Pancreas-derived DPP4 is not essential for glucose homeostasis under metabolic stress.

Mice systemically lacking dipeptidyl peptidase-4 (DPP4) have improved islet health, glucoregulation, and reduced obesity with high-fat diet (HFD) feeding compared to wild-type mice. Some, but not all, of this improvement can be linked to the loss of DPP4 in endothelial cells (ECs), pointing to the contribution of non-EC types. The importance of intra-islet signaling mediated by α to ß cell communication is becoming increasingly clear; thus, our objective was to determine if ß cell DPP4 regulates insulin secretion and glucose tolerance in HFD-fed mice by regulating the local concentrations of insulinotropic peptides. Using ß cell double incretin receptor knockout mice, ß cell- and pancreas-specific Dpp4-/- mice, we reveal that ß cell incretin receptors are necessary for DPP4 inhibitor effects. However, although ß cell DPP4 modestly contributes to high glucose (16.7 mM)-stimulated insulin secretion in isolated islets, it does not regulate whole-body glucose homeostasis.

Glucagon-Like Peptide-1 Receptor Agonists in Adult Patients With Type 2 Diabetes: Review of Cardiovascular Outcome Trials.

People with type 2 diabetes are at heightened risk for developing cardiovascular (CV) events. CV disease is the leading cause of premature death among adults with type 2 diabetes. Unfortunately, historically, some antidiabetes agents were implicated in worsening CV function, despite improving glycemic and metabolic control. Accordingly, over a decade ago, health regulatory bodies modified approval requirements for novel antidiabetes pharmacotherapies, requiring prospective evaluation of CV safety through cardiovascular outcome trials (CVOTs). To meet regulatory requirements, CVOTs were primarily designed around establishing CV safety by demonstrating noninferiority to placebo in addition to standard of care, without significant differences in blood glucose. If appropriately designed and powered, however, these CVOTs could also determine superiority, and hence CV protection. Although many of these CVOTs were initiated several years ago, the recent reporting of the results for these CVOTs has been pivotal and practice-changing. Glucagon-like peptide-1 receptor agonists (GLP-1RAs) are one such class of antidiabetes therapies, wherein multiple GLP-1RA CVOTs, but interestingly, not all, have demonstrated CV benefits. In this review, we provide a comprehensive summary of all the reported CVOTs completed with GLP-1RAs to date. Although it remains unclear why some GLP-1RAs are associated with reducing CV events, whereas others have been consistent with CV safety alone, we highlight and provide an overview of some key differences between the various GLP-1RAs and their respective CVOTs and possible implications of study design differences. We also speculate on potential mechanisms of action for glucagon-like peptide-1 receptor signalling in the CV system.

Loss of Glp2r signaling activates hepatic stellate cells and exacerbates diet-induced steatohepatitis in mice.

A glucagon-like peptide-2 (GLP-2) analog is used in individuals with intestinal failure who are at risk for liver disease, yet the hepatic actions of GLP-2 are not understood. Treatment of high-fat diet-fed (HFD-fed) mice with GLP-2 did not modify the development of hepatosteatosis or hepatic inflammation. In contrast, Glp2r-/- mice exhibited increased hepatic lipid accumulation, deterioration in glucose tolerance, and upregulation of biomarkers of hepatic inflammation. Both mouse and human liver expressed the canonical GLP-2 receptor (GLP-2R), and hepatic Glp2r expression was upregulated in mice with hepatosteatosis. Cell fractionation localized the Glp2r to hepatic stellate cells (HSCs), and markers of HSC activation and fibrosis were increased in livers of Glp2r-/- mice. Moreover, GLP-2 directly modulated gene expression in isolated HSCs ex vivo. Taken together, these findings define an essential role for the GLP-2R in hepatic adaptation to nutrient excess and unveil a gut hormone-HSC axis, linking GLP-2R signaling to control of HSC activation.

Hematopoietic cell- versus enterocyte-derived dipeptidyl peptidase-4 differentially regulates triglyceride excursion in mice.

