• Acylated apelin-13 amide analogues exhibit enzyme resistance and prolonged insulin releasing, glucose lowering and anorexic properties

      O'Harte, Finbarr P. M.; Parthsarathy, Vadivel; Hogg, Christopher; Flatt, Peter R. (Elsevier, 2017-10-04)
      The adipokine, apelin has many biological functions but its activity is curtailed by rapid plasma degradation. Fatty acid derived apelin analogues represent a new and exciting avenue for the treatment of obesity-diabetes. This study explores four novel fatty acid modified apelin-13 analogues, namely, (Lys8GluPAL)apelin-13 amide, pGlu(Lys8GluPAL)apelin-13 amide, Lys8GluPAL(Tyr13)apelin-13 and Lys8GluPAL(Val13)apelin-13. Fatty acid modification extended the half-life of native apelin-13 to >24 h in vitro. pGlu(Lys8GluPAL)apelin-13 amide was the most potent insulinotropic analogue in BRIN-BD11 cells and isolated islets with maximal stimulatory effects of up to 2.7-fold (p < .001). (Lys8GluPAL)apelin-13 amide (1.9-fold) and Lys8GluPAL(Tyr13)apelin-13 (1.7-fold) were less effective, whereas Lys8GluPAL(Val13)apelin-13 had an inhibitory effect on insulin secretion. Similarly, pGlu(Lys8GluPAL)apelin-13 amide was most potent in increasing beta-cell intracellular Ca2+ concentrations (1.8-fold, p < .001) and increasing glucose uptake in 3T3-L1 adipocytes (2.3-fold, p < .01). Persistent biological action was observed with both pGlu(Lys8GluPAL)apelin-13 amide and (Lys8GluPAL)apelin-13 amide significantly reducing blood glucose (39–43%, p < .01) and enhancing insulin secretion (43–56%, p < .001) during glucose tolerance tests in diet-induced obese mice. pGlu(Lys8GluPAL)apelin-13 amide and (Lys8GluPAL)apelin-13 amide also inhibited feeding (28–40%, p < .001), whereas Lys8GluPAL(Val13)apelin-13 increased food intake (8%, p < .05) in mice. These data indicate that novel enzymatically stable analogues of apelin-13 may be suitable for future development as therapeutic agents for obesity-diabetes.
    • Apelin-13 analogues show potent in vitro and in vivo insulinotropic and glucose lowering actions

      O'Harte, Finbarr P. M.; Parthsarathy, Vadivel; Hogg, Christopher; Flatt, Peter R. (Elsevier, 2018-02-03)
      Nine structurally modified apelin-13 analogues were assessed for their in vitro and acute in vivo antidiabetic potential. Stability was assessed in mouse plasma and insulinotropic efficacy tested in cultured pancreatic BRIN-BD11 cells and isolated mouse pancreatic islets. Intracellular Ca2+ and cAMP production in BRIN-BD11 cells was determined, as was glucose uptake in 3T3-L1 adipocytes. Acute antihyperglycemic effects of apelin analogues were assessed following i.p. glucose tolerance tests (ipGGT, 18 mmol/kg) in normal and diet-induced-obese (DIO) mice and on food intake in normal mice. Apelin analogues all showed enhanced in vitro stability (up to 5.8-fold, t½ = 12.8 h) in mouse plasma compared to native apelin-13 (t½ = 2.1 h). Compared to glucose controls, stable analogues exhibited enhanced insulinotropic responses from BRIN-BD11 cells (up to 4.7-fold, p < 0.001) and isolated mouse islets (up to 5.3-fold) for 10−7 M apelin-13 amide (versus 7.6-fold for 10−7 M GLP-1). Activation of APJ receptors on BRIN-BD11 cells increased intracellular Ca2+ (up to 3.0-fold, p < 0.001) and cAMP (up to 1.7-fold, p < 0.01). Acute ipGTT showed improved insulinotropic and glucose disposal responses in normal and DIO mice (p < 0.05 and p < 0.01, respectively). Apelin-13 amide and (pGlu)apelin-13 amide were the most effective analogues exhibiting acute, dose-dependent and persistent biological actions. Both analogues stimulated insulin-independent glucose uptake by differentiated adipocytes (2.9 to –3.3-fold, p < 0.05) and inhibited food intake (26-–33%, p < 0.001), up to 180 min in mice, versus saline. In contrast, (Ala13)apelin-13 and (Val13)apelin-13 inhibited insulin secretion, suppressed beta-cell signal transduction and stimulated food intake in mice. Thus, stable analogues of apelin-13 have potential for diabetes/obesity therapy.
    • Atlantic salmon (Salmo salar) co-product-derived protein hydrolysates: A source of antidiabetic peptides

