Fig. 7: BCFA increases hepatic glucose production and activates the mTORC1/S6K1 pathway in vitro.

Branched-chain fatty acids (BCFA) effect on hepatic glucose production in hepatocytes (FAO) in basal or insulin (1 nM) condition with (a) isovaleric (isoV) or (b) isobutyric (IsoB) acid (1–1000 µM); data corrected for total protein. For IsoV treatment in basal condition (dark red): n = 6 independent experiments for 1 µM, 500 µM and 750 µM, n = 7 for 10 µM and 250 µM, n = 11 for 1 mM, and n = 13 for 100 nM. For IsoV treatment in insulin condition (light purple): n = 5 independent experiments for 750 µM, n = 6 for 1 µM, n = 7 for 10 µM, n = 8 for 250 µM and 500 µM and n = 12 for 100 µM and 1 mM. For IsoB treatment in basal condition (blue-green): n = 6 independent experiments for 250 µM, n = 7 for 1 µM, 10 µM, 500 µM and 750 µM, n = 13 for 100 µM, n = 15 for 1 mM. For IsoB treatment in insulin condition (khaki green): n = 6 independent experiments for 750 µM, n = 7 for 1 µM, 10 µM and 250 µM, n = 9 for 500 µM and n = 14 for 100 µM and 1 mM. Immunoblots and quantification of densitometry analyses for pS6 S240-244 and total S6 in FAO cells exposed to 1 mM of (c, d) isovaleric (dark red) and (e, f) isobutyric (blue-green) acid. eEF2 has been used as a loading control and n = 4 independent experiments. Data are means ± s.e.m. A Kruskal-Wallis test followed by a two-tailed Dunn’s post-hoc test versus Vehicle/Vehicle + Insulin was performed in each condition and a general p-value is recorded under the title of each graph. Detailed significant differences detected by post-hoc test are recorded as follows: *p < 0.05, **p < 0.01, ***p < 0.001. See also Supplementary Fig. 9.