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REvIEwS

The α-​cell in diabetes mellitus


Jesper Gromada1*, Pauline Chabosseau2 and Guy A. Rutter   2*
Abstract | Findings from the past 10 years have placed the glucagon-​secreting pancreatic α-​cell
centre stage in the development of diabetes mellitus, a disease affecting almost one in every ten
adults worldwide. Glucagon secretion is reduced in patients with type 1 diabetes mellitus,
increasing the risk of insulin-​induced hypoglycaemia, but is enhanced in type 2 diabetes mellitus,
exacerbating the effects of diminished insulin release and action on blood levels of glucose.
A better understanding of the mechanisms underlying these changes is therefore an important
goal. RNA sequencing reveals that, despite their opposing roles in the control of blood levels of
glucose, α-​cells and β-​cells have remarkably similar patterns of gene expression. This similarity
might explain the fairly facile interconversion between these cells and the ability of the α-​cell
compartment to serve as a source of new β-​cells in models of extreme β-​cell loss that mimic
type 1 diabetes mellitus. Emerging data suggest that GABA might facilitate this interconversion,
whereas the amino acid glutamine serves as a liver-​derived factor to promote α-​cell replication
and maintenance of α-​cell mass. Here, we survey these developments and their therapeutic
implications for patients with diabetes mellitus.

The history of glucagon begins in 1921 when Frederick Rare ghrelin-​producing ε-​cells also exist in the islet. It is
Banting and Charles Best tested pancreatic extracts in now clear that the human islet cytoarchitecture is similar
depancreatized dogs and detected transient hypergly- to that of rodents7. The islets are made up of subunits
caemia that preceded insulin-​induced hypoglycaemia1. with a central core of β-​cells and a mantle of non-​β-cells,
Murlin et al.2 suggested 2 years later that the early hyper- including α-​cells7. Moreover, good evidence indicates
glycaemia was due to a contaminant in the pancreatic that the portal blood flow relationships are similar in
extract that had glucose agonist properties. In 1948, humans and rodents7. The anatomical similarities are
Sutherland and de Duve3 established that the α-​cells important for understanding paracrine regulation
of the endocrine pancreas are the source of glucagon. of hormone secretion and for extrapolating decades of
Glucagon is a regulator of glucose homeostasis in ani- rodent islet research to humans. In particular, the key
mals and humans. This hormone stimulates glycogen- observation of β-​cells being upstream from α-​cells was
olysis and gluconeogenesis by the liver and is the key shown in a study in which insulin antibody was infused
counter-​regulatory hormone responsible for opposing into perfused rat pancreases8 (see later section). This
the glucose-​lowering effects of insulin. The physiology finding was followed by subsequent studies demonstrat-
and pharmacology of glucagon in health and disease ing that the same mechanism was present in the pancre-
have been covered in recent reviews4,5. ases of dogs, rats, non-​human primates and humans9.
Here, we summarize the role of the glucagon-​ Thus, good evidence suggests that local insulin secretion
secreting pancreatic α-​cell in the development and pro- has a key role in the suppression of glucagon secretion by
1
Regeneron Pharmaceuticals, gression of diabetes mellitus. We review advances in our hyperglycaemia in representative mammalian species.
Tarrytown, NY, USA. understanding of the α-​cell gene expression profile and As discussed below, the remaining puzzle is whether
2
Section of Cell Biology and signalling mechanisms and discuss how the α-​cell might insulin’s suppressive effects are exerted via somatostatin
Functional Genomics, Division
become a more prominent target for the treatment secretion from δ-​cells or by direct effects of insulin on
of Diabetes, Endocrinology
and Metabolism, Department of diabetes mellitus by promoting the reprogramming of α-​cells or, as may well be the case, by both. Using large-​
of Medicine, Imperial College these cells into insulin-​secreting β-​like-cells. scale single-​islet cell quantification, the ratio of α-​cells to
London, Hammersmith β-​cells was shown to be higher in human than in mouse
Hospital Campus, The islet of Langerhans islets, whereas the number of δ-​cells and PP-​secreting
London, UK.
The pancreatic α-​cell was discovered in 1907 (REF.6) cells is similar in these species (Fig. 1). However, during
*e-​mail: jesper.gromada@
and is one of four major types of endocrine cells in the characterization of pancreatic islet preparations for
regeneron.com;
g.rutter@imperial.ac.uk the islets of Langerhans: glucagon-​secreting α-​cells, clinical trials, it was found that the number of β-​cells
https://doi.org/10.1038/ insulin-​ producing β-​ c ells, somatostatin-​ releasing is three times higher than the number of non-​β-islet
s41574-018-0097-y δ-​cells and pancreatic polypeptide (PP)-secreting cells. cells, including α-​cells10. Interestingly, α-​cells might

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α-​Cell function and mass in T2DM


