Advances in basic research on glucagon and alpha cells

Abstract

The regulation of plasma amino acid levels by glucagon in humans first attracted the attention of researchers in the 1980s. Recent basic research using animal models of glucagon deficiency suggested that a major physiological role of glucagon is the regulation of amino acid metabolism rather than to increase blood glucose levels. In this regard, novel feedback regulatory mechanisms that are mediated by glucagon and amino acids have recently been described between islet alpha cells and the liver. Increasingly, hyperglucagonemia in humans with diabetes and/or nonalcoholic fatty liver diseases is reported to likely be a compensatory response to hepatic glucagon resistance. Severe glucagon resistance due to a glucagon receptor mutation in humans causes hyperaminoacidemia and islet alpha cell expansion combined with pancreatic hypertrophy. Notably, a recent report showed that the restoration of glucagon resistance by liver transplantation resolved not only hyperglucagonemia, but also pancreatic hypertrophy and other metabolic disorders. The mechanisms that regulate islet cell proliferation by amino acids largely remain unelucidated. Clarification of such mechanisms will increase our understanding of the pathophysiology of diseases related to glucagon.

Introduction

The characterization of glucagon as a hyperglycemic substance present in an aqueous extract of the pancreas was first made in 1923 [1]. Since then, it has been widely accepted that the major physiological role of glucagon is to increase plasma glucose levels. However, advances in basic research on glucagon and alpha cells in recent decades highlighted the importance of interorgan communication between islet alpha cells and the liver mediated by glucagon and amino acids.

Glucagon and plasma amino acids in the twentieth century

The first indication of a possible regulation of plasma amino acid levels by glucagon was described in a report on a glucagonoma syndrome [2]. In the report, nine cases with necrolytic migratory erythema and pancreatic tumors were described. Plasma glucagon and amino acid levels were analyzed in four cases, with all four showing hyperglucagonemia and hypoaminoacidemia. In preceding reports in the late 1960s, the infusion of amino acids was shown to stimulate glucagon secretion [3]. In 1980, an increase in plasma amino acid levels in nine pancreatectomized subjects was detailed [4]. Following these reports, Boden et al. demonstrated that the administration of glucagon lowered plasma amino acid levels in healthy volunteers. They also determined that the inhibition of glucagon secretion after the administration of somatostatin resulted in an increase in plasma amino acid levels, thereby demonstrating that glucagon plays pivotal roles in the regulation of plasma amino acid levels [5]. However, because of the higher cost in analyzing blood amino acid levels compared to blood glucose levels, the regulation of amino acid metabolism by glucagon has not been further explored in detail. As a result, the physiological importance of the regulation of plasma amino acids by glucagon has remained under-recognized from a diabetology viewpoint during the last century.

Glucagon as a “foe”: a glucagonocentric view of diabetes

Animal models deficient in the prohormone convertase 2 (pcsk2^−/−) and glucagon receptor (gcgr^−/−) are limited in the production and activity/signaling of glucagon, respectively [6, 7]. Both models displayed lower blood glucose levels and the hyperplasia of islet alpha cells. However, plasma amino acid levels were not analyzed in these models in the earliest reports. Nevertheless, since gcgr^−/− mice were resistant to developing diabetes after the destruction of islet beta cells by streptozotocin, glucagonocentric restructuring of diabetes was proposed by Unger, a pioneer in glucagon research who established the first glucagon radioimmunoassay, and colleagues [8–10]. From a glucagonocentric viewpoint, the hyperglucagonemia often observed in patients with diabetes is considered to be due to the insufficient suppression of glucagon secretion caused by insulin deficiency and/or islet dysfunction. Therefore, Unger et al. proposed that glucagon suppression or inactivation should be considered to control blood glucose levels in patients in addition to insulin supplementation.

Role of GLP-1 in resistance to diabetes in glucagon deficiency

Whereas mice deficient in glucagon receptors display resistance to streptozotocin-induced diabetes [8], mice deficient in receptors for both glucagon and glucagon-like peptide-1 (GLP-1) are sensitive [11]. As glucagon and GLP-1 share a precursor, proglucagon, which is encoded by the glucagon gene, not only glucagon but GLP-1 is also overproduced in gcgr^−/− mice [7]. Glucagon-like peptide-1 is produced through cleavage by pcsk1, but not by pcsk2. Therefore, the production of GLP-1 is not only attenuated, but rather enhanced in gcgr^−/− and pcsk2^−/− mice [12]. As GLP-1 is one of the major incretins, enhanced GLP-1 production in these animal models plays pivotal roles in their resistance to diabetes. In addition to GLP-1, a recent report described how overproduced glucagon acting in the GLP-1 receptor appears to be involved in the development of resistance to diabetes [13].

An animal model deficient in the glucagon gene (gcg^gfp/gfp), which thereby lacks both glucagon and GLP-1, develops diabetes by streptozotocin administration, reinforcing that glucagon deficiency per se is not sufficient to prevent diabetes [14, 15]. Intriguingly, mice deficient in both receptors for glucagon and GLP-1 (gcgr^−/−glp1r^−/−), as well as gcg^gfp/gfp mice, are normoglycemic, suggesting that GLP-1 signaling plays important roles in also lowering blood glucose levels in glucagon deficiency [11, 14, 16, 17].

The destruction of beta cells by diphtheria toxin led to hyperglycemia and death in gcgr^−/− mice that expressed the diphtheria toxin receptor under the control of the insulin promoter [18]. Therefore, in addition to GLP-1, residual beta cells that endure streptozotocin treatment may play important roles in the control of blood glucose levels.

