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Endothelial NO/cGMP/VASP signaling attenuates Kupffer cell activation and hepatic insulin resistance induced by high-fat feeding.

Tateya S, Rizzo NO, Handa P, Cheng AM, Morgan-Stevenson V, Daum G, Clowes AW, Morton GJ, Schwartz MW, Kim F - Diabetes (2011)

Bottom Line: We sought to determine whether reduced endothelial nitric oxide (NO) signaling contributes to the effect of high-fat feeding to increase hepatic inflammatory signaling and if so, whether this effect 1) involves activation of Kupffer cells and 2) is ameliorated by increased NO signaling.Targeted deletion of vasodilator-stimulated phosphoprotein (VASP), a key downstream target of endothelially derived NO, similarly predisposes to hepatic and Kupffer cell inflammation and abrogates the protective effect of NO signaling in both macrophages and hepatocytes studied in a cell culture model.Our findings also identify the NO/VASP pathway as a novel potential target for the treatment of obesity-associated liver insulin resistance.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of Washington, Seattle, Washington, USA.

ABSTRACT

Objective: Proinflammatory activation of Kupffer cells is implicated in the effect of high-fat feeding to cause liver insulin resistance. We sought to determine whether reduced endothelial nitric oxide (NO) signaling contributes to the effect of high-fat feeding to increase hepatic inflammatory signaling and if so, whether this effect 1) involves activation of Kupffer cells and 2) is ameliorated by increased NO signaling.

Research design and methods: Effect of NO/cGMP signaling on hepatic inflammation and on isolated Kupffer cells was examined in C57BL/6 mice, eNos(-/-) mice, and Vasp(-/-) mice fed a low-fat or high-fat diet.

Results: We show that high-fat feeding induces proinflammatory activation of Kupffer cells in wild-type mice coincident with reduced liver endothelial nitric oxide synthase activity and NO content while, conversely, enhancement of signaling downstream of endogenous NO by phosphodiesterase-5 inhibition protects against high fat-induced inflammation in Kupffer cells. Furthermore, proinflammatory activation of Kupffer cells is evident in eNos(-/-) mice even on a low-fat diet. Targeted deletion of vasodilator-stimulated phosphoprotein (VASP), a key downstream target of endothelially derived NO, similarly predisposes to hepatic and Kupffer cell inflammation and abrogates the protective effect of NO signaling in both macrophages and hepatocytes studied in a cell culture model.

Conclusions: These results collectively imply a physiological role for endothelial NO to limit obesity-associated inflammation and insulin resistance in hepatocytes and support a model in which Kupffer cell activation during high-fat feeding is dependent on reduced NO signaling. Our findings also identify the NO/VASP pathway as a novel potential target for the treatment of obesity-associated liver insulin resistance.

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Effect of VASP deficiency on hepatic and Kupffer cell inflammation and hepatic insulin signaling. A: Hepatic PKG expression and phosphorylation of VASP Ser 239 from wild-type (WT) mice fed either a low-fat (LF) or high-fat (HF) diet for 4 weeks from age 8 weeks (n = 5). B–E: Vasp−/− mice and littermate control mice were fed an LF diet for 4 weeks. B: Body weight (BW) (g). C: Body composition (%). Liver lysates were analyzed for IκB-α phosphorylation by Western blot (D), and IL-6 mRNA expression was measured by RT-PCR (E) (n = 8). *P < 0.05. F: Kupffer cells were isolated from Vasp−/− mice or WT control mice on an LF diet for 4 weeks. Inflammatory markers as measured by RT-PCR (n = 5). *P < 0.05. G and H: In parallel experiments in Vasp−/− (n = 7) and littermate control (n = 9) mice, hepatic insulin (Ins) signaling was assessed following intraperitoneal injection of insulin. IRS-1 and IRS-2 tyrosine phosphorylation and Akt serine phosphorylation. *P < 0.05. I: Hepatic triglyceride (TG) from Vasp−/− and control (n = 5). *P < 0.05. Veh, vehicle. IB, immunoblot; kD, kilodalton.
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Figure 4: Effect of VASP deficiency on hepatic and Kupffer cell inflammation and hepatic insulin signaling. A: Hepatic PKG expression and phosphorylation of VASP Ser 239 from wild-type (WT) mice fed either a low-fat (LF) or high-fat (HF) diet for 4 weeks from age 8 weeks (n = 5). B–E: Vasp−/− mice and littermate control mice were fed an LF diet for 4 weeks. B: Body weight (BW) (g). C: Body composition (%). Liver lysates were analyzed for IκB-α phosphorylation by Western blot (D), and IL-6 mRNA expression was measured by RT-PCR (E) (n = 8). *P < 0.05. F: Kupffer cells were isolated from Vasp−/− mice or WT control mice on an LF diet for 4 weeks. Inflammatory markers as measured by RT-PCR (n = 5). *P < 0.05. G and H: In parallel experiments in Vasp−/− (n = 7) and littermate control (n = 9) mice, hepatic insulin (Ins) signaling was assessed following intraperitoneal injection of insulin. IRS-1 and IRS-2 tyrosine phosphorylation and Akt serine phosphorylation. *P < 0.05. I: Hepatic triglyceride (TG) from Vasp−/− and control (n = 5). *P < 0.05. Veh, vehicle. IB, immunoblot; kD, kilodalton.