Postprandial triglycerides (TGs) are elevated in people with type 2 diabetes (T2D). Glucose-lowering agents, such as glucagon-like peptide-1 (GLP-1) receptor agonists and dipeptidyl peptidase-4 (DPP-4) inhibitors, also reduce postprandial TG excursion. Although the glucose-lowering mechanisms of DPP-4 have been extensively studied, how the reduction of DPP-4 activity improves lipid tolerance remains unclear. Here, we demonstrate that gut-selective and systemic inhibition of DPP-4 activity reduces postprandial TG excursion in young mice. Genetic inactivation of Dpp4 simultaneously within endothelial cells and hematopoietic cells using Tie2-Cre reduced intestinal lipoprotein secretion under regular chow diet conditions. Bone marrow transplantation revealed a key role for hematopoietic cells in modulation of lipid responses arising from genetic reduction of DPP-4 activity. Unexpectedly, deletion of Dpp4 in enterocytes increased TG excursion in high-fat diet-fed (HFD-fed) mice. Moreover, chemical reduction of DPP-4 activity and increased levels of GLP-1 were uncoupled from TG excursion in older or HFD-fed mice, yet lipid tolerance remained improved in older Dpp4-/- and Dpp4EC-/- mice. Taken together, this study defines roles for specific DPP-4 compartments, age, and diet as modifiers of DPP-4 activity linked to control of gut lipid metabolism.

Plasma levels of DPP4 activity and sDPP4 are dissociated from inflammation in mice and humans.

Dipeptidyl peptidase-4 (DPP4) modulates inflammation by enzymatic cleavage of immunoregulatory peptides and through its soluble form (sDPP4) that directly engages immune cells. Here we examine whether reduction of DPP4 activity alters inflammation. Prolonged DPP4 inhibition increases plasma levels of sDPP4, and induces sDPP4 expression in lymphocyte-enriched organs in mice. Bone marrow transplantation experiments identify hematopoietic cells as the predominant source of plasma sDPP4 following catalytic DPP4 inhibition. Surprisingly, systemic DPP4 inhibition increases plasma levels of inflammatory markers in regular chow-fed but not in high fat-fed mice. Plasma levels of sDPP4 and biomarkers of inflammation are lower in metformin-treated subjects with type 2 diabetes (T2D) and cardiovascular disease, yet exhibit considerable inter-individual variation. Sitagliptin therapy for 12 months reduces DPP4 activity yet does not increase markers of inflammation or levels of sDPP4. Collectively our findings dissociate levels of DPP4 enzyme activity, sDPP4 and biomarkers of inflammation in mice and humans.

The gut hormone receptor GIPR links energy availability to the control of hematopoiesis.

OBJECTIVE: Glucose-dependent insulinotropic polypeptide (GIP) conveys information from ingested nutrients to peripheral tissues, signaling energy availability. The GIP Receptor (GIPR) is also expressed in the bone marrow, notably in cells of the myeloid lineage. However, the importance of gain and loss of GIPR signaling for diverse hematopoietic responses remains unclear. METHODS: We assessed the expression of the Gipr in bone marrow (BM) lineages and examined functional roles for the GIPR in control of hematopoiesis. Bone marrow responses were studied in (i) mice fed regular or energy-rich diets, (ii) mice treated with hematopoietic stressors including acute 5-fluorouracil (5-FU), pamsaccharide (LPS), and Pam3CysSerLys4 (Pam3CSK4), with or without pharmacological administration of a GIPR agonist, and (iii) mice with global (Gipr-/-) or selective deletion of the GIPR (GiprTie2-/-) with and without bone marrow transplantation (BMT). RESULTS: Gipr is expressed within T cells, myeloid cells, and myeloid precursors; however, these cell populations were not different in peripheral blood, spleen, or BM of Gipr-/- and GiprTie2-/- mice. Nevertheless, gain and loss of function studies revealed that GIPR signaling controls the expression of BM Toll-like receptor (TLR) and Notch-related genes regulating hematopoiesis. Loss of the BM GIPR attenuates the extent of adipose tissue inflammation and dysregulates the hematopoietic response to BMT. GIPR agonism modified BM gene expression profiles following 5-FU and Pam3CSK4 whereas loss of the Gipr altered the hematopoietic responses to energy excess, two TLR ligands, and 5-FU. However, the magnitude of the cellular changes in hematopoiesis in response to gain or loss of GIPR signaling was relatively modest. CONCLUSION: These studies identify a functional gut hormone-BM axis positioned for the transduction of signals linking nutrient availability to the control of TLR and Notch genes regulating hematopoiesis. Nevertheless, stimulation or loss of GIPR signaling has minimal impact on basal hematopoiesis or the physiological response to hematopoietic stress.