Key points
Type 2 diabetes mellitus (T2DM) is often thought of as
• The mechanisms involved in the control of glucagon secretion in pancreatic α-​cells a bi-​hormonal disease characterized by hypoinsulinae-
have now been identified. mia and hyperglucagonaemia with elevated blood levels
• The pancreatic α-​cell has a role in the development of diabetes mellitus. of glucose. The hypoinsulinaemia results from insuffi-
• Physiological and pharmacological activators and inhibitors of glucagon secretion cient insulin secretion and is associated with a 25–50%
might provide therapeutic targets. decrease in β-​cell mass12–18. In contrast to the effects on
• Single α-​cell gene expression profiling in health and disease has resulted in new levels of insulin, patients with T2DM often show persis-
insights about the function of α-​cells. tent fasting hyperglucagonaemia and lack of suppression
• Advances in understanding α-​cell to β-​cell reprogramming could lead to new of glucagon levels in the postprandial state19. This hyper-
therapeutic strategies for diabetes mellitus. glucagonaemia aggravates the hyperglycaemia induced
by the hypoinsulinaemia, as the hyperglucagonaemia
enhances hepatic glucose production20. These data sug-
have distinct roles in humans and rodents in responses gest a causal role for glucagon in the pathophysiology of
to parasympathetic innervation of the islet, serving in T2DM and support the development of pharmacological
the former (only) as a source of acetylcholine that then agents that inhibit glucagon secretion (or action) to alle-
‘primes’ β-​cells11. viate hyperglycaemia in patients with T2DM. The status
Despite a key role for insulin in suppression of gluca- of these agents has been reviewed elsewhere4,5. Briefly,
gon secretion, the reason for the differences in the ratio studies in humans using small-​molecule inhibitors or
of human α-​cells to β-​cells versus that seen in the rodent blocking antibodies towards the glucagon receptor have
is unknown. In this context, it is also important to further uncovered strong effects on lowering blood levels of
investigate potential differences in the α-​cell-to-​β-cell glucose but also on-target adverse effects. The essential
ratio observed between human islet isolation centres. question for drug development is whether these features
can be separated sufficiently to provide a wide enough
therapeutic window to allow for safe long-​term use.
Mouse Human In particular, blockade of glucagon receptors is asso-
ciated with α-​cell hyperplasia, abnormal lipid metabo-
lism, hepatic steatosis and elevated plasma levels of liver
enzymes, making inhibition of α-​cell hypersecretion an
appealing strategy for treatment of T2DM4,5. The mech-
anism for the link between inhibition of glucagon recep-
tors in the liver and induction of α-​cell hyperplasia is
discussed in a subsequent section. This effect is a safety
concern as α-​cell hyperplasia increases the risk of devel-
oping glucagonoma21. Glucagon is known to have potent
hypolipaemic effects22,23, including causing a decrease in
50 μm 50 μm triglyceride and VLDL release by the liver24,25, a reduc-
tion in plasma levels of cholesterol23,25 and stimulation of
DAPI Insulin Glucagon free fatty acid β-​oxidation in the liver26. The decrease in
triglyceride and VLDL release is secondary to reduced
VLDL apoprotein production22. Glucagon also reduces
levels of LDL27. Therefore, it is not surprising that gluca-
gon receptor inhibition is associated with increases in
circulating levels of triglycerides and LDL cholesterol
in preclinical models and humans28. The increase in levels
of LDL cholesterol is also attributed to overexpression of
genes with protein products involved in cholesterol syn-
thesis29 and re-​absorption30. LDL cholesterol adversely
influences the risk of cardiovascular disease and increases
levels of LDL cholesterol in the setting of T2DM (the
target population of patients for therapies that block
glucagon receptors), which is a cause for concern as
this effect could potentiate the already elevated risk of
cardiovascular disease in these patients. The increased
α-Cell β-Cell δ-Cell PP cell
risk of developing hepatic steatosis following glucagon
receptor inhibition could be due to reduced free fatty acid
Fig. 1 | The localization and number of α-​cells differ between mouse and human
pancreatic islets. Mouse islets have a β-​cell-rich core (insulin stain in green), which is
β-​oxidation in the liver31. The reason for the increase in
surrounded by a low number of α-​cells (glucagon stain in red). Human islets are plasma levels of liver enzymes is unknown. It is impor-
composed of substructures, each with an arrangement similar to that of mouse islets. tant to emphasize that although drugs that antagonize
Cell nuclei are stained in blue. The pie charts show the proportions of the major glucagon might be of great benefit for the treatment of
endocrine cell types in mouse and human islets. DAPI, 4′,6-diamidino-2-phenylindole; T2DM, sufficient levels of basal insulin are required for
PP, pancreatic polypeptide. their maximal therapeutic effects32,33.

