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. 2011 Nov 29;153(1):92–100. doi: 10.1210/en.2011-1442

Fig. 4.

Fig. 4.

Targeting of hOGG1 to mitochondria in L6 myotubes ameliorated palmitate-mediated inhibition of insulin-induced. A, C tyrosine phosphorylation of IRS-1. D and E, Akt (Ser473) phosphorylation. G and F, GLUT4 translocation to the PM. MTS-hOGG1- or GFP-transduced L6 myotubes were exposed to control medium (C) (2% BSA) or to medium containing 1 mm palmitate (P) for 16 h and then treated with insulin as described in Materials and Methods. Total cell lysates or PM (as specified) were isolated and analyzed by Western blot analysis with the indicated antibodies. A, top panel, IRS-1 was immunoprecipitated from 200 μg of total cell lysates, and Western blottings were performed using an pTyr antibody. Bottom, Representative Western blotting of total IRS-1. B, Densitometry data for the total IRS-1 protein levels normalized to GFP control data (n ≥ 3). *, P < 0.05 vs. all other groups. C, Densitometry data for insulin dependent (pTyr-IRS-1) were normalized to GFP (control plus insulin) data and presented as means ± se (n ≥ 3). *, P < 0.05 vs. GFP-transduced cells treated with palmitate. D, Representative blots from at least three independent experiments for phosphorylation of Akt (Ser473) are shown. Total cell lysates were used. E, The values from densitometry from at least three (pAkt) independent experiments were normalized to the level of total Akt and expressed as fold of difference after addition of insulin normalized to the GFP (control plus insulin) data. The mean results ± se are shown. *, P < 0.05 vs. GFP-transduced cells treated with palmitate. G, PM were isolated from adenovirus-transduced L6 myotubes, as described in Materials and Methods, and proteins were analyzed by Western blotting using GLUT4 antibody. Immunodetection of the α-subunit of Na+/K+-ATPase was performed to confirm PM localization. The values from densitometry performed on three to four independent GLUT4 translocation to PM independent experiments were normalized to the level of α-subunit of Na+/K+-ATPase and then normalized to the GFP (control plus insulin) data ± se (n ≥ 3). *, P < 0.05 vs. GFP-transduced cells treated with palmitate.