01 and ^***P<0.001).