The glucagon gene is expressed along the GI tract in highly specialized gut endocrine cells, designated L cells. The majority of L cells are classically thought to be located in the distal gut, predominantly the ileum and colon.
Although the principal function of enteroendocrine cells is classically to synthesize and secrete gut hormones, enteroendocrine cells may also express TLRs which are important for innate immunity in response to environmental challenge. The TLR ligands LPS, flagellin, or CpG oligonucleotides activate their cognate TLRs on STC-1 cells and enhance L cell secretion leading to release of gut hormones, chemokines and b-defensin-2 both in vitro and in vivo, as described in Activation of Enteroendocrine Cells via TLRs Induces Hormone, Chemokine, and Defensin Secretion. J Immunol. 2007 Apr 1;178(7):4296-303.
Two commonly studied enteroendocrine cell lineages include GIP-secreting K cells and GLP-1-secreting L cells. As gut K cells are sparse and difficult to study, there is little known about the molecular biology of the GIP-secreting K cell. Parker and colleagues isolated purified populations of non-immortalized murine K cells from transgenic mice expressing a yellow fluorscent protein under the control of the GIP promoter. Murine K cells were found to express Kir6.2, Sur1, Sglt1, GPR40, GPR119, and GPR120 and GIP secretion was stimulated by glucose, cyclic AMP and linoleic acid as well as tolbutamide but not by modulation of tast receptor activity. GIP+ cells also expressed the proglucagon gene, and at a much higher level, the Pyy gene. See Nutrient-dependent secretion of glucose-dependent insulinotropic polypeptide from primary murine K cells. Diabetologia. 2009 Feb;52(2):289-98.
The origin of L cell or enteroendocrine cell lineages remains a challenging research issue. In an elegant paper Targeted ablation of secretin-producing cells in transgenic mice reveals a common differentiation pathway with multiple enteroendocrine cell lineages in the small intestine. Development. 1999 Sep;126(18):4149-56. Rindi and Leiter addressed this question by ablating cells expressing a secretin-HSVTK transgene with gancylovir. Remarkably, cells expressing CCK, PYY/enteroglucagon as well as secretin were markedly reduced in number. These studies suggest a developmental relationship between these cell types, possibly invoking a multipotential endocrine stem cell precursor that gives rise to multiple lineages.
Although the "master regulators" of enteroendocrine cell lineage remain unclear, emerging evidence supports a role for Pdx-1 as a key transcription factor for specific gut endocrine cell lineages. Introduction of pdx-1 into rat IEC-6 intestinal epithelial cells induced differentiation into an enteroendocrine phenotype, with induction of chromogranin A, serotonin, CCK, gastrin and somatostatin. See Differentiation of immature enterocytes into enteroendocrine cells by Pdx1 overexpression Am. J. Physiol. 2001;281 229-236
Several genes appear important for both islet and endocrine cell development. For example, mice with the SEYNEU mutation exhibit defective islet organization and markedly reduced numbers of glucagon- immunopositive L cells as described in Mol Endocrinol 1999 Sep;13(9):1474-86 Essential requirement for Pax6 in control of enteroendocrine proglucagon gene transcription
Similarly, the Notch signaling pathway appears essential for the regulation of islet and enteroendocrine cell development, see Nat Genet 2000 Jan;24(1):36-44 Control of endodermal endocrine development by Hes-1
The neurogenin gene is important not only for the formation of islet cell lineages, but also determines specification of gastric endocrine cell lineages in the mouse, including glucagon-immunopositive gastric endocrine cells, as shown in Neurogenin 3 is essential for the proper specification of gastric enteroendocrine cells and the maintenance of gastric epithelial cell identity. Genes Dev. 2002 Jun 15;16(12):1488-97 Similarly, disruption of Ngn-3 produces abnormalities in multiple intestinal endocrine cell lineages as shown in Neurogenin3 is differentially required for endocrine cell fate specification in the intestinal and gastric epithelium. EMBO J. 2002 Dec 2;21(23):6338-47
There appear to be distinct populations of L cells that express GLP-1 alone, or cells that co-express GLP-1/PYY or GLP-1/CCK.