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β-Cell δ-Cell patients with this disease47. Whereas glucagon release is


normally regulated reciprocally to that of insulin, being
stimulated as blood concentrations of glucose fall, the
Glucose ↑Insulin Glucose response to low blood levels of glucose is progressively
diminished in T1DM48,49. Given the preserved α-​cell
mass, the insufficient glucagon secretion could be due
Insulin to ‘hypoglycaemia blindness’ in the T1DM α-​cell pop-
Somatostatin
ulation. The mechanism underlying the inability of the
Insulin ↑Somatostatin
receptor α-​cells in patients with T1DM to sense or respond to low
Somatostatin blood levels of glucose is unknown.
α-Cell receptor Although the molecular mechanisms involved in
Glucagon the regulation of insulin secretion are increasingly well
↓cAMP and understood50, our knowledge of those that mediate the
PKA signalling inhibition of glucagon release remains fragmentary.
Glucose Glucose suppresses glucagon secretion by direct signal-
ling mechanisms involving its uptake and metabolism51
but also indirectly through controlling the release of par-
acrine factors derived from β-​cells (or other islet cells)
(Fig. 2) and via glucose-​sensing neurons in the brain52.
β-Adrenergic These mechanisms are discussed in subsequent sections
receptor
and outlined in Supplementary Table 1.
Parasympathetic tone Glucose
Adrenaline and GABAergic neurons
Direct effects of glucose on α-​cells
α-​Cells display spontaneous oscillations in cytosolic lev-
Fig. 2 | intracellular and intercellular mechanisms implicated in the suppression of
els of Ca2+ at low (<3 mM) concentrations of glucose,
glucagon secretion by glucose. Glucose suppresses glucagon secretion by direct
signalling mechanisms involving its uptake and metabolism but also indirectly through
and increases in the glucose concentration lead to both a
controlling the release of paracrine factors derived from β-​cells and/or other islet cells decrease in electrical activity53 and a fall in cytosolic con-
and via glucose-​sensing neurons in the brain. PKA , protein kinase A. centrations and oscillations of Ca2+ (REFS54–56). Although
these changes in Ca2+ are the opposite of those observed
in β-​cells21, evidence suggests that the initial metabolic
Whether α-​c ell mass is unchanged 34,35 or sensing of glucose by the α-cells might be similar to that of
increased15,17,36,37 in patients with T2DM is not clear. β-​cells57 (Supplementary Table 1). In particular, gluco­
Importantly, even in the setting of unchanged α-​cell kinase (encoded by GCK in humans and Gck in rodents)
number, the ratio of α-​cells to β-​cells is higher in patients is expressed in both cell types58, a feature expected to
with T2DM owing to the reduced β-​cell mass compared allow α-​cells to respond metabolically to changes in
with people who do not have T2DM34. Evidence from blood concentrations of glucose in the physiological
the past few years obtained in mouse and non-​human (3–11 mM) and immediate subphysiological (2–3 mM)
primate models of diabetes mellitus and in humans with range. Correspondingly, knockout of Gck in α-​cells leads
T2DM suggests that β-​cells dedifferentiate and adopt to hyperglucagonaemia and progressive development of
α-​cell characteristics38–40. This finding is substantiated hyperglycaemia59. Interestingly, the glucose transporter
by the detection of insulin and glucagon bi-​hormonal GLUT2 (encoded by SLC2A2 or Slc2a2 in humans and
cells in islets from patients with T2DM41. However, it rodents, respectively), which has an affinity for glucose
remains to be established whether the dedifferentiated in the physiological range, is largely absent from α-​cells,
β-​cells make a notable contribution to decreased insulin whereas the higher-​affinity glucose transporter GLUT1
secretion and whether they transdifferentiate into α-​cells (encoded by SLC2A1 or Slc2a1 in humans and rodents,
and secrete glucagon. respectively) is present in both mouse60,61 and human62
In patients with type 1 diabetes mellitus (T1DM), α-​cells. Glucose 6-phosphatase catalytic subunit 2, which
where most (although often not all)42,43 β-​cells are lost, might dephosphorylate glucose to establish a futile
α-​cells make up three-​quarters of the total cell number cycle in β-​cells, is also present in human63, although not
in islets44; however, the absolute mass and therefore the mouse61, α-​cells. The nutrient-​sensitive protein kinases
ratio of α-​cells to δ-​cells to PP cells does not seem to AMP-​a ctivated protein kinase (AMPK)64 and PAS
be altered37. Interestingly, α-​cells might be an important domain-​containing serine/threonine-​protein kinase
source of glucagon-​like peptide 1 (GLP1), and other (PASK)65 are also implicated in α-​cell glucose sensing.
peptides45,46, capable of stimulating insulin secretion to Thus, metabolic sensing of glucose by the α-​cell is likely
lessen the symptoms of T2DM. to be similar to that of β-​cells but acts through differ-
ent glucose transporters. The differential expression of
Glucagon secretion is regulated by glucose GLUT1 and GLUT2 in α-​cells versus β-​cells might affect
Appropriate stimulation of glucagon release from α-cells the detection glucose range for the sugar and, conceivably,
is particularly important to minimize the impact of acute downstream cellular responses.
insulin-​induced hypoglycaemia. This form of hypo­ Mitochondrial metabolism of glucose is central to
glycaemia is a major complication of T1DM, and it is the stimulation of insulin release from β-​cells66,67 and
estimated to be responsible for up to 4% of all deaths in is abnormal in patients with T2DM68. Both α-​cells and

