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. 2014 May 8;5(5):e1209. doi: 10.1038/cddis.2014.175

Figure 2.

Figure 2

OKA, a PP2A inhibitor, prevents metformin-induced reduction of phospho-Ser129 α-syn. SH-SY5Y cells were pretreated with retinoic acid and tetracycline. Cells were then incubated for 16 h with no addition or in the presence of OKA alone (15 nM), metformin alone (2.5 mM) or both OKA and metformin. Levels of hα-syn, phospho-Ser129 hα-syn and tubulin were assayed by immunoblotting (a). Semiquantitative analysis of band intensities is expressed as phospho-Ser129 hα-syn/hα-syn ratio (b). In separate experiments, cells were treated with no addition or with 300 nM OKA for 4 h, and levels of hα-syn, phospho-Ser129 hα-syn and tubulin were assayed by western blotting (c, first two lanes). In cultures preincubated with OKA, the medium was removed, cells were washed with fresh medium and incubations were continued for 16 h in the absence or presence of 2.5 mM metformin. Levels of hα-syn, phospho-Ser129 hα-syn and tubulin were assayed by western blotting (c, third and fourth lanes). Analysis of band intensities is expressed as phospho-Ser129 hα-syn/hα-syn ratio (d). Data in (b and d) are the means±S.E.M. of at least three independent experiments. *P<0.05, **P<0.01 and ***P<0.001 versus the respective control group (black bar). †P<0.05 versus the group treated with OKA alone (b). ##P<0.01 versus the group treated with both OKA and metformin (b). †††P<0.001 versus the group pretreated with OKA for 4 h (d). #P<0.05 versus the group pretreated with OKA and then incubated in the absence of metformin (d)