Mentions: In tissues such as vascular smooth muscle cells and platelets, NO/cGMP signaling activates PKG, which in turn phosphorylates VASP on Ser 239. To determine whether high-fat feeding is associated with changes in PKG or phospho-VASP (Ser 239), we performed Western blot analysis on liver lysates from wild-type mice fed either a low-fat or high-fat diet for 8 weeks. As expected, we found that high-fat feeding is associated with reduced liver phospho-VASP levels, whereas total PKG protein levels were not significantly altered (Fig. 4A). This finding suggests that in mouse liver, the effect of high-fat feeding to reduce endothelial NO signaling decreases the basal level of VASP activation—an effect that in turn could contribute to the proinflammatory response to reduced endothelial NO signaling.


Endothelial NO/cGMP/VASP signaling attenuates Kupffer cell activation and hepatic insulin resistance induced by high-fat feeding.

Tateya S, Rizzo NO, Handa P, Cheng AM, Morgan-Stevenson V, Daum G, Clowes AW, Morton GJ, Schwartz MW, Kim F - Diabetes (2011)

Effect of VASP deficiency on hepatic and Kupffer cell inflammation and hepatic insulin signaling. A: Hepatic PKG expression and phosphorylation of VASP Ser 239 from wild-type (WT) mice fed either a low-fat (LF) or high-fat (HF) diet for 4 weeks from age 8 weeks (n = 5). B–E: Vasp−/− mice and littermate control mice were fed an LF diet for 4 weeks. B: Body weight (BW) (g). C: Body composition (%). Liver lysates were analyzed for IκB-α phosphorylation by Western blot (D), and IL-6 mRNA expression was measured by RT-PCR (E) (n = 8). *P < 0.05. F: Kupffer cells were isolated from Vasp−/− mice or WT control mice on an LF diet for 4 weeks. Inflammatory markers as measured by RT-PCR (n = 5). *P < 0.05. G and H: In parallel experiments in Vasp−/− (n = 7) and littermate control (n = 9) mice, hepatic insulin (Ins) signaling was assessed following intraperitoneal injection of insulin. IRS-1 and IRS-2 tyrosine phosphorylation and Akt serine phosphorylation. *P < 0.05. I: Hepatic triglyceride (TG) from Vasp−/− and control (n = 5). *P < 0.05. Veh, vehicle. IB, immunoblot; kD, kilodalton.
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Related In: Results  -  Collection