Glucagon deficiency and islet alpha cell proliferation

Multiple animal models of glucagon deficiency have shown hyperplasia of islet alpha cells. In glucagon-gene null models and gcgr^−/−glp1r^−/− mice, hyperplasia of islet cells suggested that lower blood glucose and GLP-1 levels are not prerequisites for the proliferation of alpha cells [14, 16, 19]. However, a mouse model of the liver-specific ablation of the glucagon receptor displays alpha cell hyperplasia, indicating that glucagon activity in the liver plays a central role in the regulation of alpha cell proliferation [20].

The hyperplasia of islet alpha cells has been reported not only in genetically modified models, but also in models administered blocking antibodies to glucagon receptors [21, 22]. Therefore, islet alpha cell proliferation is inducible by glucagon deficiency or glucagon resistance after maturation. However, a study employing a glucagon receptor antagonist suggested that the potential of alpha cells to proliferate appears to decline with age [23].

Regulation of alpha cell proliferation

The cell populations and functions of endocrine organs, such as those of the thyroid, adrenal glands and gonads, are regulated by “trophins” secreted by trophic cells in the pituitary. However, exploration of the regulatory mechanisms involved in the proliferation of islet endocrine cells has been challenging. Candidates for specific “trophins” secreted by the liver, such as angiopoietin-like proteins, have been identified. However, no specific “trophin” has been reproducibly shown to regulate the proliferation of islet endocrine cells [24–26].

In gcg^gfp/gfp mice that lack both glucagon and GLP-1, the expression levels of enzymes involved in amino acid catabolism are decreased in the liver, leading to mice displaying hyperaminoacidemia [27]. Similarly, gcgr^−/− mice also show hyperaminoacidemia [28], indicating that glucagon deficiency results in increased plasma amino acid levels, regardless of the presence or absence of GLP-1. In the two above-mentioned independent studies using blocking antibodies for the glucagon receptor, animals also developed hyperaminoacidemia in addition to islet alpha cell proliferation [21, 22]. The inevitable linkage of hyperaminoacidemia and the hyperplasia/proliferation of islet alpha cells in glucagon deficiency led to the emergence of the notion that amino acids themselves may regulate alpha cell proliferation.

Interorgan communication between liver and islet alpha cells mediated by glucagon and amino acids

Glucagon plays pivotal roles in the regulation of amino acid metabolism. Consequently, the liver, the major target organ of glucagon, has a central role in amino acid metabolism and ureagenesis [21, 22, 27]. At present, mutual feedback regulation between the liver and islet alpha cells mediated by glucagon and amino acids is well established in animal models. A rigorous exploration of the molecular mechanisms involved is ongoing as summarized in recent reviews [29–32].

Glucagon as a “friend” and hyperglucagonemia as compensation for hepatic glucagon resistance

In the insulin-resistant state, both blood glucose and plasma insulin levels increase. In this case, increased insulin production is considered a compensatory response to insulin resistance. Wewer-Albrechtsen et al. reported that both plasma glucagon and amino acid levels, especially alanine, increase in humans with relatively high homeostatic model assessment of insulin resistance [33].

Hyperglucagonemia is often observed in patients with diabetes and/or nonalcoholic fatty liver disease [32, 34, 35]. If glucagon levels are increased due to the insufficient suppression of glucagon secretion and/or islet dysfunction as proposed in a glucagonocentric view of diabetes, the suppression of glucagon secretion and/or blocking glucagon activity seem to be appropriate therapeutic maneuvers [9]. However, the restoration of glucagon resistance or its supplementation would be regarded as physiologically suitable treatments if hyperglucagonemia is compensation for glucagon resistance. In regard to the latter viewpoint, glucagon agonists are expected to have a beneficial effect on metabolism. Indeed, enhanced glucagon signaling in the liver by a GLP-1R/GcgR agonist reduced lipid content and improved mitochondrial function in the mouse liver [36].

Human cases with glucagon deficiency and liver transplantation

Loss of glucagon receptor function leads to severe glucagon resistance resulting in hyperglucagonemia and hyperaminoacidemia [37]. The hyperplasia of islet alpha cells has been reported in multiple cases with glucagon receptor mutations [38]. As hepatic glucagon resistance is sufficient to induce the hyperplasia of islet alpha cells in animal models [20], restoring glucagon receptor function in the liver is expected to normalize hyperglucagonemia, hyperaminoacidemia, and alpha cell hyperplasia. Liver transplantation is considered an effective treatment for a wide variety of disorders based predominantly on the liver, whereas gene therapy for liver diseases is still challenging [39, 40].

Very recently, the first case of liver transplantation in a patient with a loss of glucagon function was reported [41]. Orthotopic transplantation of liver normalized plasma glucagon levels to a normal range 9 months after transplant surgery. The resolution of hyperglucagonemia was accompanied by a reduction in pancreas size, and the normalization of other metabolic parameters and symptoms. This historical case underscored the critical roles of the liver in glucagon signaling and the regulation of proliferation of islet alpha cells.

Conclusion

In the last few decades, the development of animal models of glucagon deficiency has highlighted the crucial role glucagon has in the regulation of amino acid metabolism. Detailed mechanisms for the regulation of islet cell proliferation and/or glucagon secretion by amino acids largely remain to be elucidated. Efforts to clarify such mechanisms should bring further paradigm shifts in understanding the pathophysiological roles of glucagon in subsequent decades.

Declarations

Conflict of interest

The author declares no conflicts of interest associated with this manuscript.

Research involves human and animal participants

This article does not contain any studies with human or animal subjects performed by any of the authors.