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Investigations on the metabolic role of the Wnt signaling pathway and hepatic transcription factor 7-like 2 (TCF7L2) have generated opposing views. While some studies demonstrated a repressive effect of TCF7L2 on hepatic gluconeogenesis, a recent study using liver-specific Tcf7l2(-/-) mice suggested the opposite. As a consequence of redundant and bidirectional actions of transcription factor (TCF) molecules and other complexities of the Wnt pathway, knockout of a single Wnt pathway component may not effectively reveal a complete metabolic picture of this pathway. To address this, we generated the liver-specific dominant-negative (DN) TCF7L2 (TCF7L2DN) transgenic mouse model LTCFDN. These mice exhibited progressive impairment in response to pyruvate challenge. Importantly, LTCFDN hepatocytes displayed elevated gluconeogenic gene expression, gluconeogenesis, and loss of Wnt-3a-mediated repression of gluconeogenesis. In C57BL/6 hepatocytes, adenovirus-mediated expression of TCF7L2DN, but not wild-type TCF7L2, increased gluconeogenesis and gluconeogenic gene expression. Our further mechanistic exploration suggests that TCF7L2DN-mediated inhibition of Wnt signaling causes preferential interaction of β-catenin (β-cat) with FoxO1 and increased binding of β-cat/FoxO1 to the Pck1 FoxO binding site, resulting in the stimulation of Pck1 expression and increased gluconeogenesis. Together, our results using TCF7L2DN as a unique tool revealed that the Wnt signaling pathway and its effector β-cat/TCF serve a beneficial role in suppressing hepatic gluconeogenesis., (© 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.)

p21-activated protein kinase-1 (Pak1) plays a role in insulin secretion and glucagon-like peptide-1 (GLP-1) production. Pak1(-/-) mice were found to carry a defect in ip pyruvate tolerance test (IPPTT), leading us to speculate whether Pak1 represses hepatic gluconeogenesis. We show here that the defect in IPPTT became more severe in aged Pak1(-/-) mice. In primary hepatocytes, 2,2'-dihydroxy-1,1'-dinaphthyldisulfide, a potent inhibitor of group I Paks, reduced basal glucose production (GP), attenuated forskolin- or glucagon-stimulated GP, and attenuated the stimulation of forskolin on the expression of Pck1 and G6pc. In addition, the capacity of primary hepatocytes isolated from Pak1(-/-) mice in GP at the basal level is significantly lower than that of the control littermates. These in vitro observations imply that the direct effect of Paks in hepatocytes is the stimulation of gluconeogenesis and that the impairment in IPPTT in Pak1(-/-) mice is due to the lack of Pak1 elsewhere. Consecutive ip injection of forskolin for 2 weeks increased gut proglucagon expression, associated with improved IPPTT in aged Pak1(-/-) mice and wild-type controls. In addition, administration of the DPP-IV (dipeptidyl peptidase-4) inhibitor sitagliptin for 1 week reversed the defect in IPPTT in aged Pak1(-/-) mice, associated with increased plasma GLP-1 levels. Our observations indicate a potential role of Pak1 in the gut/pancreas/liver axis in controlling glucose disposal and affirmed the therapeutic application of GLP-1 and DPP-IV inhibitors in attenuating hepatic gluconeogenesis.

The type 2 diabetes risk gene TCF7L2 is the effector of the Wnt signaling pathway. We found previously that in gut endocrine L-cell lines, TCF7L2 controls transcription of the proglucagon gene (gcg), which encodes the incretin hormone glucagon-like peptide-1 (GLP-1). Whereas peripheral GLP-1 stimulates insulin secretion, brain GLP-1 controls energy homeostasis through yet-to-be defined mechanisms. We aim to determine the metabolic effect of a functional knockdown of TCF7L2 by generating transgenic mice that express dominant-negative TCF7L2 (TCF7L2DN) specifically in gcg-expressing cells. The gcg-TCF7L2DN transgenic mice showed reduced gcg expression in their gut and brain, but not in pancreas. Defects in glucose homeostasis were observed in these mice, associated with attenuated plasma insulin levels in response to glucose challenge. The defect in glucose disposal was exacerbated with high-fat diet. Brain Wnt activity and feeding-mediated hypothalamic AMP-activated protein kinase (AMPK) repression in these mice were impaired. Peripheral injection of the cAMP-promoting agent forskolin increased brain β-cat Ser675 phosphorylation and brain gcg expression and restored feeding-mediated hypothalamic AMPK repression. We conclude that TCF7L2 and Wnt signaling control gut and brain gcg expression and glucose homeostasis and speculate that positive cross-talk between Wnt and GLP-1/cAMP signaling is an underlying mechanism for brain GLP-1 in exerting its metabolic functions.

