Figure 5: Oxidative stress and monoamine oxidase (MAO) transcription in the four lines of induced pluripotent stem cell (iPSC)-derived neurons.
(a,b) Western blot analysis (a) and quantification (b) of protein carbonyls in the four lines of iPSC-derived neurons treated without or with dopamine (75 μM for 4 h). DNP, 2,4-dinitrophenyl; p150, p150glued, a dynactin subunit as loading control. *P<0.01 versus C001 or C002 with DA treatment, n=8, Student's t-test, unpaired, two-tailed. (c) Quantitative reverse transcriptase PCR (qRT–PCR) measurement of MAO-A and MAO-B transcripts in the four lines of iPSC-derived neuronal cultures. *P<0.01 versus C001 or C002, n=9, Student's t-test, unpaired, two-tailed. (d) MAO-A and MAO-B activities in the four lines of iPSC-derived neurons. *P<0.01 versus C001 or C002, n=6–11, Student's t-test, unpaired, two-tailed. (e,f) qRT–PCR measurements of MAO-A and MAO-B messenger RNA (mRNA) levels in P001 (e) or P002 (f) neurons infected with lentivirus expressing green fluorescent protein (GFP), parkin or T240R mutant parkin. (g,h) MAO-A and MAO-B activities in P001 (g) or P002 (h) neurons infected with lentivirus expressing GFP, parkin or T240R mutant parkin. (i–k) Western blot analysis (i) and quantification (k) of protein carbonyls in P001 or P002 neurons infected with lentivirus expressing GFP, parkin or T240R mutant parkin. Total cell lysates were blotted with antibodies against GFP or FLAG or p150glued (j). *P<0.05 versus GFP-expressing neurons, n=3–4 for (e) to (k), Student's t-test, unpaired, two-tailed. mg, milligrams of total cellular proteins. Error bars represent s.e.m.