The Drucker lab has isolated and studied a mouse enteroendocrine cell line, designated GLUTag. GLUTag cells respond to the same secretagogues that normally regulate rat enteroendocrine cell function, and appear highly differentiated when compared to the behavior of normal rat intestinal endocrine cells. For a direct comparison of FRIC cultures and GLUTag cells, see Endocrinology 1998 Oct;139(10):4108-14 Regulation of glucagon-like peptide-1 synthesis and secretion in the GLUTag enteroendocrine cell line . L cells appear to be glucose-sensitive and contain multiple ion channels, glucose transporters and glucokinase Glucose-sensing in glucagon-like peptide-1-secreting cells Diabetes. 2002 Sep;51(9):2757-63. Nevertheless, GLP-1 secretion appears normal in human subjects with glucokinase mutations, hence this molecule is unlikely to be essential for glucose-stimulated GLP-1 secretion in L cells Glucokinase, the pancreatic glucose sensor, is not the gut glucose sensor. Diabetologia. 2009 Jan;52(1):154-9.
For a more detailed overview of the properties of GLUTag cells, see GLUTag
A major challenge in the study of the factors that stimulate GLP-1/GLP-2 secretion remains the difficulty in isolating large numbers of viable non-transformed L cells. Gribble and colleagues used a transgenic approach to isolate non-transformed L cells using a fluorescent protein expressed in L cells under the control of the proglucagon promoter. Isolated sorted GLP-1-secreting L cells were analyzed in primary culture. These cells were directly glucose-responsive, and secreted GLP-1 in response to agents causing membrane depolarization or neurotransmitter-stimulated signal transduction. Moreover, comparison of key genes and proteins expressed in and signal transduction pathways operative in L cells with the same genes, proteins, and pathways utilized by the GLUTag cell line revealed remarkable similarities between non-immortalized L cells and GLUTag cells. See Glucose sensing in L cells: a primary cell study Cell Metab. 2008 Dec;8(6):532-9
A human intestinal cell line derived from an adenocarcinoma has been characterized in some detail and exhibits endocrine properties, including chromogranin and proglucagon expression. NCI-H716 cells secrete GLP-1 in response to a number of neurotransmitters, nutrients and secretagogues, as described in A human cellular model for studying the regulation of glucagon-like peptide-1 secretion. Endocrinology. 2001 Oct;142(10):4522-8. Nevertheless, the control of proglucagon gene expression and gene transcription is highly aberrant in NCI-H716 and does not at all recapitulate regulation of gene expression inferred from studies of primary intestinal cell cultures. Hence, the utility of these cells as a differentiated experimental model for the human gut L cell lineage is questionable, as outlined in Aberrant Regulation of Human Intestinal Proglucagon Gene Expression in the NCI-H716 Cell Line. Endocrinology. 2003 May;144(5):2025-33
The molecular mechanisms linking fatty acids to GLP-1 secretion from gut endocrine cells remain incompletely understood. Hirasawa and colleagues have identified a GPCR, designated GPR120 that serves as a receptor for fatty acids on gut endocrine cells. Fatty acids activate the receptor in vitro in association with stimulation of GLP-1 secretion. Whether GPR120 is essential for fatty acid-stimulated GLP-1 secretion in vivo remains unclear. See Free fatty acids regulate gut incretin glucagon-like peptide-1 secretion through GPR120. Nat Med. 2005 Jan;11(1):90-4. Epub 2004 Dec 26
A role for PKCz in the oleic acid-induced stimulation of GLP-1 secretion has been described in studies using gut endocrine cells. Reduction of PKCz expression in GLUTag cells, or addition of a PKCz inhibitor significantly diminished the secretory response to oleic acid, as shown in Protein kinase C{zeta} is required for oleic acid-induced secretion of glucagon-like peptide-1 by intestinal endocrine L cells. Endocrinology. 2006 Nov 16;in press
Similar studies using the STC-1 cell line have identified a receptor for bile acids, designated TGR5 (also known as BG37) which is coupled to stimulation of GLP-1 secretion in a cAMP-dependent manner, as described in Bile acids promote glucagon-like peptide-1 secretion through TGR5 in a murine enteroendocrine cell line STC-1. Biochem Biophys Res Commun. 2005 Apr 1;329(1):386-390
GPR119 was originally described as an orphan G protein coupled receptor expressed on islet β-cells, as outlined in Deorphanization of a G protein-coupled receptor for oleoylethanolamide and its use in the discovery of small-molecule hypophagic agents Cell Metab. 2006 Mar;3(3):167-75. Subsequent studies demonstrated that GPR119 was also directly expressed in human and rodent gut endocrine cells, and in GLUTag cells.
