Supplementary Figure 5: TNF induces insulin resistance in part via ligand-independent activation of the GR. | Nature Cell Biology

Supplementary Figure 5: TNF induces insulin resistance in part via ligand-independent activation of the GR.

From: Identification of nuclear hormone receptor pathways causing insulin resistance by transcriptional and epigenomic analysis

Supplementary Figure 5

(a) Expression of Nr3c1 (GR) in cells 6 days after transduction with lentiviruses carrying shRNAs directed against GR or a scrambled control (shScr) (data represent mean of n = 3 dishes, data from 1 additional independent experiment shown in Source Data Table). (b) Expression of Rela (p65) in cells 6 days after transduction with lentiviruses carrying shRNAs directed against p65 or a scrambled control (shScr) (data represent mean of n = 3 dishes, data from 1 additional independent experiment shown in Source Data Table). (c) Several shRNAs against p65 (vs. scrambled shRNA: shScr) were delivered to mature adipocytes via lentivirus, and cells were then treated with Dex or TNF and assessed for insulin-stimulated glucose uptake. Shown is the percent of insulin-stimulated glucose uptake rescued by p65 knockdown (data represent mean of n = 6 dishes, data from 2 additional independent experiments shown in Source Data Table); ND = not determined because of cell death). (d) Expression of Hsd11b1 was assessed in mature 3T3-L1 adipocytes by Q-PCR after treatment with Dex and TNF for the indicated period of time (data represent mean of n = 3 dishes, data from 1 additional independent experiment shown in Source Data Table). (e) Carbenoxolone at the indicated dose was treated to fully differentiated adipocytes with or without Dex or TNF for 6 days and then assessed for insulin-stimulated glucose uptake (data represent mean of n = 6 dishes, data from 1 additional independent experiment shown in Source Data Table). (f) The effect of carbenoxolone (70 μM) was tested on glucocorticoid-dependent gene expression in mature 3T3-L1 adipocytes. Cortisol or cortisone (1 uM each) were used to induce expression of known targets Sgk1 and Dusp1; carbenoxolone blocks the effect of cortisone, which must be activated by 11-β-HSD1, but not cortisol, which does not require activation, (data represent mean of n = 3 dishes, data from 1 additional independent experiment shown in Source Data Table).

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