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Physiological activators Pharmacological


• GIP activators
• GRP • SGLT2 inhibitor
• Adrenaline • Sulfonylurea
• Acetylcholine
• Cholecystokinin
• Ghrelin
α-Cell

Glucagon

β-Cell
Physiological Pharmacological
inhibitors inhibitors
• Insulin • Sulfonylurea
• Leptin • DPP4 inhibitor
• Secretin • GLP1
• Somatostatin • Amylin
• Serotonin
• GABA
• GLP1

Fig. 3 | Physiological and pharmacological activators and inhibitors of α-​cell function and glucagon secretion.
Physiological activators and inhibitors of α-​cell function and glucagon secretion can act via paracrine mechanisms (for
example, insulin, somatostatin and GABA) or as circulating hormones (such as glucagon-​like peptide 1 (GLP1), glucose-​
dependent insulinotropic peptide (GIP) and ghrelin) or be released from nerve endings within the islets (for example,
acetylcholine and cholecystokinin). Sulfonylureas are therapeutic agents used in the treatment of type 2 diabetes mellitus
(T2DM) and have been reported to both stimulate and inhibit glucagon secretion depending on the experimental
conditions. GLP1 and inhibitors of dipeptidyl peptidase 4 (DPP4), an enzyme that degrades GLP1, are commonly used
agents in the management of T2DM. Part of the glucose-​lowering effect of these agents results from inhibition of
glucagon secretion. The sodium–glucose cotransporter 2 (SGLT2) inhibitor dapagliflozin stimulates glucagon secretion
by an unknown mechanism. GRP, gastrin-releasing peptide.

β-​cells express fairly low levels of lactate dehydro­ Insulin. Several studies have provided evidence that
genase (LDHA)69,70 and the monocarboxylate transporter insulin suppresses glucagon secretion75–77. This effect
MCT1 (encoded by Slc16a1)71, which are both ‘disal- could be mediated directly via insulin receptors on the
lowed’ islet genes (that is, genes for which the expres- α-​cells or indirectly via increased somatostatin secretion
sion is suppressed compared with the majority of cells from neighbouring δ-​cells78 (Fig. 2). The suppression of
in the body)72,73. Thus, oxidative metabolism of glyco­ glucagon secretion by insulin is, at least in part, mediated
lytically derived NADH and pyruvate might be crit­ by a reduction of intracellular cAMP levels and weak-
ical for suppressing glucagon secretion. Nonetheless, ened protein kinase A (PKA) signalling79. α-​Cells express
a study of fluorescence-​activated cell sorting (FACS)- somatostatin receptors, and their activation suppresses
purified cells61 revealed that Ldha (sixfold) and Slc16a1 glucagon secretion78. In T2DM, altered α-​cell function
(threefold to fourfold) mRNA levels are significantly could result from a combination of reduced insulin
higher in mouse α-​cells than β-​cells61, suggestive of at secretion and impaired insulin action80–82.
least partially distinct signalling mechanisms for glu-
cose in the two cell types. As discussed in a subsequent Zinc ions. Zn2+ has an important role in the storage of
section, RNA sequencing of single human α-​cells did insulin in β-​cell secretory granules. In 2003, Ishihara
not reveal changes in the expression of genes associated and colleagues83 provided evidence that, in the per-
with glucose metabolism or related pathways in T2DM; fused rat pancreas, glucose-​stimulated release of Zn2+
however, the limited sensitivity of this approach might from β-​cells contributed to the inhibition of glucagon
preclude the detection of changes in the expression of secretion. However, a similar role for Zn2+ in the con-
disallowed genes. trol of glucagon secretion from mouse islets has not
been found51. Moreover, glucagon secretion was normal
Paracrine control of glucagon secretion when measured in islets from mice with inactivation of
In addition to glucose, numerous paracrine, hormonal the secretory granule Zn2+ transporter ZnT8 (Slc30a8)84
and nervous signals fine-​tune glucagon secretion under under conditions where release of Zn2+ from the β-​cell
different physiological conditions 74. Possible roles is essentially eliminated85. Interestingly, presumed loss-​
for key paracrine signalling are presented in Fig. 2 and of-function variants in the SLC30A8 gene are associ-
discussed in this section. ated with reduced risk of T2DM86, and inactivation87

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a
β-Cell δ-Cell
Microbeads

α-Cell
PP cell
ε-Cell
Sequencer
Oil
Isolated pancreas Dissociated islet cells Single-cell capture and sequencing

b c 104 d α-Cell β-Cell


20,000 GCG IAPP

β-Cell (average UMI + 1)


