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E4orf1 induction in adipose tissue promotes insulin-independent signaling in the adipocyte.

Kusminski CM, Gallardo-Montejano VI, Wang ZV, Hegde V, Bickel PE, Dhurandhar NV, Scherer PE - Mol Metab (2015)

Bottom Line: At the whole body level, this leads to reduced body-weight gain under a high fat diet challenge.Nevertheless, they are protected from diet-induced hepatic steatosis.The resulting systemic phenotype is complex, yet highlights the powerful nature of manipulating selective branches of the insulin signaling network within the adipocyte.

View Article: PubMed Central - PubMed

Affiliation: Touchstone Diabetes Center, Department of Internal Medicine, The University of Texas Southwestern Medical Center, Dallas, TX, USA.

ABSTRACT

Background/purpose: Type 2 diabetes remains a worldwide epidemic with major pathophysiological changes as a result of chronic insulin resistance. Insulin regulates numerous biochemical pathways related to carbohydrate and lipid metabolism.

Methods: We have generated a novel mouse model that allows us to constitutively activate, in an inducible fashion, the distal branch of the insulin signaling transduction pathway specifically in adipocytes.

Results: Using the adenoviral 36 E4orf1 protein, we chronically stimulate locally the Ras-ERK-MAPK signaling pathway. At the whole body level, this leads to reduced body-weight gain under a high fat diet challenge. Despite overlapping glucose tolerance curves, there is a reduced requirement for insulin action under these conditions. The mice further exhibit reduced circulating adiponectin levels that ultimately lead to impaired lipid clearance, and inflamed and fibrotic white adipose tissues. Nevertheless, they are protected from diet-induced hepatic steatosis. As we observe constitutively elevated p-Akt levels in the adipocytes, even under conditions of low insulin levels, this pinpoints enhanced Ras-ERK-MAPK signaling in transgenic adipocytes as a potential alternative route to bypass proximal insulin signaling events.

Conclusion: We conclude that E4orf1 expression in the adipocyte leads to enhanced baseline activation of the distal insulin signaling node, yet impaired insulin receptor stimulation in the presence of insulin, with important implications for the regulation of adiponectin secretion. The resulting systemic phenotype is complex, yet highlights the powerful nature of manipulating selective branches of the insulin signaling network within the adipocyte.

No MeSH data available.


Related in: MedlinePlus

E4orf1-Tg mice exhibit lower body-weight gain, systemic insulin-sparing effects, inflamed and fibrotic WAT with minimal hepatic steatosis. (A) Body-weight gain (g) of male C57/Bl6 WT mice versus E4orf1-Tg mice during Dox-HFD (600 mg/kg) feeding, (n = 7 per group). (B) Glucose levels (left) and insulin levels (right) during an oral glucose tolerance test (OGTT) (2.5 g/kg body-weight glucose; single gavage) on male WT mice versus E4orf1-Tg mice, (n = 7 per group). (C) Triglyceride clearance test on WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding. Mice were gavaged (20 ul/g body-weight 20% Intra-lipid) following an overnight (14–16 h fast), (n = 7 per group). (D) Ad libitum and 24 h fasted systemic glucose, TG and adiponectin levels in WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD (600 mg/kg) feeding, (n = 7 per group). (E) Representative images of H&E staining of sWAT (top left) and gWAT (middle left) tissues from WT versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding. Representative images of perilipin immunohistochemical (IHC) staining of sWAT (bottom left) derived from WT versus E4orf1-Tg mice. Mac2 IHC staining (top right) and Trichrome staining (middle right) of sWAT, in addition to H&E staining of liver tissues (bottom right) from WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding. Scale bar: 32 μm. (F) Fat-pad (sWAT and gWAT) and liver tissue weights (mg) normalized to body-weight (g), in WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding, (n = 7 per group). (G) Hepatic TG content (mg/g) in livers derived from WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding, (n = 7 per group). (Student's t-test, *P < 0.05; **P < 0.01; ***P < 0.001).
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fig2: E4orf1-Tg mice exhibit lower body-weight gain, systemic insulin-sparing effects, inflamed and fibrotic WAT with minimal hepatic steatosis. (A) Body-weight gain (g) of male C57/Bl6 WT mice versus E4orf1-Tg mice during Dox-HFD (600 mg/kg) feeding, (n = 7 per group). (B) Glucose levels (left) and insulin levels (right) during an oral glucose tolerance test (OGTT) (2.5 g/kg body-weight glucose; single gavage) on male WT mice versus E4orf1-Tg mice, (n = 7 per group). (C) Triglyceride clearance test on WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding. Mice were gavaged (20 ul/g body-weight 20% Intra-lipid) following an overnight (14–16 h fast), (n = 7 per group). (D) Ad libitum and 24 h fasted systemic glucose, TG and adiponectin levels in WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD (600 mg/kg) feeding, (n = 7 per group). (E) Representative images of H&E staining of sWAT (top left) and gWAT (middle left) tissues from WT versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding. Representative images of perilipin immunohistochemical (IHC) staining of sWAT (bottom left) derived from WT versus E4orf1-Tg mice. Mac2 IHC staining (top right) and Trichrome staining (middle right) of sWAT, in addition to H&E staining of liver tissues (bottom right) from WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding. Scale bar: 32 μm. (F) Fat-pad (sWAT and gWAT) and liver tissue weights (mg) normalized to body-weight (g), in WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding, (n = 7 per group). (G) Hepatic TG content (mg/g) in livers derived from WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding, (n = 7 per group). (Student's t-test, *P < 0.05; **P < 0.01; ***P < 0.001).

