Abstract
Hypersecretion and alterations in the biological activity of the incretin hormone, glucose-dependent insulinotropic polypeptide (GIP), have been postulated as contributing factors in the development of obesity-related diabetes. However, recent studies also point to weight-reducing effects of GIP receptor activation. Therefore, generating precise experimental tools, such as specific and effective GIP receptor (GIPR) antagonists, is of key significance to better understand GIP physiology. Thus, the primary aim of the current study was to uncover improved GIPR antagonists for use in rodent studies, using human and mouse GIP sequences with N- and C-terminal deletions. Initial in vitro studies revealed that the GIPR agonists, human (h) GIP(1-42), hGIP(1-30) and mouse (m) GIP(1-30), stimulated (P < 0.01 to P < 0.001) insulin secretion from rat BRIN-BD11 cells. Analysis of insulin secretory effects of the N- and C-terminally cleaved GIP peptides, including hGIP(3-30), mGIP(3-30), h(Pro 3)GIP(3-30), hGIP(5-30), hGIP(3-42) and hGIP(5-42), revealed that these peptides did not modulate insulin secretion. More pertinently, only hGIP(3-30), mGIP(3-30) and h(Pro 3)GIP(3-30) were able to significantly (P < 0.01 to P < 0.001) inhibit hGIP(1-42)-stimulated insulin secretion. The human-derived GIPR agonist sequences, hGIP(1-42) and hGIP(1-30), reduced (P < 0.05) glucose levels in mice following conjoint injection with glucose, but mGIP(1-30) was ineffective. None of the N- and C-terminally cleaved GIP peptides affected glucose homeostasis when injected alone with glucose. However, hGIP(5-30) and mGIP(3-30) significantly (P < 0.05 to P < 0.01) impaired the glucose-lowering action of hGIP(1-42). Further evaluation of these most effective sequences demonstrated that mGIP(3-30), but not hGIP(5-30), effectively prevented GIP-induced elevations of plasma insulin concentrations. These data highlight, for the first time, that mGIP(3-30) represents an effective molecule to inhibit GIPR activity in mice.
| Original language | English |
|---|---|
| Pages (from-to) | 1-9 |
| Number of pages | 9 |
| Journal | Clinical Medicine Insights: Endocrinology and Diabetes |
| Volume | 12 |
| Issue number | 1-9 |
| DOIs | |
| Publication status | Published - 12 Sep 2019 |
Bibliographical note
Funding Information:Recent work has revealed hGIP(3-30) as an efficacious GIPR antagonist in humans. 33 This accords with the view that only amino acids from position 3-30 of GIP are necessary for GIPR binding. 21 However, the same research group has also established that hGIP(3-30) is selective for primate, but not rodent, GIPRs. 34 Given the plethora of potentially translatable information acquired from GIP studies conducted in rodents, and particularly mice, it is imperative to determine an effective GIPR antagonist for such rodent-based studies. Indeed, because GIP is regarded as the major physiological incretin hormone in man, 38 as suggested over 15 years previously in rodents, 26 the need is even more imperative. - In terms of GIPR agonism, the first two N-terminal amino acids of GIP are known to be essential for agonist properties. 21 In keeping with this, and the idea that only amino acids 3-30 are required for GIPR interaction, several studies have established hGIP(1-30) as a full GIPR agonist at rat 39 41 and human 30 receptors. This is despite knowledge that the final 12 C-terminal amino acid residues of GIP are believed to enhance intrinsic receptor activity. 19 However, GIP(1-30) is expressed in pancreatic α-cells and within some GIP-producing intestinal K cells, 42 suggesting physiological importance. In keeping with this, hGIP(1-30) and mGIP(1-30) augmented insulin secretion from the rodent-derived BRIN-BD11 pancreatic beta cell line in the current study. However, in mice, mGIP(1-30) appeared to be ineffective in terms of regulating glucose homeostasis, whereas both hGIP(1-42) and hGIP(1-30) displayed good bioactivity. This was unexpected, especially given the reported species specificity of the GIPR signalling system. 21 , 30 Moreover, mGIP(1-30) is naturally expressed in mice 42 and long-acting forms display good bioactivity when administered subchronically to diabetic mice 43 , 44 ; thus, our observations could relate to the strain of mouse or peptide dose employed. Nonetheless, together this suggests that omission of the 31-42 C-terminal sequence of GIP should have no detrimental effect on GIPR interaction of prospective antagonistic GIP-derived peptide sequences. Consistent with this view, the current study demonstrated that GIP peptides retaining C-terminal residues 31-42, namely hGIP(3-42) and hGIP(5-42), were unable to counter GIP-induced insulin secretion in vitro , or GIP-mediated reductions in glucose levels in mice. Interestingly, GIP(3-42) represents the naturally occurring dipeptidyl peptidase-4 (DPP-4) degradation product of GIP(1-42) and is recognised to circulate at relatively high levels in the bloodstream. 45 It was initially considered as a probable GIPR antagonist and postulated to physiologically moderate the insulin secreting and metabolic actions of GIP in vivo . 