Number of detected genes

INS TTR INS


103
15,000 CRYBA2 HADH
TTR TM4SF4 DLK1
102
10,000 GCG TMEM176B RBP4
PCSK2 NPTX2
5,000 101 ALDH1A1 SHISAL2B
α-Cell β-Cell GC UCHL1
0 100 CHGB GAD2
0 2,500 5,000 7,500 10,000 100 101 102 103 104 FXYD3 SMAD11
Number of cells α-Cell (average UMI + 1)
Fig. 4 | rnA sequencing of single human islet cells reveals a few genes that are enriched in α-​cells and β-​cells
among a large number of detected genes. a | Schematic of workflow from donor to isolation of single human islet cells
being loaded onto a capture and sequencing platform. Dissociated human islet cells, including glucagon-​secreting α-​cells,
insulin-​producing β-​cells, somatostatin-​releasing δ-​cells and pancreatic polypeptide (PP)-secreting cells, as well as rare
ghrelin-​producing ε-​cells, were obtained from isolated pancreatic islets from non-​diabetic donors. The single islet cells,
reagents and single gel beads containing barcoded oligonucleotides were encapsulated into nanolitre-​sized particles
using a capture and sequencing platform. Lysis and barcoded reverse transcription of RNAs from single cells were
performed inside each nanolitre-​sized particle. High-​quality next-​generation sequencing libraries are finished in a single
bulk reaction. b | The number of genes detected in human α-​cells and β-​cells increases with the number of sequenced
cells. c | Scatterplot of all detected genes in human α-​cells and β-​cells (n = 20,757). The top enriched β-​cell (INS) and α-​cell
genes (GCG and TTR) are highlighted. d | The top ten enriched genes for human α-​cells and β-​cells. Please see
Supplementary Table 2 for further details. The data were obtained from REFS103,104. UMI, unique molecular identifier.

or overexpression88 of Slc30a8 selectively in the α-​cell parasympathetic and GABAergic neurons is controlled
enhances or reduces hypoglycaemia-​induced glucagon by glucose in the brain (Fig. 2).
release, respectively, suggesting a possible contribution
to altered T2DM risk. Of note, we have also reported Activators and inhibitors. Figure  3 lists physiolog­
that selective deletion of the T2DM-​associated gene ical and pharmacological activators and inhibitors of
Tcf7l2, which encodes transcription factor 7-like-2 and glucagon secretion. For example, catecholamines stim-
is involved in WNT signalling, from the α-​cell in mice89 ulate glucagon secretion via the activation of adren­
impairs the normal regulation of glucagon secretion ergic receptors on both rodent and human α-​cells94.
by glucose, possibly by increasing Slc30a8 expression Sulfonylureas have been reported to both stimulate and
(as observed after Tcf7l2 silencing in islets)90. Thus, inhibit glucagon release depending on the experimental
the actions of the T2DM-​associated variants present conditions. The intestinal-​derived glucose-​dependent
at the risk loci described here, and some of the ~400 insulinotropic polypeptide (GIP) stimulates glucagon
other T2DM risk loci in humans91, might also, at least secretion and contributes to hyperglycaemia in T2DM95.
in part, be mediated via altered release of glucagon. This Accordingly, an antagonist of the GIP receptor dampens
possibility is an exciting current area of research. hyperglycaemia in T2DM95. Another intestinally derived
peptide, GLP1, inhibits glucagon secretion via para­crine
GABA. GABA92,93 is stored in specialized vesicles in the mechanisms involving stimulation of somatostatin
β-​cells, and GABA release contributes to the inhibition and potentially insulin secretion96. Inhibitors of dipep-
of glucagon secretion in rodents and humans. As men- tidyl peptidase 4 (DDP4), which degrades and inacti-
tioned in a subsequent section, GABA might have a role vates GLP1, also inhibit glucagon secretion4,5. GLP1
in the reprogramming of α-​cells into β-​like-cells. This receptor agonists and DPP4 inhibitors are widely used
effect of GABA holds promise for replenishing the pool anti-​diabetic drugs4,5. Of note, inhibition of the sodium–
of functional β-​cells in diabetes mellitus. glucose cotransporter 2 (SGLT2) with dapagliflozin, a
In addition to insulin, GABA and Zn2+ ions, gluca- treatment now widely used in T2DM to promote glucose
gon secretion is regulated by adrenaline, acting via loss through the kidney, also triggers glucagon secretion
β-​adrenergic receptors, as well as by the parasympa- from α-​cells97. The potential long-​term implications of
thetic tone and GABAergic neurons. The activity of the this unexpected effect of dapagliflozin on the α-​cell

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a Gcgr–/–
(mice)
Glucagon antibody α-Cell β-Cell Liver Gcgr–/–
(rodent)
(mice)
Glucagon receptor antibody Gcg–/–
(rodent and monkey) (mice)
Glucagon receptor antisense
oligonucleotide knockdown Pcsk2–/–
(rodent) (mice)

Loss-of-function variants Liver Gs–/–


in glucagon receptor (mice)
(human) PP cell
δ-Cell

Inhibition of glucagon action

b Increased α-cell mass


Mouse α-cell Human α-cell

↑Glutamine ↑Amino acids Amino acid


and/or alanine
SLC38A5 ? ? ?

