Treatment with SRT1720, a SIRT1 activator, ameliorates fatty liver with reduced expression of lipogenic enzymes in MSG mice.

Nonalcoholic fatty liver disease (NAFLD) is an abnormal liver metabolism often observed with insulin resistance and metabolic syndrome. Calorie restriction is a useful treatment for NAFLD and reportedly prolongs the life spans of several species in which sirtuin plays an important role. In this study, we examined whether the activation of SIRT1, a mammalian ortholog of sirtuin, may ameliorate the development of NAFLD. Monosodium glutamate (MSG) mice, which exhibited obesity and insulin resistance, were treated with SRT1720, a specific SIRT1 activator from the age of 6-16 wk. Sixteen-week-old MSG mice exhibited increased liver triglyceride content and elevated levels of aminotransferase. SRT1720 treatment significantly reduced these levels without affecting body weight or food intake. These results suggested that the administration of SRT1720 ameliorated the development of NAFLD in MSG mice. The expressions of lipogenic genes, such as sterol regulatory element-binding protein-1c, acetyl-CoA carboxylase, and fatty acid synthase, and the serum lipid profiles, including free fatty acids, were elevated in MSG mice and were reduced by SRT1720 treatment. SRT1720 treatment also reduced the expressions of lipogenic genes in cultured HepG2 cells. Furthermore, SRT1720 treatment decreased the expressions of marker genes for oxidative stress and inflammatory cytokines in the liver of MSG mice. Taken together, SRT1720 treatment may reduce liver lipid accumulation, at least in part, by directly reducing the expressions of lipogenic genes. The reduction of oxidative stress and inflammation may also be involved in the amelioration of NAFLD.

Effects of pioglitazone on suppressor of cytokine signaling 3 expression: potential mechanisms for its effects on insulin sensitivity and adiponectin expression.

Pioglitazone is widely used for the treatment of diabetic patients with insulin resistance. The mechanism of pioglitazone to improve insulin sensitivity is not fully understood. Recent studies have shown that the induction of suppressor of cytokine signaling 3 (SOCS3) is related to the development of insulin resistance. Here, we examined whether the insulin-sensitizing effect of pioglitazone affects the SOCS induction. In db/db mice and high-fat-fed mice, expression of SOCS3 mRNA in fat tissue was increased compared with that in lean control mice, and pioglitazone suppressed SOCS3 levels. In 3T3-L1 adipocytes, mediators of insulin resistance such as tumor necrosis factor-alpha (TNF-alpha), interleukin-6, growth hormone, and insulin increased SOCS3 expression, which was partially inhibited by pioglitazone. The ability of pioglitazone to suppress SOCS3 induction by TNF-alpha was greatly augmented by peroxisome proliferator-activated receptor gamma overexpression. SOCS3 overexpression and tyrphostin AG490, a Janus kinase 2 inhibitor, or dominant-negative STAT3 expression partially inhibited adiponectin secretion and was accompanied by decreased STAT3 phosphorylation. Conversely, pioglitazone increased adiponectin secretion and STAT3 phosphorylation in fat tissue of db/db mice and in 3T3-L1 adipocytes. These results suggest that pioglitazone exerts its effect to improve whole-body insulin sensitivity in part through the suppression of SOCS3, which is associated with the increase in STAT3 phosphorylation and adiponectin production in fat tissue.

Sirt1 activator induces proangiogenic genes in preadipocytes to rescue insulin resistance in diet-induced obese mice.

Sirt1 plays an important role in regulating glucose and lipid metabolism in obese animal models. Impaired adipose tissue angiogenesis in the obese state decreases adipogenesis and thereby contributes to glucose intolerance and lipid metabolism. However, the mechanism by which Sirt1 activation affects obesity-associated impairments in angiogenesis in the adipose tissue is not fully understood. Here, we show that SRT1720 treatment induces angiogenic genes in cultured 3T3-L1 preadipocytes and ex vivo preadipocytes. siRNA-mediated knockdown of Sirt1 in 3T3-L1 preadipocytes downregulated angiogenic genes in the preadipocytes. SRT1720 treatment upregulated metabolically favorable genes and reduced inflammatory gene expressions in the adipose tissue of diet-induced obese (DIO) mice. Collectively, these findings suggest a novel role of SRT1720-induced Sirt1 activation in the induction of angiogenic genes in preadipocytes, thereby reducing inflammation and fibrosis in white adipose tissue (WAT) and promoting insulin sensitivity.

Publisher Correction: Sirt1 activator induces proangiogenic genes in preadipocytes to rescue insulin resistance in diet-induced obese mice.

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Chronic tumor necrosis factor-alpha treatment causes insulin resistance via insulin receptor substrate-1 serine phosphorylation and suppressor of cytokine signaling-3 induction in 3T3-L1 adipocytes.