Mentions: To apply a metabolic challenge, we fed E4orf1-Tg mice Dox high-fat diet (Dox-HFD) (600 mg/kg) and monitored body-weights. Figure 2A shows that E4orf1-Tg mice exhibit significantly lower body-weight gain during Dox-HFD feeding. Following six weeks Dox-HFD feeding, we performed an oral glucose tolerance test (OGTT) and observed no marked differences in glucose tolerance between WT mice and E4orf1-Tg mice (Figure 2B), although baseline acute fasting (3 h) glucose levels were significantly lower in E4orf1-Tg mice compared to WT littermates (Figure 2B), suggesting possible protection from HFD-induced fasting hyperglycemia. Interestingly, E4orf1-Tg mice exhibit a profoundly lower transient insulin spike in response to the glucose load during the OGTT, in comparison to WT mice. Insulin levels were significantly lower both at baseline and throughout the duration of the OGTT (Figure 2B). We ruled out any pancreatic β-cell defects in E4orf1-Tg mice, given their normal capacity to clear a glucose bolus from circulation. In vitro studies have documented that E4orf1 enhances cellular glucose uptake in the absence of insulin, either by bypassing the insulin receptor or by sustaining activation of downstream insulin signaling components in the face of an insulin receptor knockdown [20–22], suggesting potential unique ‘insulin sparing’ actions of E4orf1-mediated signaling. Here, given that E4orf1-Tg mice clear exogenous glucose efficiently while not harboring any β-cell defects, yet exhibit severely lower insulin levels in doing so, our data suggest that E4orf1-Tg mice may have a lower requirement for insulin. They effectively preserve whole-body glucose clearance capacity and the distal branch of the insulin signaling cascade, with a lower requirement for insulin. We further assessed lipid clearance efficacy and observed that E4orf1-Tg mice exhibit impaired triglyceride (TG) clearance, evidenced by significantly higher systemic TG levels following an oral lipid gavage (Figure 2C). This suggests that E4orf1-Tg mice harbor an unfavorable systemic lipid profile. We subsequently measured various systemic parameters under ad libitum and fasted (24 h) conditions. While E4orf1-Tg mice weigh markedly less than their WT counterparts, no differences in the fasting-induced body-weight changes were apparent following a 24 h fast (data not shown). Secondly, E4orf1-Tg mice exhibit significantly lower fasting blood glucose levels (Figure 2D). Such fasting-induced hypoglycemia is consistent with in vitro studies demonstrating that E4orf1 confers anti-hyperglycemic actions by improving cellular glucose disposal [20,23]. Here, such actions of E4orf1 appear to be more effective under fasted conditions. In line with the impaired TG clearance capacity, we observed markedly higher circulating TG levels and FFA levels in E4orf1-Tg mice, when compared with WT mice (Figure 2D), implicating some degree of fasting-induced lipolysis present in metabolically challenged E4orf1-Tg mice. Similar to Dox-chow fed conditions, both in the Dox-HFD-fed-state and during fasting, E4orf1-Tg mice exhibit lower systemic adiponectin levels in comparison to their WT littermates (Figure 2D). Given that it has been established that circulating adiponectin levels are lower in states of obesity and T2DM [29–31], the reduction in adiponectin in E4orf1-Tg mice that exhibit a leaner phenotype in comparison to their WT counterparts suggests impaired production of adiponectin from WAT and further implicates that E4orf1 induction may promote WAT dysfunction and metabolically ‘unhealthy’ WAT expansion.