Liver

mTOR Glutamine ↓Amino acid uptake mTOR?


and/or and catabolism
alanine

Glucagon
α-Cell expansion receptor α-Cell expansion
inhibition

↑Glucagon secretion ↑Glucagon secretion

Fig. 5 | The liver–α-​cell axis. a | Genetic or pharmacological disruption of hepatic glucagon signalling is invariably linked
to α-​cell hyperplasia in rodents, non-​human primates and humans. b | Amino acids promote glucagon secretion and α-​cell
expansion to increase amino acid uptake and metabolism by the liver. In mouse α-​cells, the amino acid transporter
SLC38A5 and the amino acid sensor mTOR regulate expansion of α-​cell mass. The amino acid transporters regulating
amino acid uptake in human α-​cells and the involvement of mTOR remain to be established. The green circles represent
the amino acids glutamine or alanine, whereas the pink circles represent any amino acid. PP, pancreatic polypeptide.

remain to be determined. Other activators of α-​cells Human α-​cells and β-​cells express at least 17,000–18,000
and glucagon secretion not mentioned elsewhere in this genes (Fig. 4b). More genes are likely to be detected once
Review include gastrin-releasing peptide (GRP), ace- more sensitive RNA sequencing methods become avail-
tylcholine from parasympathetic nerves and ghrelin. In able. The scatterplot of all expressed genes reveals that
addition, leptin, secretin and serotonin inhibit glucagon the transcriptomes of human α-​cells and β-​cells are
secretion. The relative contributions of these regulators strikingly similar. In fact, using a stringent report, we
of human α-​cell glucagon secretion in physiology and found that only 97 genes (0.47%; n = 20,757 for com-
pathophysiology remain to be determined. bined gene sets) are enriched or unique to either cell
type (Fig. 4c). The top ten enriched or unique α-​cell and
Single α-​cell gene expression profiling β-​cell genes103,104 are shown in Fig. 4d. A brief description
A better understanding of the mechanisms involved in of each gene and its role in α-​cell and β-​cell function is
the control of glucagon secretion, and its dysregulation provided in Supplementary Table 2. Thus, the pheno-
in T2DM, has been facilitated by our growing under- types of α-​cells and β-​cells are determined by a fairly
standing of the molecular identity of the α-​cell. In par- small number of genes, supporting the observation that
ticular, advances in the past 5 years in single-​cell RNA the cells of the endocrine pancreas are highly plastic in
sequencing technologies have provided unprecedented nature105,106. Similar overlap exists between mouse α-​cell
insights into the gene expression profiles of human and and β-​cell transcriptomes61,107,108. Correspondingly, an
mouse α-​cells63,98–102. Figure 4a exemplifies the workflow analysis of the expression of islet disallowed genes has
for large-​scale single human islet cell RNA sequencing. revealed broad overlap between these two cell types in

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a Replication

Glucagon secretion
α-Cell
Transcription factors
• ↓ARX and DNMT1
ARX • ↑PDX1 and homeobox
protein Nkx-6.1

Arx–/– Circulating factors


Pax4–/– • ↑Activin A
Foregut Pancreatic Endocrine • ↑IGFBP1
endoderm progenitor progenitor • ↑GABA
PAX4
Extreme β-cell loss
β-Cell
Insulin secretion

Replication

T1DM

GABA
Artemisinins

α/β-Cell GABAAR

Rescue of T1DM

Fig. 6 | α-​cell development and its possible modulation as a therapy in type 1 diabetes mellitus. a | Key genes
and transcription factors regulating α-​cell and β-​cell development, as well as transdifferentiation of α-​cells to β-​cells and
β-​cells to α-​cells. The transcription factor ARX is important for α-​cell development, and lack of Arx expression in mice
(Arx−/−) promotes differentiation of endocrine progenitor cells into β-​cells. PAX4 is important for β-​cell development,
and lack of this transcription factor in mice (Pax4−/−) favours conversion of endocrine progenitor cells into α-​cells. Mature
α-​cells can be reprogrammed into β-​like-cells by reducing the expression of ARX and DNMT1 or increasing the
expression of PDX1 and Nkx-6.1. Extreme β-​cell loss or increasing the concentrations of the circulating factors activin A ,
IGF-binding protein 1 (IGFBP1) and GABA have been shown to promote the reprogramming of α-​cells into β-​cells.
b | β-​cells (green) lost in type 1 diabetes mellitus (T1DM) might be replaced from α-​cell-derived precursors (α/β cells;
shown in purple) in response to treatment with GABA and artemisinins, the latter increasing GABA receptor expression
on α-​cells122. Part b adapted from REF.147, Springer Nature Limited.

