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. 2020 Oct 19;9(10):2323. doi: 10.3390/cells9102323

Figure 1.

Figure 1

Pregnenolone and TSPO oscillations are dependent on a functional circadian clock in A172 cells. (A) P5 (pregnenolone) concentration was quantified in lysates from synchronized A172 glioma cells using a direct pregnenolone ELISA test. Time points from 12 to 36 h post-synchronization are shown. All data are normalized to 1 and presented as mean ± SD (three independent experiments, seven time points, n = 12 for each time point). (B) P5 concentration in synchronized PER1/PER2 siRNA transfected A172 glioma cells. Time points from 12 to 36 h post-synchronization are shown. All data are normalized to 1 and presented as mean ± SD (three independent experiments, seven time points, n = 3–5 for each time point). (C) Translocator protein (TSPO) levels were analyzed in cell lysates from synchronized A172 glioma cells by immunoblotting. Time points from 12 to 36 h post-synchronization are shown. All data are normalized to 1 and presented as mean ± SD (three independent experiments, seven time points, n = 4–6 for each time point). (D) TSPO protein levels in synchronized PER1/PER2 siRNA transfected A172 glioma cells. Time points from 12 to 36 h post-synchronization are shown. All data are normalized to 1 and presented as mean ± SD (two independent experiments, seven time points, n = 3–5 for each time point). (E) Representative immunoblots of synchronized A172 glioma cell lysates stained with antibodies specific for TSPO and VDAC (voltage-dependent anion-selective channel = control). Time points from 12 to 36 h post-synchronization are shown. (F) Representative immunoblots of synchronized PER1/PER2 siRNA transfected A172 cells stained with antibodies specific for TSPO and VDAC (control). Time points from 12 to 36 h post-synchronization are shown.