The glucoregulatory actions of GPR119 may include a direct role for activation of the receptor in β-cells leading to direct stimulation of insulin secretion. Indeed, GPR119 agonists activate cyclic formation and may enhance insulin secretion from b-cells and b-cell lines Lysophosphatidylcholine enhances glucose-dependent insulin secretion via an orphan G-protein-coupled receptor Biochem Biophys Res Commun. 2005 Jan 28;326(4):744-51. Erratum in: Biochem Biophys Res Commun. 2005 Apr 1;329(1):417 and A role for beta-cell-expressed G protein-coupled receptor 119 in glycemic control by enhancing glucose-dependent insulin release Endocrinology. 2007 Jun;148(6):2601-9. Epub 2007 Feb 8. Erratum in: Endocrinology. 2007 Oct;148(10):4753. However, the observation that GPR119 is also expressed in gut endocrine cells, and in enteroendocrine cell lines such as GLUTag cells, suggests that activation of GPR119 signaling in enteroendocrine cells may lead to increased secretion of gut hormones such as GIP and GLP-1 as demonstrated by Chu and colleagues A Role for Intestinal Endocrine Cell-Expressed GPR119 in Glycemic Control by Enhancing GLP-1 and GIP Release Endocrinology. 2008 May;149(5):2038-47. Although GPR 119 agonists stimulate the secretion of both GLP-1 and GIP in mice, the actions on GIP secretion may be indirect, as GPR119 receptors have not yet been detected on GIP+ gut enteroendocrine K cells. The identity of the endogenous physiological ligand for GPR119 remains uncertain, but one candidate is oleoylethanolamide (OEA). Administration of OEA enhances GLP-1 secretion from GLUTag cells in a PKA-and GPR119-dependent dependent manner and enteral administration of OEA in the rat small bowel increased GLP-1 and insulin secretion in a glucose-dependent manner as outlined in GPR119 is essential for oleoylethanolamide-induced glucagon-like peptide-1 secretion from the intestinal enteroendocrine L-cell Diabetes. 2009 Feb 10. [Epub ahead of print]
Consistent with the importance of GPR119 in the gut endocrine cell, GPR119 agonists appear to preferentially stimulate insulin secretion and lower blood glucose following enteral administration. Accordingly, GPR119 agonists may exert their glucoregulatory actions at least in part indirectly via stimulation of GIP and GLP-1 secretion, which in turn, promote glucose-dependent insulin secretion. Moreover, at least part of the glucoregulatory actions of GPR119 agonists are diminished by co-administration of the GLP-1R antagonist, exendin (9-39), as shown in A Role for Intestinal Endocrine Cell-Expressed GPR119 in Glycemic Control by Enhancing GLP-1 and GIP Release Endocrinology. 2008 Jan 17; [Epub ahead of print]. The functional properties of different synthetic GPR119 agonists, as well as the naturally ocurring molecules, have been characterized in studies of signal transduction and insulin secretion in Endogenous and synthetic agonists of GPR119 differ in signalling pathways and their effects on insulin secretion in MIN6c4 insulinoma cells. Br J Pharmacol. 2008 Aug 25. [Epub ahead of print]
Although limited information is available about the role of GPR119 in the control of human GLP-1 and/or insulin secretion, a preliminary report by press release describes glucose-lowering properties of a GPR119 agonist designated APD668, in single dose and 14 day administration studies of human subjects, as outlined in the Jan 7 2008 Press Release
The related GPCR designated GPR40 may also play a role in the control of murine gut hormone secretion. GPR40 was localized to subsets of GIP and GLP-1+ endocrine cells in the murine gut and GPR4-/- mice exhibit defective incretin secretion in response to acute administration of fat, whereas GIP and GLP-1 secretion is normal in GPR40-/- mice See Gpr40 is expressed in enteroendocrine cells and mediates FFA stimulation of incretin secretion. Diabetes. 2008 Jun 2. [Epub ahead of print]
Are there examples of enteroendocrine cell deficiency in human disease states? Although admittedly rare, patients with autoimmune polyglandular endocrine failure may exhibit a deficiency of gut endocrine cells, leading to clinical disease. For example, review the case report describing a marked reduction in the number of CCK+ enteroendocrine cells in a human patient with fat malabsorption and diarrhea in the NEJM; Malabsorption due to cholecystokinin deficiency in a patient with autoimmune polyglandular syndrome type I. N Engl J Med. 2001 Jan 25;344(4):270-4.
Are there examples of enteroendocrine cell hyperfunction leading too increased circulating levels of gut peptides that contribute to the development of syndromes in human subjects? See Gut hormones and Bariatric Surgery for an overview