mouse islets72. This observation is an important aspect liver–α-​cell axis by genetic (using glucagon receptor-
when considering approaches for reprogramming deficient or glucagon-deficient mice or liver-​specific
α-​cells into β-​like-cells as a novel treatment approach for deletion of Gcgr or the G protein Gs in mice) or phar-
patients with diabetes mellitus. macological inhibition of glucagon action using anti-
bodies to glucagon or the glucagon receptor, as well as
The liver–α-​cell loop knockdown of the glucagon receptor using an antisense
As discussed in the next section, α-​cells might provide oligonucleotide approach, has invariably been linked to
a source of new β-​cells to replace those that are lost or increased α-​cell mass (Fig. 5a). This finding is supported
become dysfunctional in T1DM and T2DM. However, by the observation that rare inactivating mutations in
to avoid a depletion of the α-​cell pool itself, insights are GCGR in humans are associated with glucagonoma21.
required as to how the proliferation of these cells might Amino acids, and in particular glutamine, are critical for
be controlled. Amino acids constitute an integral part promoting α-​cell expansion following disruption of the
of a liver–α-​cell axis by promoting glucagon secretion glucagon signalling pathway110–113. In mouse α-​cells,
and serving as substrates in the process of gluconeogen- the amino acid transporter SLC38A5 and the amino acid
esis to produce glucose in the liver109. Disruption of the sensor mTOR have important roles in regulating α-​cell

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expansion following inhibition of glucagon action111–113 fully mimic those used in the former study122. The physi-
(Fig. 5b). mTOR is also important for α-​cell expansion ological role of GABA for α-​cell to β-​cell reprogramming
during development in mice114, and the increase in α-​cell remains to be established.
mass observed after α-​cell-selective deletion of AMPK An additional reason why α-​cells could be a good
might be explained by a resulting activation of mTORC1, source of β-​cells is that α-​cell mass is maintained in the
which is involved in the regulation of cell size115. Little diabetic state34,35 (see previous sections). Thus, the rate
is known about the pathways regulating human α-​cell of reprogramming of α-​cells into β-​like-cells is unlikely
proliferation. Human α-​cells express several amino acid to be limited by the availability of α-​cells, whereas the
transporters, but their contribution to α-​cell hyperplasia pool of α-​cells will be depleted and affect blood levels
following disruption of hepatic glucagon signalling is a of glucose. This view is supported by an α-​cell ablation
topic for future research113 (Fig. 5b). study in mice demonstrating that only 2% of the α-​cell
mass is required for normal α-​cell function, glucagon
α-​Cell to β-​cell reprogramming signalling and maintenance of blood levels of glucose125.
Cell reprogramming strategies for the generation of Furthermore, the mechanisms that regulate DNA
insulin-​producing β-​like-cells for the treatment of diabe- methylation and chromatin modifications are similar
tes mellitus have been investigated and hold promise for in human α-​cells and β-​cells, which facilitates α-​cell to
the treatment of T1DM and possibly T2DM. The results β-​cell reprogramming126–132. In addition, human α-​cells
are encouraging, and restoration of normal glucose proliferate at a higher rate than other islet cell types and
levels has been achieved in diabetic animal models116. can be induced to proliferate following disruption of the
Several reasons could explain why α-​cells might be a hepatic glucagon signalling pathway133–136. α-​Cell prolif-
good source for new β-​cells. First, α-​cells and β-​cells eration could help maintain the α-​cell mass in a setting
have the same developmental origin, and these cell types of accelerated reprogramming to β-​like-cells. However,
mostly express similar genes104. Second, α-​cell to β-​cell this maintenance might not be necessary as enhanced
reprogramming has been accomplished by repression or glucagon signalling contributes to the diabetic state and
expression of key transcription factors39,117–119 (Fig. 6a), a partial reduction in α-​cell mass could be beneficial
after extreme β-​cell loss120 or following treatment of for maintaining optimal glycaemic control. Finally, the
α-​cells with circulating factors and pharmacological remaining α-​cells might secrete factors (for example,
agents, including GABA and artemisinins121–123 (Fig. 6b). GLP1) that could accelerate the maturation and increase
The latter studies have also demonstrated that glucagon the function of the newly formed β-​like-cells137,138.
secretion and detection via an autocrine loop help main- These data support the view that further research is
tain the α-​cell phenotype. Thus, GABA-​induced inhibi- needed to understand the detailed molecular mecha-
tion of glucagon secretion contributes to the instability nisms that govern α-​cell and β-​cell fates and how to safely
of the α-​cell phenotype and facilitates reprogramming promote α-​cell to β-​cell reprogramming. In patients with
into β-​cells. Particularly encouraging are transcrip- T1DM, efforts towards β-​cell replacement might need to
tional data indicating that the phenotypes of human be combined with immune suppression; however, α-​cell
α-​cells and β-​cells are very similar, which might facilitate to β-​cell reprogramming studies in autoimmune non-​
reprogramming101. It is important to note, however, that obese diabetic mice did not require immune suppres-
the findings using the artemisinin artemether have been sion to achieve long-​term normalization of blood levels
questioned124, although it is unclear whether the condi- of glucose116.
tions used in the latter study (notably the culture period)
Gene expression in T2DM α-​cells
5.0
RNA sequencing studies of single human islet cells have
log2-fold change versus non-diabetic donors