46 However, the GIP inhibitory effects of hGIP(3-42) are weak, as described here and elsewhere, 47 discounting it as a pharmacologically useful GIPR antagonistic. Given the above, it may be expected that GIP(3-30) should also be discoverable in the circulation, given the ubiquitous expression of DPP-4, 48 but surprisingly this has yet to be confirmed. Nonetheless, mGIP(3-30) and hGIP(3-30), as well as h(Pro 3 )GIP(3-30), countered the insulinotropic action of GIP in vitro . Thus, although (Pro 3 )GIP(1-42) was shown to have agonist activity at hGIP receptors, 30 it is considered a competitive antagonist at rat and mGIP receptors. 20 , 30 In the same experimental setting, hGIP(5-30) was less effective, despite suggestion that it functions as a high-affinity competitive GIPR antagonist. 21 However, such differences could be related to the reported species- or tissue-specific actions of GIP peptides. 30 , 31 As such, substitution of histidine in human GIP, for arginine in mouse and rat GIP at position 18, could have some importance in terms of modulation of the GIPR in our experimental systems. In addition, assessment of receptor binding affinities of the peptides may also help to further understand any differences in GIPR inhibitory activity. When administered conjointly with GIP to mice, only mGIP(3-30) and hGIP(5-30) fully countered the glucose-lowering action of GIP. Interestingly, hGIP(3-30) and hGIP(5-30) have previously been identified as competitive antagonists of the human GIPR. 21 Given the primary physiological action of GIP is glucose-dependent insulin secretion, 49 we further investigated the ability of mGIP(3-30) and hGIP(5-30) to modulate this response in mice. In keeping with species specificity of GIP-mediated biological actions, 30 only mGIP(3-30) effectively prevented GIP-induced elevations of insulin secretion in mice. Thus, hGIP(5-30) is perhaps less useful as a GIPR antagonist in murine systems. Indeed, rat GIP(3-30) is recognised as a high affinity competitive antagonist at the level of the rat, but not human, GIPR. 32 It may also have been interesting to consider the effect of mGIP(3-30) and hGIP(5-30) on GIP-induced glucagon secretion, 50 and further studies in this regard would be required. As well as being useful tools to enable better understanding of GIP physiology, there is important possible therapeutic application of GIPR antagonists for obesity. 12 For instance, studies employing GIPR blockade in rodents through genetic deletion of the GIPR, 51 , 52 active or passive immunisation, 7 , 9 , 53 small molecular weight receptor antagonists 54 , 55 or peptide-based GIP inhibitors 19 , 20 , 23 , 56 all provide clear evidence for anti-obesity effects of attenuation of GIPR signalling. In humans, GIP induces cytokine expression, lipolysis and insulin resistance in adipocytes, 57 and hGIP(3-30) inhibits GIP-induced increases in abdominal adipose tissue blood flow and decreases adipose tissue triacylglyceride uptake. 58 , 59 Furthermore, knowledge that highly effective bariatric weight loss surgeries are, in part, linked to surgical removal of GIP-secreting K cells and compromised GIP secretion 13 , 14 strongly suggests translatable benefits of GIPR antagonists for human obesity. Notably, the dose of GIPR antagonists employed for the current study is well beyond normal circulating levels of GIP, 60 implying that such regimens would effectively annul the biological actions of endogenously released GIP. In conclusion, the current studies support the concept of species-specific activity of GIP in different mammalian systems. 21 , 30 , 32 , 34 Given the potential therapeutic application of peptide-based GIPR antagonists in human obesity and diabetes, 61 the origin of such GIP peptides needs to be carefully considered. In this regard, we present mGIP(3-30) as a highly effective molecule to inhibit GIPR activity in mice. Utilisation of mGIP(3-30) for murine studies should better reflect the expected impact of GIPR inhibition with human GIP sequences, such as hGIP(3-30), in the human setting. Funding: The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: This work was supported by a PhD studentship (awarded to RAP) from the Department for the Economy (DfE) Northern Ireland and University of Ulster strategic research funding. Declaration of Conflicting Interests: The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article. Author Contributions NI, VAG and PRF conceived the study, drafted the manuscript and revised it critically for intellectual content. RAP, SLC and MTN participated in the analysis and interpretation of data, drafted the manuscript and revised it critically for intellectual content. All authors approved the final version of the manuscript. ORCID iD SL Craig https://orcid.org/0000-0001-7871-1468
Publisher Copyright:
© The Author(s) 2019.
Copyright:
Copyright 2019 Elsevier B.V., All rights reserved.
Keywords
- GIP
- insulin secretion
- glucose homeostasis
- Species Specificity
- Glucose-dependent insulinotropic polypeptide (GIP)
- species specificity