In gut endocrine L cells, the Wnt signaling pathway effector β-catenin (β-cat)/transcription factor 7-like 2 mediates the stimulatory effect of insulin on proglucagon (gcg) expression and glucagon-like peptide-1 (GLP-1) production. In several other cell lineages, insulin is able to stimulate p21-activated protein kinase 1 (Pak1). Here we determined the role of Pak1 in gcg expression and the effect of Pak1 deletion on glucose homeostasis. Insulin stimulated Pak1 activation through increasing its Thr423 phosphorylation in gut gcg-expressing cell lines, associated with increased gcg mRNA levels. This stimulation was attenuated by the Pak inhibitor 2,2'-dihydroxy-1,1'-dinaphthyldisulfide (IPA3) or dominant-negative Pak1. Both insulin and cAMP-promoting agents activated β-cat Ser675 phosphorylation, which was attenuated by IPA3 or protein kinase A inhibition, respectively. Gut gcg levels were reduced in male Pak1(-/-) mice, associated with impaired glucose tolerance after an ip or oral glucose challenge. These mice had lower circulating active GLP-1 levels after a glucose challenge as well as reduced distal ileum GLP-1 content after insulin treatment. Finally, the Pak1(-/-) mice exhibited reduced brainstem gcg level and abolished β-cat Ser675 phosphorylation in brain neurons after insulin treatment. We suggest that Pak1 mediates the cross talk between insulin and Wnt signaling pathways on gut and brain gcg expression, and its ablation impairs glucose homeostasis.

Background: Dietary polyphenols including anthocyanins target multiple organs.Objective: We aimed to assess the involvement of glucagon-like peptide 1 (GLP-1), leptin, insulin and fibroblast growth factor 21 (FGF21) in mediating metabolic beneficial effects of purified anthocyanin cyanidin-3-glucoside (Cy3G).Methods: Intestinal proglucagon gene (Gcg; encoding GLP-1) and liver Fgf21 expression were assessed in 6-wk-old male C57BL-6J mice fed a low-fat-diet (LFD; 10% of energy from fat), alone or with 1.6 mg Cy3G/L in drinking water for 3 wk [experiment (Exp.) 1; n = 5/group]. Similar mice were fed the LFD or a high-fat diet (HFD; 60% energy from fat) with or without Cy3G for 20 wk. Half of the mice administered Cy3G also received 4 broad-spectrum antibiotics (ABs) in drinking water between weeks 11 and 14, for a total of 6 groups (n = 8/group). Metabolic tolerance tests were conducted between weeks 2 and 16. Relevant hormone gene expression and plasma hormone concentrations were assessed mainly at the end of 20 wk (Exp. 2).Results: In Exp. 1, Cy3G administration increased ileal but not colonic Gcg level by 2-fold (P < 0.05). In Exp. 2, Cy3G attenuated HFD-induced body-weight gain (20.3% at week 16), and improved glucose tolerance (26.5% at week 15) but not insulin tolerance. Although Cy3G had no effect on glucose tolerance in LFD mice, LFD/Cy3G/AB mice showed better glucose tolerance than LFD/Cy3G mice (23%). In contrast, HFD/Cy3G/AB mice showed worse glucose tolerance compared with HFD/Cy3G mice (15%). Beneficial effects of Cy3G in HFD mice were not associated with changes in plasma leptin, insulin or GLP-1 concentrations. However, Cy3G increased hepatic Fgf21 expression in mice in Exp. 1 by 4-fold and attenuated Fgf21 overexpression in HFD mice (Exp. 2, 22%), associated with increased expression of genes that encode FGFR1 and β-klotho (>3-fold, P < 0.05).Conclusions: Dietary Cy3G may reduce body weight and exert metabolic homeostatic effects in mice via changes in hepatic FGF21. [ABSTRACT FROM AUTHOR]

Abstract: Thioredoxin interacting protein (TxNIP) functions as an effector of glucotoxicity in pancreatic β-cells. Exendin-4 (Ex-4), a long-term effective GLP-1 receptor agonist, reduces TxNIP level in pancreatic β-cells. Mechanisms underlying this reduction, however, remain largely unknown. We show here that Ex-4, 8-bromo-cAMP, the cAMP promoting agent forskolin, as well as activators of protein kinase A (PKA) and exchange protein activated by cAMP (Epac), all attenuated the effect of high glucose (20mM) on TxNIP level in the pancreatic β-cell line Ins-1. Forskolin and Ex-4 also reduced TxNIP level in cultured primary rat islets. This repressive effect is at least partially mediated via stimulating proteasome-dependent TxNIP degradation, since the proteasomal inhibitor MG132, but not the lysosomal inhibitor chloroquine, significantly blocked the repressive effect of forskolin. Furthermore, forskolin enhanced TxNIP ubiquitination. Both PKA inhibition and Epac inhibition partially blocked the repressive effect of forskolin on TxNIP level. In addition, forskolin and Ex-4 protected Ins-1 cells from high glucose-induced apoptotic activity, assessed by measuring caspase 3 activity. Finally, knockdown of TxNIP expression led to reduced caspase 3 expression levels and blunted response to forskolin treatment. We suggest that proteasome-dependent TxNIP degradation is a novel mechanism by which Ex-4-cAMP signaling protects pancreatic β cells. [Copyright &y& Elsevier]