Distinct Neural Sites of GLP-1R Expression Mediate Physiological versus Pharmacological Control of Incretin Action.

Glucagon-like peptide 1 (GLP-1) receptors are widely distributed throughout the nervous system, enabling physiological and pharmacological control of glucose and energy homeostasis. Here we elucidated the importance of Glp1r expression within cellular domains targeted by expression of Wnt1-Cre2 or Phox2b-Cre. Widespread loss of neural Glp1r in Glp1rΔWnt1-/- mice had no effect on basal food intake, gastric emptying, and glucose homeostasis. However, the glucoregulatory actions of GLP-1R agonists, but not gut-selective DPP-4 inhibition, were preserved in Glp1rΔWnt1-/- mice. Unexpectedly, selective reduction of Glp1r expression within neurons targeted by Phox2b-Cre impaired glucose homeostasis and gastric emptying and attenuated the extent of weight loss achieved with sustained GLP-1R agonism. Collectively, these studies identify discrete neural domains of Glp1r expression mediating GLP-1-regulated control of metabolism and the gut-brain axis and reveal the unexpected importance of neuronal Phox2b+ cells expressing GLP-1R for physiological regulation of gastric emptying, islet hormone responses, and glucose homeostasis.

Physiological roles of the GIP receptor in murine brown adipose tissue.

OBJECTIVE: Glucose-dependent insulinotropic polypeptide (GIP) is secreted from the gut in response to nutrient ingestion and promotes meal-dependent insulin secretion and lipid metabolism. Loss or attenuation of GIP receptor (GIPR) action leads to resistance to diet-induced obesity through incompletely understood mechanisms. The GIPR is expressed in white adipose tissue; however, its putative role in brown adipose tissue (BAT) has not been explored. METHODS: We investigated the role of the GIPR in BAT cells in vitro and in BAT-specific (GiprBAT-/-) knockout mice with selective elimination of the Gipr within the Myf5+ expression domain. We analyzed body weight, adiposity, glucose homeostasis, insulin and lipid tolerance, energy expenditure, food intake, body temperature, and iBAT oxygen consumption ex vivo. High-fat diet (HFD)-fed GiprBAT-/- mice were studied at room temperature (21 °C), 4 °C, and 30 °C ambient temperatures. RESULTS: The mouse Gipr gene is expressed in BAT, and GIP directly increased Il6 mRNA and IL-6 secretion in BAT cells. Additionally, levels of thermogenic, lipid and inflammation mRNA transcripts were altered in BAT cells transfected with Gipr siRNA. Body weight gain, energy expenditure, and glucose and insulin tolerance were normal in HFD-fed GiprBAT-/- mice housed at room temperature. However, GiprBAT-/- mice exhibited higher body temperatures during an acute cold challenge and a lower respiratory exchange ratio and impaired lipid tolerance at 21 °C. In contrast, body weight was lower and iBAT oxygen consumption was higher in HFD-fed mice housed at 4 °C but not at 30 °C. CONCLUSIONS: The BAT GIPR is linked to the control of metabolic gene expression, fuel utilization, and oxygen consumption. However, the selective loss of the GIPR within BAT is insufficient to recapitulate the findings of decreased weight gain and resistance to obesity arising in experimental models with systemic disruption of GIP action.

Fibroblast activation protein is dispensable for control of glucose homeostasis and body weight in mice.