E4orf1 induction in adipose tissue promotes insulin-independent signaling in the adipocyte.

Kusminski CM, Gallardo-Montejano VI, Wang ZV, Hegde V, Bickel PE, Dhurandhar NV, Scherer PE - Mol Metab (2015)

E4orf1-Tg mice exhibit lower body-weight gain, systemic insulin-sparing effects, inflamed and fibrotic WAT with minimal hepatic steatosis. (A) Body-weight gain (g) of male C57/Bl6 WT mice versus E4orf1-Tg mice during Dox-HFD (600 mg/kg) feeding, (n = 7 per group). (B) Glucose levels (left) and insulin levels (right) during an oral glucose tolerance test (OGTT) (2.5 g/kg body-weight glucose; single gavage) on male WT mice versus E4orf1-Tg mice, (n = 7 per group). (C) Triglyceride clearance test on WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding. Mice were gavaged (20 ul/g body-weight 20% Intra-lipid) following an overnight (14–16 h fast), (n = 7 per group). (D) Ad libitum and 24 h fasted systemic glucose, TG and adiponectin levels in WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD (600 mg/kg) feeding, (n = 7 per group). (E) Representative images of H&E staining of sWAT (top left) and gWAT (middle left) tissues from WT versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding. Representative images of perilipin immunohistochemical (IHC) staining of sWAT (bottom left) derived from WT versus E4orf1-Tg mice. Mac2 IHC staining (top right) and Trichrome staining (middle right) of sWAT, in addition to H&E staining of liver tissues (bottom right) from WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding. Scale bar: 32 μm. (F) Fat-pad (sWAT and gWAT) and liver tissue weights (mg) normalized to body-weight (g), in WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding, (n = 7 per group). (G) Hepatic TG content (mg/g) in livers derived from WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding, (n = 7 per group). (Student's t-test, *P < 0.05; **P < 0.01; ***P < 0.001).
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fig2: E4orf1-Tg mice exhibit lower body-weight gain, systemic insulin-sparing effects, inflamed and fibrotic WAT with minimal hepatic steatosis. (A) Body-weight gain (g) of male C57/Bl6 WT mice versus E4orf1-Tg mice during Dox-HFD (600 mg/kg) feeding, (n = 7 per group). (B) Glucose levels (left) and insulin levels (right) during an oral glucose tolerance test (OGTT) (2.5 g/kg body-weight glucose; single gavage) on male WT mice versus E4orf1-Tg mice, (n = 7 per group). (C) Triglyceride clearance test on WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding. Mice were gavaged (20 ul/g body-weight 20% Intra-lipid) following an overnight (14–16 h fast), (n = 7 per group). (D) Ad libitum and 24 h fasted systemic glucose, TG and adiponectin levels in WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD (600 mg/kg) feeding, (n = 7 per group). (E) Representative images of H&E staining of sWAT (top left) and gWAT (middle left) tissues from WT versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding. Representative images of perilipin immunohistochemical (IHC) staining of sWAT (bottom left) derived from WT versus E4orf1-Tg mice. Mac2 IHC staining (top right) and Trichrome staining (middle right) of sWAT, in addition to H&E staining of liver tissues (bottom right) from WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding. Scale bar: 32 μm. (F) Fat-pad (sWAT and gWAT) and liver tissue weights (mg) normalized to body-weight (g), in WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding, (n = 7 per group). (G) Hepatic TG content (mg/g) in livers derived from WT mice versus E4orf1-Tg mice post eight-weeks Dox-HFD feeding, (n = 7 per group). (Student's t-test, *P < 0.05; **P < 0.01; ***P < 0.001).
Mentions: To apply a metabolic challenge, we fed E4orf1-Tg mice Dox high-fat diet (Dox-HFD) (600 mg/kg) and monitored body-weights. Figure 2A shows that E4orf1-Tg mice exhibit significantly lower body-weight gain during Dox-HFD feeding. Following six weeks Dox-HFD feeding, we performed an oral glucose tolerance test (OGTT) and observed no marked differences in glucose tolerance between WT mice and E4orf1-Tg mice (Figure 2B), although baseline acute fasting (3 h) glucose levels were significantly lower in E4orf1-Tg mice compared to WT littermates (Figure 2B), suggesting possible protection from HFD-induced fasting hyperglycemia. Interestingly, E4orf1-Tg mice exhibit a profoundly lower transient insulin spike in response to the glucose load during the OGTT, in comparison to WT mice. Insulin levels were significantly lower both at baseline and throughout the duration of the OGTT (Figure 2B). We ruled out any pancreatic β-cell defects in E4orf1-Tg mice, given their normal capacity to clear a glucose bolus from circulation. In vitro studies have documented that E4orf1 enhances cellular glucose uptake in the absence of insulin, either by bypassing the insulin receptor or by sustaining activation of downstream insulin signaling components in the face of an insulin receptor knockdown [20–22], suggesting potential unique ‘insulin sparing’ actions of E4orf1-mediated signaling. Here, given that E4orf1-Tg mice clear exogenous glucose efficiently while not harboring any β-cell defects, yet exhibit severely lower insulin levels in doing so, our data suggest that E4orf1-Tg mice may have a lower requirement for insulin. They effectively preserve whole-body glucose clearance capacity and the distal branch of the insulin signaling cascade, with a lower requirement for insulin. We further assessed lipid clearance efficacy and observed that E4orf1-Tg mice exhibit impaired triglyceride (TG) clearance, evidenced by significantly higher systemic TG levels following an oral lipid gavage (Figure 2C). This suggests that E4orf1-Tg mice harbor an unfavorable systemic lipid profile. We subsequently measured various systemic parameters under ad libitum and fasted (24 h) conditions. While E4orf1-Tg mice weigh markedly less than their WT counterparts, no differences in the fasting-induced body-weight changes were apparent following a 24 h fast (data not shown). Secondly, E4orf1-Tg mice exhibit significantly lower fasting blood glucose levels (Figure 2D). Such fasting-induced hypoglycemia is consistent with in vitro studies demonstrating that E4orf1 confers anti-hyperglycemic actions by improving cellular glucose disposal [20,23]. Here, such actions of E4orf1 appear to be more effective under fasted conditions. In line with the impaired TG clearance capacity, we observed markedly higher circulating TG levels and FFA levels in E4orf1-Tg mice, when compared with WT mice (Figure 2D), implicating some degree of fasting-induced lipolysis present in metabolically challenged E4orf1-Tg mice. Similar to Dox-chow fed conditions, both in the Dox-HFD-fed-state and during fasting, E4orf1-Tg mice exhibit lower systemic adiponectin levels in comparison to their WT littermates (Figure 2D). Given that it has been established that circulating adiponectin levels are lower in states of obesity and T2DM [29–31], the reduction in adiponectin in E4orf1-Tg mice that exhibit a leaner phenotype in comparison to their WT counterparts suggests impaired production of adiponectin from WAT and further implicates that E4orf1 induction may promote WAT dysfunction and metabolically ‘unhealthy’ WAT expansion.