      Harnedy, Pàdraigín A.; Parthsarathy, Vadivel; McLaughlin, Chris M.; O'Keeffe, Martina B.; Allsopp, Philip J.; McSorley, Emeir M.; O'Harte, Finbarr P. M.; FitzGerald, Richard J. (Elsevier, 2018-02-06)
      Large quantities of low-value protein rich co-products, such as salmon skin and trimmings, are generated annually. These co-products can be upgraded to high-value functional ingredients. The aim of this study was to assess the antidiabetic potential of salmon skin gelatin and trimmingderived protein hydrolysates in vitro. The gelatin hydrolysate generated with Alcalase 2.4L and Flavourzyme 500L exhibited significantly higher (p<0.001) insulin and GLP-1 secretory activity from pancreatic BRIN-BD11 and enteroendocrine GLUTag cells, respectively, when tested at 2.5 mg/mL compared to hydrolysates generated with Alcalase 2.4L or Promod 144MG. The gelatin hydrolysate generated with Alcalase 2.4L and Flavourzyme 500L showed significantly more potent (p<0.01) DPP-IV inhibitory activity than those generated with Alcalase 2.4L or Promod 144MG. No significant difference was observed in the insulinotropic activity mediated by any of the trimming-derived hydrolysates when tested at 2.5 mg/mL. However, the trimmings hydrolysate generated with Alcalase 2.4L and Flavourzyme 500L exhibited significantly higher DPP-IV inhibitory (p<0.05:Alcalase 2.4L and p<0.01:Promod 144MG) and GLP-1 (p<0.001, 2.5 mg/mL) secretory activity than those generated with Alcalase 2.4L or Promod 144MG. The salmon trimmings hydrolysate generated with Alcalase 2.4L and Flavourzyme 500L when subjected to simulated gastrointestinal digestion (SGID) was shown to retain its GLP-1 secretory and DPP-IV inhibitory activities, in addition to improving its insulin secretory activity. However, the gelatin hydrolysate generated with Alcalase 2.4L and Flavourzyme 500L was shown to lose GLP-1 secretory activity following SGID. A significant increase in membrane potential (p<0.001) and intracellular calcium (p<0.001) by both co-product hydrolysates generated with Alcalase 2.4L and Flavourzyme 500L suggest that both hydrolysates mediate their insulinotropic activity through the KATP channel-dependent pathway. Additionally, by stimulating a significant increase in intracellular cAMP release (p<0.05) it is likely that the trimming-derived hydrolysate may also mediate insulin secretion through the protein kinase A pathway. The results presented herein demonstrate that salmon co-product hydrolysates exhibit promising in vitro antidiabetic activity.
    • Beneficial long-term antidiabetic actions of N- and C-terminally modified analogues of apelin-13 in diet-induced obese diabetic mice.

      Parthsarathy, Vadivel; Hogg, Christopher; Flatt, Peter R.; O'Harte, Finbarr P. M. (Wiley, 2017-07-20)
      AimsThis study investigated the chronic effects of twice daily administration of stable apelin analogues, apelin-13 amide and (pGlu)apelin-13 amide, on metabolic parameters in glucose intolerant and insulin resistant diet-induced obese (DIO) mice fed a high-fat diet for 150 days.Study Design & MethodsGroups of mice received twice daily (09:00 and 17:00 h) injections of saline vehicle, apelin-13 amide, (pGlu)apelin-13 amide or exendin-4(1–39) for 28 days (all at 25 nmol/kg). Energy intake, body weight, non-fasting blood glucose, plasma insulin, glucose tolerance, metabolic response to feeding and insulin sensitivity together with pancreatic hormone content and biochemical parameters such as lipids and total GLP-1 were monitored. Dual-energy X-ray absorptiometry (DXA) analysis and indirect calorimetry were also performed.ResultsAdministration of apelin-13 amide, (pGlu)apelin-13 amide or exendin-4 significantly decreased bodyweight, food intake, blood glucose and increased plasma insulin compared with high-fat fed saline treated controls (P < 0.05 and P < 0.001), Additionally, all peptide treated groups exhibited improved glucose tolerance (oral and ip), metabolic responses to feeding and associated insulin secretion. (pGlu)apelin-13 amide also significantly improved HbA1c and insulin sensitivity after 28 days. Both (pGlu)apelin-13 amide and exendin-4 increased bone mineral content and decreased respiratory exchange ratio (RER), whereas only (pGlu)apelin-13 amide increased energy expenditure. All treatment groups displayed reduced circulating triglycerides and increased GLP-1 concentrations, although only (pGlu)apelin-13 amide significantly reduced LDL-cholesterol, total body fat, and increased pancreatic insulin content.ConclusionThese data indicate the therapeutic potential of stable apelin-13 analogues with effects equivalent to or better than exendin-4.
    • Blue whiting (Micromesistius poutassou) muscle protein hydrolysate with in vitro and in vivo antidiabetic properties

      Harnedy, Pàdraigín A.; Parthsarathy, Vadivel; McLaughlin, Chris M.; O'Keeffe, Martina B.; Allsopp, Philip J.; McSorley, Emeir M.; O'Harte, Finbarr P. M.; Fitzgerald, Richard J. (Elsevier, 2017-10-29)
      A blue whiting (Micromesistius poutassou) protein hydrolysate generated using Alcalase 2.4L and Flavourzyme 500L and its simulated gastrointestinal digestion (SGID) sample was assessed for antidiabetic potential in vitro and in vivo. In addition to inhibiting dipeptidyl peptidase-IV (DPP–IV), the hydrolysates mediated insulin and glucagon-like peptide-1 (GLP-1) release from BRIN-BD11 and GLUTag cells, respectively. No significant difference was observed in insulinotropic and DPP-IV inhibitory activity following SGID, while GLP-1 secretion increased significantly (p < 0.01). SGID resulted in a significant increase in membrane potential, intracellular calcium and cyclic AMP concentration (p < 0.001) versus a glucose control, indicating that insulin secretion may be mediated by the KATP channel-dependent and the protein kinase A pathways. Additionally, acute (90–120min) and persistent (4h) glucose-lowering effects of the blue whiting hydrolysate were observed in normal healthy mice. These results demonstrate that the blue whiting protein hydrolysate had significant metabolic effects relevant to glucose control in vivo.