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. 2022 Feb 19;298(4):101748. doi: 10.1016/j.jbc.2022.101748

Figure 6.

Figure 6

Feeding-linked insulin activity directly upregulates TGF-β1 expression in adipocytes.A, Western blot image of TGF-β1 protein after 24 h of insulin treatment in 3T3-L1 adipocytes. B, relative gene expression of TGF-β1 in 3T3-L1 adipocytes after 48 h of insulin treatment with/without an AKT inhibitor (MK2206: 2 μM), mTOR inhibitor (rapamycin: 0.2 μM), or MEK inhibitor (U0126: 2 μM) (n = 4). C, relative gene expression of TGF-β1 in chicken adipose tissue in response to insulin neutralization (GSE35581, n = 4). D, relative gene expression of TGF-β1 in 3T3-L1 adipocytes treated with a SMAD3 inhibitor (SIS3 HCl: 10 μM) for 12 h with/without 2 ng/ml recombinant TGF-β1. E and F, transcriptome analysis of SMAD3 knockout mice (E) and TGF-β inhibitory antibody-injected mice (F) (GDS3985). G, SMAD3 ChIP-Seq peaks in human and mouse TGF-β1 gene regions (human SUM159 cells: GSM3736834; mouse embryonic stem cells: GSM3563733). The y-axes indicate RPM (reads per million mapped reads) units. H, relative gene expression of TGF-β1 and ECM remodeling genes in 3T3-L1 adipocytes after 18 h of TGFBR inhibitor treatment (SB431542: 10 μM), followed by 3 h of insulin responses (0.1 nM) (n = 3). I, relative gene expression of lipogenesis-related genes in 3T3-L1 adipocytes after 18 h of TGFBR inhibitor treatment (SB431542: 10 μM), followed by 3 h of insulin response (0.1 nM) (n = 3). Data are presented as the mean ± SEM. #p < 0.1, ∗p < 0.05, ∗∗p < 0.01. ChIP-Seq, chromatin immunoprecipitation sequencing; ECM, extracellular matrix; MEK, MAPK/ERK kinase; mTOR, mammalian target of rapamycin; SMAD3, SMAD family member 3; TGF-β1, transforming growth factor β1; TGFBR, transforming growth factor β receptor.