Bottom Line: At the whole body level, this leads to reduced body-weight gain under a high fat diet challenge.Nevertheless, they are protected from diet-induced hepatic steatosis.The resulting systemic phenotype is complex, yet highlights the powerful nature of manipulating selective branches of the insulin signaling network within the adipocyte.

View Article: PubMed Central - PubMed

Affiliation: Touchstone Diabetes Center, Department of Internal Medicine, The University of Texas Southwestern Medical Center, Dallas, TX, USA.

ABSTRACT

Background/purpose: Type 2 diabetes remains a worldwide epidemic with major pathophysiological changes as a result of chronic insulin resistance. Insulin regulates numerous biochemical pathways related to carbohydrate and lipid metabolism.

Methods: We have generated a novel mouse model that allows us to constitutively activate, in an inducible fashion, the distal branch of the insulin signaling transduction pathway specifically in adipocytes.

Results: Using the adenoviral 36 E4orf1 protein, we chronically stimulate locally the Ras-ERK-MAPK signaling pathway. At the whole body level, this leads to reduced body-weight gain under a high fat diet challenge. Despite overlapping glucose tolerance curves, there is a reduced requirement for insulin action under these conditions. The mice further exhibit reduced circulating adiponectin levels that ultimately lead to impaired lipid clearance, and inflamed and fibrotic white adipose tissues. Nevertheless, they are protected from diet-induced hepatic steatosis. As we observe constitutively elevated p-Akt levels in the adipocytes, even under conditions of low insulin levels, this pinpoints enhanced Ras-ERK-MAPK signaling in transgenic adipocytes as a potential alternative route to bypass proximal insulin signaling events.

Conclusion: We conclude that E4orf1 expression in the adipocyte leads to enhanced baseline activation of the distal insulin signaling node, yet impaired insulin receptor stimulation in the presence of insulin, with important implications for the regulation of adiponectin secretion. The resulting systemic phenotype is complex, yet highlights the powerful nature of manipulating selective branches of the insulin signaling network within the adipocyte.

No MeSH data available.


Related in: MedlinePlus