Serine phosphorylation of insulin receptor substrate (IRS)-1 and the induction of suppressor of cytokine signaling 3 (SOCS3) is recently well documented as the mechanisms for the insulin resistance. However, the relationship between these two mechanisms is not fully understood. In this study, we investigated the involvement of SOCS3 and IRS-1 serine phosphorylation in TNFalpha-induced insulin resistance in 3T3-L1 adipocytes. TNFalpha transiently stimulated serine phosphorylation of IRS-1 from 10 min to 1 h, whereas insulin-stimulated IRS-1 tyrosine phosphorylation was inhibited only after TNFalpha treatment longer than 4 h. These results suggest that serine phosphorylation of IRS-1 alone is not the major mechanism for the inhibited insulin signaling by TNFalpha. TNFalpha stimulation longer than 4 h enhanced the expression of SOCS3 and signal transducer and activator of transcription-3 phosphorylation, concomitantly with the production of IL-6. Anti-IL-6 neutralizing antibody ameliorated suppressed insulin signaling by 24 h TNFalpha treatment, when it partially decreased SOCS3 induction and signal transducer and activator of transcription-3 phosphorylation. These results suggest that SOCS3 induction is involved in inhibited insulin signaling by TNFalpha. However, low-level expression of SOCS3 by IL-6 or adenovirus vector did not affect insulin-stimulated IRS-1 tyrosine phosphorylation. Interestingly, when IRS-1 serine phosphorylation was enhanced by TNFalpha or anisomycin in the presence of low-level SOCS3, IRS-1 degradation was remarkably enhanced. Taken together, both IRS-1 serine phosphorylation and SOCS3 induction are necessary, but one of the pair is not sufficient for the inhibited insulin signaling. Chronic TNFalpha may inhibit insulin signaling effectively because it causes both IRS-1 serine phosphorylation and the following SOCS3 induction in 3T3-L1 adipocytes.

Interleukin-1alpha inhibits insulin signaling with phosphorylating insulin receptor substrate-1 on serine residues in 3T3-L1 adipocytes.

Proinflammatory cytokines are recently reported to inhibit insulin signaling causing insulin resistance. IL-1alpha is also one of the proinflammatory cytokines; however, it has not been clarified whether IL-1alpha may also cause insulin resistance. Here, we investigated the effects of IL-1alpha treatment on insulin signaling in 3T3-L1 adipocytes. IL-1alpha treatment up to 4 h did not alter insulin-stimulated insulin receptor tyrosine phosphorylation, whereas tyrosine phosphorylation of insulin receptor substrate (IRS)-1 and the association with phosphatidylinositol 3-kinase were partially inhibited with the maximal inhibition in around 15 min. IRS-1 was transiently phosphorylated on some serine residues around 15 min after IL-1alpha stimulation, when several serine kinases, IkappaB kinase, c-Jun-N-terminal kinase, ERK, and p70S6K were activated. Chemical inhibitors for these kinases inhibited IL-1alpha-induced serine phosphorylation of IRS-1. Tyrosine phosphorylation of IRS-1 was recovered only by the IKK inhibitor or JNK inhibitor, suggesting specific involvement of these two kinases. Insulin-stimulated Akt phosphorylation and 2-deoxyglucose uptake were not inhibited only by IL-1alpha. Interestingly, Akt phosphorylation was synergistically inhibited by IL-1alpha in the presence of IL-6. Taken together, short-term IL-1alpha treatment transiently causes insulin resistance at IRS-1 level with its serine phosphorylation. IL-1alpha may suppress insulin signaling downstream of IRS-1 in the presence of other cytokines, such as IL-6.

Telmisartan improves insulin resistance and modulates adipose tissue macrophage polarization in high-fat-fed mice.

Diet-induced obesity is reported to induce a phenotypic switch in adipose tissue macrophages from an antiinflammatory M2 state to a proinflammatory M1 state. Telmisartan, an angiotensin II type 1 receptor blocker and a peroxisome proliferator-activated receptor-γ agonist, reportedly has more beneficial effects on insulin sensitivity than other angiotensin II type 1 receptor blockers. In this study, we studied the effects of telmisartan on the adipose tissue macrophage phenotype in high-fat-fed mice. Telmisartan was administered for 5 wk to high-fat-fed C57BL/6 mice. Insulin sensitivity, macrophage infiltration, and the gene expressions of M1 and M2 markers in visceral adipose tissues were then examined. An insulin- or a glucose-tolerance test showed that telmisartan treatment improved insulin resistance, decreasing the body weight gain, visceral fat weight, and adipocyte size without affecting the amount of energy intake. Telmisartan reduced the mRNA expression of CD11c and TNF-α, M1 macrophage markers, and significantly increased the expressions of M2 markers, such as CD163, CD209, and macrophage galactose N-acetyl-galactosamine specific lectin (Mgl2), in a quantitative RT-PCR analysis. A flow cytometry analysis showed that telmisartan decreased the number of M1 macrophages in visceral adipose tissues. In conclusion, telmisartan improves insulin sensitivity and modulates adipose tissue macrophage polarization to an antiinflammatory M2 state in high-fat-fed mice.