revealed that <0.5% of all detected genes are affected


in T2DM63,101. Stringent inclusion criteria were applied in
2.5 the studies to minimize the contribution of, for instance,
donor–donor variability, differences in medication and
glucose control to the observed changes in gene expres-
0.0 sion. Of the differentially regulated genes, only a few have
an assigned role in islet cell development, growth or func-
tion. None of the detected genes are involved in nutrient
–2.5 sensing or intermediary metabolism. Interestingly, 40%
of the affected genes have been implicated in cell growth
in other cell types, whereas 28% have no known func-
–5.0 tion101. This finding is of interest as reduced β-​cell mass
10–4 10–2 100 102 104 106 in patients with T2DM contributes to onset and progres-
Gene expression (normal counts) sion of the disease (see previously). Figure 7 shows the
directionality of change of the affected genes in α-​cells
Fig. 7 | differentially regulated genes in single α-cells
from donors with type 2 diabetes mellitus. Gene from patients with T2DM. Future studies of the affected
expression profiles were compared between single α-cells genes might provide exciting insights into the disease
from donors with or without type 2 diabetes mellitus. aetiology and unravel druggable pathways for correction
Differentially regulated genes are highlighted in red. of defects in glucagon secretion and maintenance of cell
Adapted with permission from REF.101, Elsevier. number and phenotype in patients with T2DM.

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Reviews

? population from mice102 and humans63 (reviewed in


H REF.141). Importantly, β-​cells in the intact mouse islet
display functional heterogeneity, with a hierarchy exist-
ing in which subsets of interconnected cells behave as
‘hubs’ and ‘followers’. The former are able to serve as
T1DM pacemakers to control overall islet Ca2+ dynamics and
insulin secretion142,143. Whether similar α-​cell–α-​cell
α-Cell ? connectivity exists and controls overall glucagon secre-
tion remains unclear56 (Fig. 8). Nonetheless, indirect evi-
H
dence in support of this view includes the remarkably
penetrant phenotypes of genetic modifications affecting

Secretion
H only a minority (~14%) of α-​cells in the mouse87. β-​Cell
connectivity is altered in human obesity142, in in vitro
models of T2DM143 and after disruption of several genes
associated with T2DM144–146. Whether similar changes
β-Cell might affect the α-​cell population is an intriguing pro-
?
posal (Fig. 8). Of note, α-​cell heterogeneity and special-
Glucose ization should be borne in mind when considering
Glucagon H therapies that could deplete the α-​cell pool to generate
Insulin new β-​cells.
T2DM
H
Secretion

Conclusions
In this article, we have presented an overview of nutri-
ent and stimulus sensing by the α-​cell and compared the
properties of the α-​cell to those of the β-​cell. We also
described changes that occur in each cell type in T2DM
and highlighted critical differences between them in this
Secretion

disease setting. Finally, we discussed the potential for


α-​cells to serve as a source of new β-​like-cells.
Soon, 100 years will have passed since we learned that
glucagon has an important role in the maintenance of
glucose homeostasis by opposing the glucose-​lowering
Fig. 8 | Potential role of α-​cell–α-​cell connectivity in the control of glucagon effects of insulin. Although long thought of as culprits,
secretion. β-​Cells in islets display functional heterogeneity , with a hierarchy existing in the dysfunction of which contributes to the pathology
which subsets of interconnected cells behave as ‘hubs’ (indicated with an ‘H’). Hub cells of both T1DM and T2DM, it now appears that α-​cells
are able to serve as pacemakers to control overall insulin secretion. A similar α-​cell–α-​cell
might also be saviours, able to provide a new source of
connectivity could exist. Intercellular connectivity is potentially affected in diabetes
β-cells. This theory is supported by exciting studies using
mellitus (dashed lines from the hub cell). β-​Cells are shown in green, and α-​cells are
shown in red. T1DM, type 1 diabetes mellitus; T2DM, type 2 diabetes mellitus. pharmacological tools to promote the reprogramming
of α-​cells into insulin-​secreting β-​cells in a preclinical
model of T1DM, resulting in normalization of the blood
A novel perspective levels of glucose116. A demonstration of this provocative
Work published in the late 1980s and early 1990s139,140, concept in clinical practice might ultimately herald a
as well as more recent studies using RNA sequencing new era in diabetes mellitus research and treatment.
and other omics approaches at the single-​cell level, have
demonstrated substantial heterogeneity in the β-​cell Published online xx xx xxxx

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