Palmitic acid up regulates Gal‑3 and induces insulin resistance in macrophages by mediating the balance between KLF4 and NF‑κB

Figures

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  Figure 1. Induction of THP-1 differentiation into macrophages by PMA. (A) Results of hematoxylin and eosin staining. The left image of each group was obtained at a magnification of x100 (left-hand image scale bars, 200 µm). The images on right were obtained at a magnification of x400 (right-hand image scale bars 50 µm). (B) The morphology of THP-1 cells under the indicate treatments were observed under phase-contrast microscopy. (C) Images of microsphere phagocytosis assay. (D) Phagocytic rate of macrophages based on (C). PMA, phorbol 12-myristate 13-acetate. *P<0.05 vs. THP-1.

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  Figure 2. PA-induced insulin resistance downregulates the expression of KLF4 in macrophages. (A) 2-deoxy-2-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]-D-glucose uptake detection. (B) Representative western blots and (C) semi-quantification of the protein expression levels of insulin resistance-related proteins. (D) Representative western blots and (E) semi-quantification of KLF4 protein expression levels. ELISA detection of the levels of (F) IL-6, (G) IL-1β and (H) TNF-α following treatment with different concentrations of PA. *P<0.05 and **P<0.01 vs. control. IL, interleukin; INSR, insulin receptor; IRS1, insulin receptor substrate 1; KLF4, Krüppel-like factor 4; p, phosphorylated; PA, palmitic acid; TNF-α, tumor necrosis factor α.

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  Figure 3. PA treatment promotes the expression of Gal-3. (A) Representative western blots and (B) semi-quantification of the protein expression levels of Gal-3 in different tissue samples. (C) Representative western blots and (D) semi-quantification of the protein expression levels of Gal-3 in THP-1 macrophages treated with different concentrations of PA. (E) Reverse transcription-quantitative PCR detection of Gal-3 mRNA expression levels. (F) MTT detection of the effect of different concentrations of PA on cell viability. *P<0.05 and **P<0.01 vs. control. Gal-3, galectin-3; PA, palmitic acid; T2DM, type 2 diabetes mellitus.

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  Figure 4. Gal-3 promotes insulin resistance of macrophages. Macrophages were treated with PA, the Gal-3 inhibitor GB1107 or both. (A) 2-deoxy-2- [(7-nitro-2,1,3-benzoxadiazol- 4-yl) amino]-D-glucose uptake test results. (B) Representative western blots and (C) semi-quantitation of Gal-3 protein expression levels. (D) Immunofluorescence results of Gal-3. (E) Histogram shown the relative fluorescence density of Gal-3. **P<0.01 vs. control group. ##P<0.01 as indicated. Gal-3, galectin-3; PA, palmitic acid.

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  Figure 5. PA promotes the activation of the TLR4/NF-κB pathway by downregulating KLF4 and promoting Gal-3 expression. Macrophages were treated with PA, TAK-242 and both. (A) Representative western blots and (B) semi-quantification of protein expression levels. (C) Main protein interaction map. **P<0.01 vs. control. Gal-3, galectin-3; KLF4, Krüppel-like factor 4; LGALS3, galectin-3; NF-κB, nuclear factor-κB; p, phosphorylated; PA, palmitic acid; TLR4, toll-like receptor 4; IFNA1, Interferon α 1; NOX4, NADPH οxidase 4.