OBJECTIVE: Fibroblast Activation Protein (FAP), an enzyme structurally related to dipeptidyl peptidase-4 (DPP-4), has garnered interest as a potential metabolic drug target due to its ability to cleave and inactivate FGF-21 as well as other peptide substrates. Here we investigated the metabolic importance of FAP for control of body weight and glucose homeostasis in regular chow-fed and high fat diet-fed mice. METHODS: FAP enzyme activity was transiently attenuated using a highly-specific inhibitor CPD60 and permanently ablated by genetic inactivation of the mouse Fap gene. We also assessed the FAP-dependence of CPD60 and talabostat (Val-boroPro), a chemical inhibitor reportedly targeting both FAP and dipeptidyl peptidase-4 RESULTS: CPD60 robustly inhibited plasma FAP activity with no effect on DPP-4 activity. Fap gene disruption was confirmed by assessment of genomic DNA, and loss of FAP enzyme activity in plasma and tissues. CPD60 did not improve lipid tolerance but modestly improved acute oral and intraperitoneal glucose tolerance in a FAP-dependent manner. Genetic inactivation of Fap did not improve glucose or lipid tolerance nor confer resistance to weight gain in male or female Fap-/- mice fed regular chow or high-fat diets. Moreover, talabostat markedly improved glucose homeostasis in a FAP- and FGF-21-independent, DPP-4 dependent manner. CONCLUSION: Although pharmacological FAP inhibition improves glucose tolerance, the absence of a metabolic phenotype in Fap-/-mice suggest that endogenous FAP is dispensable for the regulation of murine glucose homeostasis and body weight. These findings highlight the importance of characterizing the specificity and actions of FAP inhibitors in different species and raise important questions about the feasibility of mouse models for targeting FAP as a treatment for diabetes and related metabolic disorders.

The brown adipose tissue glucagon receptor is functional but not essential for control of energy homeostasis in mice.

OBJECTIVE: Administration of glucagon (GCG) or GCG-containing co-agonists reduces body weight and increases energy expenditure. These actions appear to be transduced by multiple direct and indirect GCG receptor (GCGR)-dependent mechanisms. Although the canonical GCGR is expressed in brown adipose tissue (BAT) the importance of BAT GCGR activity for the physiological control of body weight, or the response to GCG agonism, has not been defined. METHODS: We studied the mechanisms linking GCG action to acute increases in oxygen consumption using wildtype (WT), Ucp1-/- and Fgf21-/- mice. The importance of basal GCGR expression within the Myf5+ domain for control of body weight, adiposity, glucose and lipid metabolism, food intake, and energy expenditure was examined in GcgrBAT-/- mice housed at room temperature or 4 °C, fed a regular chow diet (RCD) or after a prolonged exposure to high fat diet (HFD). RESULTS: Acute GCG administration induced lipolysis and increased the expression of thermogenic genes in BAT cells, whereas knockdown of Gcgr reduced expression of genes related to thermogenesis. GCG increased energy expenditure (measured by oxygen consumption) both in vivo in WT mice and ex vivo in BAT and liver explants. GCG also increased acute energy expenditure in Ucp1-/- mice, but these actions were partially blunted in Ffg21-/- mice. However, acute GCG administration also robustly increased oxygen consumption in GcgrBAT-/- mice. Moreover, body weight, glycemia, lipid metabolism, body temperature, food intake, activity, energy expenditure and adipose tissue gene expression profiles were normal in GcgrBAT-/- mice, either on RCD or HFD, whether studied at room temperature, or chronically housed at 4 °C. CONCLUSIONS: Exogenous GCG increases oxygen consumption in mice, also evident both in liver and BAT explants ex vivo, through UCP1-independent, FGF21-dependent pathways. Nevertheless, GCGR signaling within BAT is not physiologically essential for control of body weight, whole body energy expenditure, glucose homeostasis, or the adaptive metabolic response to cold or prolonged exposure to an energy dense diet.

Circulating Levels of Soluble Dipeptidyl Peptidase-4 Are Dissociated from Inflammation and Induced by Enzymatic DPP4 Inhibition.