Regulatory mechanisms for adipose tissue M1 and M2 macrophages in diet-induced obese mice.

OBJECTIVE: To characterize the phenotypic changes of adipose tissue macrophages (ATMs) under different conditions of insulin sensitivity. RESEARCH DESIGN AND METHODS: The number and the expressions of marker genes for M1 and M2 macrophages from mouse epididymal fat tissue were analyzed using flow cytometry after the mice had been subjected to a high-fat diet (HFD) and pioglitazone treatment. RESULTS: Most of the CD11c-positive M1 macrophages and the CD206-positive M2 macrophages in the epididymal fat tissue were clearly separated using flow cytometry. The M1 and M2 macrophages exhibited completely different gene expression patterns. Not only the numbers of M1 ATMs and the expression of M1 marker genes, such as tumor necrosis factor-alpha and monocyte chemoattractant protein-1, but also the M1-to-M2 ratio were increased by an HFD and decreased by subsequent pioglitazone treatment, suggesting the correlation with whole-body insulin sensitivity. We also found that the increased number of M2 ATMs after an HFD was associated with the upregulated expression of interleukin (IL)-10, an anti-inflammatory Th2 cytokine, in the adipocyte fraction as well as in adipose tissue. The systemic overexpression of IL-10 by an adenovirus vector increased the expression of M2 markers in adipose tissue. CONCLUSIONS: M1 and M2 ATMs constitute different subsets of macrophages. Insulin resistance is associated with both the number of M1 macrophages and the M1-to-M2 ratio. The increased expression of IL-10 after an HFD might be involved in the increased recruitment of M2 macrophages.

Angiotensin II enhances the increase in monocyte chemoattractant protein-1 production induced by tumor necrosis factor-{alpha} from 3T3-L1 preadipocytes.

Monocyte chemoattractant protein-1 (MCP-1) and angiotensin II (Ang II) in adipose tissue are thought to induce systemic insulin resistance in rodents; but the precise mechanism is not fully clarified. We examined the mechanism of Ang II-induced and/or tumor necrosis factor-alpha (TNF-alpha)-induced MCP-1 production from 3T3-L1 preadipocytes. The MCP-1 protein and MCP-1 mRNA expression in 3T3-L1 preadipocytes were increased significantly by stimulation with TNF-alpha. We found no significant increase in MCP-1 concentrations by Ang II alone; but it enhanced the TNF-alpha-induced MCP-1 mRNA expression in a dose-dependent manner. Then, we examined the effect of Ang II and/or TNF-alpha on phosphorylation of extracellular signal-regulated kinase (ERK), p38MAPK, and IkappaB-alpha. Ang II and TNF-alpha clearly enhanced ERK and p38MAPK phosphorylation. IkappaB-alpha phosphorylation was enhanced by TNF-alpha, but not by Ang II. The MCP-1 mRNA expression induced by TNF-alpha and co-stimulation with Ang II was inhibited by either ERK inhibitor, p38MAPK inhibitor or NF-kappaB inhibitor. Moreover, Ang II enhanced the activation of AP-1 (c-fos) induced by TNF-alpha. Our results suggest that Ang II may serve as an additional stimulus on the TNF-alpha-induced MCP-1 production through the ERK-and p38MAPK-dependent pathways probably due to AP-1 activation.

Isoproterenol suppresses cytokine-induced RANTES secretion in human lung epithelial cells through the inhibition of c-jun N-terminal kinase pathway.

It has been reported that beta2-agonists may potentially exert some anti-inflammatory action in addition to bronchodilation that may contribute to their beneficial effects on asthma control. Bronchial epithelial cells are well known to respond to a range of stimuli by producing various biologically active mediators that can influence airway inflammation. RANTES (regulated on activation, normal T cells expressed and secreted) plays an important role in the pathophysiology of airway inflammation of asthmatics through its chemotactic activity for eosinophils. In this study, the authors investigated whether cytokine-induced RANTES release from BEAS-2B human bronchial epithelial cells could be modulated by beta-agonist isoproterenol (ISO). The possible involvement of c-jun N-terminal kinase (JNK) pathway was also studied. Combination of tumor necrosis factor-alpha and interleukin-1beta (cytokine mix) increased RANTES release from BEAS-2B cells and stimulated JNK activity. Similar to JNK inhibitor SP600125, ISO inhibited not only the production of RANTES but also the activation of JNK pathway in cytokine mix-stimulated BEAS-2B cells. The effect of ISO was mediated by the beta2-adrenoceptor, since it was blocked by ICI 118,551, a selective beta2-receptor antagonist, but not by atenolol, a selective beta1-receptor antagonist. Adenylyl cyclase activator forskolin reproduced the effects of ISO. Isoproterenol was found to inhibit the release of RANTES from the human bronchial epithelial cells, at least in part, through the inhibition of JNK signaling pathway.