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Figure 4: Effect of VASP deficiency on hepatic and Kupffer cell inflammation and hepatic insulin signaling. A: Hepatic PKG expression and phosphorylation of VASP Ser 239 from wild-type (WT) mice fed either a low-fat (LF) or high-fat (HF) diet for 4 weeks from age 8 weeks (n = 5). B–E: Vasp−/− mice and littermate control mice were fed an LF diet for 4 weeks. B: Body weight (BW) (g). C: Body composition (%). Liver lysates were analyzed for IκB-α phosphorylation by Western blot (D), and IL-6 mRNA expression was measured by RT-PCR (E) (n = 8). *P < 0.05. F: Kupffer cells were isolated from Vasp−/− mice or WT control mice on an LF diet for 4 weeks. Inflammatory markers as measured by RT-PCR (n = 5). *P < 0.05. G and H: In parallel experiments in Vasp−/− (n = 7) and littermate control (n = 9) mice, hepatic insulin (Ins) signaling was assessed following intraperitoneal injection of insulin. IRS-1 and IRS-2 tyrosine phosphorylation and Akt serine phosphorylation. *P < 0.05. I: Hepatic triglyceride (TG) from Vasp−/− and control (n = 5). *P < 0.05. Veh, vehicle. IB, immunoblot; kD, kilodalton.
Mentions: In tissues such as vascular smooth muscle cells and platelets, NO/cGMP signaling activates PKG, which in turn phosphorylates VASP on Ser 239. To determine whether high-fat feeding is associated with changes in PKG or phospho-VASP (Ser 239), we performed Western blot analysis on liver lysates from wild-type mice fed either a low-fat or high-fat diet for 8 weeks. As expected, we found that high-fat feeding is associated with reduced liver phospho-VASP levels, whereas total PKG protein levels were not significantly altered (Fig. 4A). This finding suggests that in mouse liver, the effect of high-fat feeding to reduce endothelial NO signaling decreases the basal level of VASP activation—an effect that in turn could contribute to the proinflammatory response to reduced endothelial NO signaling.

Bottom Line: We sought to determine whether reduced endothelial nitric oxide (NO) signaling contributes to the effect of high-fat feeding to increase hepatic inflammatory signaling and if so, whether this effect 1) involves activation of Kupffer cells and 2) is ameliorated by increased NO signaling.Targeted deletion of vasodilator-stimulated phosphoprotein (VASP), a key downstream target of endothelially derived NO, similarly predisposes to hepatic and Kupffer cell inflammation and abrogates the protective effect of NO signaling in both macrophages and hepatocytes studied in a cell culture model.Our findings also identify the NO/VASP pathway as a novel potential target for the treatment of obesity-associated liver insulin resistance.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, University of Washington, Seattle, Washington, USA.

ABSTRACT

Objective: Proinflammatory activation of Kupffer cells is implicated in the effect of high-fat feeding to cause liver insulin resistance. We sought to determine whether reduced endothelial nitric oxide (NO) signaling contributes to the effect of high-fat feeding to increase hepatic inflammatory signaling and if so, whether this effect 1) involves activation of Kupffer cells and 2) is ameliorated by increased NO signaling.

Research design and methods: Effect of NO/cGMP signaling on hepatic inflammation and on isolated Kupffer cells was examined in C57BL/6 mice, eNos(-/-) mice, and Vasp(-/-) mice fed a low-fat or high-fat diet.

Results: We show that high-fat feeding induces proinflammatory activation of Kupffer cells in wild-type mice coincident with reduced liver endothelial nitric oxide synthase activity and NO content while, conversely, enhancement of signaling downstream of endogenous NO by phosphodiesterase-5 inhibition protects against high fat-induced inflammation in Kupffer cells. Furthermore, proinflammatory activation of Kupffer cells is evident in eNos(-/-) mice even on a low-fat diet. Targeted deletion of vasodilator-stimulated phosphoprotein (VASP), a key downstream target of endothelially derived NO, similarly predisposes to hepatic and Kupffer cell inflammation and abrogates the protective effect of NO signaling in both macrophages and hepatocytes studied in a cell culture model.

Conclusions: These results collectively imply a physiological role for endothelial NO to limit obesity-associated inflammation and insulin resistance in hepatocytes and support a model in which Kupffer cell activation during high-fat feeding is dependent on reduced NO signaling. Our findings also identify the NO/VASP pathway as a novel potential target for the treatment of obesity-associated liver insulin resistance.

Show MeSH
Related in: MedlinePlus