Dipeptidyl peptidase-4 (DPP-4) controls glucose homeostasis through enzymatic termination of incretin action. We report that plasma DPP-4 activity correlates with body weight and fat mass, but not glucose control, in mice. Genetic disruption of adipocyte Dpp4 expression reduced plasma DPP-4 activity in older mice but did not perturb incretin levels or glucose homeostasis. Knockdown of hepatocyte Dpp4 completely abrogated the obesity-associated increase in plasma DPP-4 activity, reduced liver cytokine expression, and partially attenuated inflammation in adipose tissue without changes in incretin levels or glucose homeostasis. In contrast, circulating levels of soluble DPP4 (sDPP4) were dissociated from inflammation in mice with endothelial-selective or global genetic inactivation of Dpp4. Remarkably, inhibition of DPP-4 enzymatic activity upregulated circulating levels of sDPP4 originating from endothelial or hematopoietic cells without inducing systemic or localized inflammation. Collectively, these findings reveal unexpected complexity in regulation of soluble versus enzymatic DPP-4 and control of inflammation and glucose homeostasis.

Cellular Sites and Mechanisms Linking Reduction of Dipeptidyl Peptidase-4 Activity to Control of Incretin Hormone Action and Glucose Homeostasis.

Pharmacological inhibition of the dipeptidyl peptidase-4 (DPP4) enzyme potentiates incretin action and is widely used to treat type 2 diabetes. Nevertheless, the precise cells and tissues critical for incretin degradation and glucose homeostasis remain unknown. Here, we use mouse genetics and pharmacologic DPP4 inhibition to identify DPP4+ cell types essential for incretin action. Although enterocyte DPP4 accounted for substantial intestinal DPP4 activity, ablation of enterocyte DPP4 in Dpp4Gut-/- mice did not produce alterations in plasma DPP4 activity, incretin hormone levels, and glucose tolerance. In contrast, endothelial cell (EC)-derived DPP4 contributed substantially to levels of soluble plasma DPP4 activity, incretin degradation, and glucose control. Surprisingly, DPP4+ cells of bone marrow origin mediated the selective degradation of fasting GIP, but not GLP-1. Collectively, these findings identify distinct roles for DPP4 in the EC versus the bone marrow compartment for selective incretin degradation and DPP4i-mediated glucoregulation.

Inhibition of Dipeptidyl Peptidase-4 Impairs Ventricular Function and Promotes Cardiac Fibrosis in High Fat-Fed Diabetic Mice.

Dipeptidyl peptidase-4 (DPP4) inhibitors used for the treatment of type 2 diabetes are cardioprotective in preclinical studies; however, some cardiovascular outcome studies revealed increased hospitalization rates for heart failure (HF) among a subset of DPP4 inhibitor-treated subjects with diabetes. We evaluated cardiovascular function in young euglycemic Dpp4(-/-) mice and in older, high fat-fed, diabetic C57BL/6J mice treated with either the glucagon-like peptide 1 receptor (GLP-1R) agonist liraglutide or the highly selective DPP4 inhibitor MK-0626. We assessed glucose metabolism, ventricular function and remodeling, and cardiac gene expression profiles linked to inflammation and fibrosis after transverse aortic constriction (TAC) surgery, a pressure-volume overload model of HF. Young euglycemic Dpp4(-/-) mice exhibited a cardioprotective response after TAC surgery or doxorubicin administration, with reduced fibrosis; however, cardiac mRNA analysis revealed increased expression of inflammation-related transcripts. Older, diabetic, high fat-fed mice treated with the GLP-1R agonist liraglutide exhibited preservation of cardiac function. In contrast, diabetic mice treated with MK-0626 exhibited modest cardiac hypertrophy, impairment of cardiac function, and dysregulated expression of genes and proteins controlling inflammation and cardiac fibrosis. These findings provide a model for the analysis of mechanisms linking fibrosis, inflammation, and impaired ventricular function to DPP4 inhibition in preclinical studies.