A Strong Stomach for Somatostatin Free

Evolution of life from single cell to complex multicellular organisms necessitated sophistication in communication between a myriad of specialized cell types. This communication is enabled by a network of neurocrine, endocrine, paracrine, and juxtracrine (contact-dependent) arrangements. The first 3 forms involve release of chemical messengers that activate receptors expressed on the surface or within the target cell to affect a response. Such an array of communication modes enables rapid (eg, neurocrine) and selective (eg, paracrine) communication, while also ensuring a broader reach of chemical substances to all cells within the system (eg, endocrine). One unique chemical within the organism that functions and is regulated by all of the aforementioned modes is somatostatin. Originally isolated from hypothalamic extracts as an inhibitory signal to GH release from the pituitary gland (1, 2), somatostatin was later found to be synthesized by an extensive network of distinct endocrine cells in the gastrointestinal (GI) tract and pancreas (3). Somatostatin subsequently was discovered to function broadly throughout the body as an inhibitor of multiple secretory processes, including insulin and glucagon from the pancreas, gastrin, secretin, cholecystokinin (CCK), vasoactive intestinal peptide, glucagon-like peptide 1, gastric inhibitory peptide, ghrelin, pepsin, and gastric acid from the digestive tract, T₃, T₄, and calcitonin from the thyroid gland, aldosterone from the adrenal gland, and monoamines from the brain (4, 5). Clinical therapeutics has taken advantage of some of these functions, resulting in the development of various somatostatin analogues for diagnosis, staging and treatment of certain neuroendocrine tumors and for treatment of acromegaly; they also hold promise to treat inflammatory conditions as well as obesity and diabetic complications (4, 6–8). Further studies on somatostatin-secreting cells, and in particular, the mechanisms regulating somatostatin release, hold similar promise.

Gastric D cells serve as a major source for circulating somatostatin and are of particular interest because of their relatively large numbers as compared with other sources and due to their distinctive ability to secrete somatostatin in response to a plethora of neurotransmitters, hormones as well as nutrients. These somatostatin-secreting cells are similar to other gastric endocrine cells, which include ghrelin cells, serotonin-secreting enterochromaffin cells, histamine-secreting enterochromaffin-like cells and gastrin-secreting G cells, in that they are sparsely distributed within the mucosa. Nestled among neighboring endocrine and exocrine cells, the D cells exert inhibitory input onto their neighbors’ secretory capacities via release of somatostatin from characteristic basal cytoplasmic peduncles. Gastric D cells are also major contributors to the picomolar concentrations of somatostatin found in circulation, even though most of the somatostatin produced in the stomach acts locally and is rapidly degraded by proteases. Somatostatin therefore lacks selectivity and precision in its role as a “hormone,” in terms of both site of origin and its target organs (9). The speckled distribution of D cells within the gastric mucosa has hindered the derivation of homogenous populations of D cells thus slowing research on their physiology. Also, lack of selective antibodies to membrane-bound G protein-coupled receptors (GPCRs) and other receptor classes has largely prevented immunohistochemical characterization of the signaling systems that directly regulate somatostatin secretion and other aspects of D-cell function. Instead, insights into somatostatin secretion from D cells so far have been obtained mostly from functional studies in whole animals or from isolated/perfused stomach preparations with pharmacological agents.

In the current issue of Endocrinology, Egerod et al (10) and Adriaenssens et al (11) independently have overcome these well-recognized anatomical and other experimental barriers to provide comprehensive transcriptome analyses of the largely uncharacterized population of somatostatin secreting gastric D cells. In particular, the 2 groups generated D cell-specific reporter mice by crossing somatostatin-selective Cre recombinase-expressing mouse lines, generated by transgenic (11) or by knock-in (12) approaches, with fluorescent reporter mice. The marking of somatostatin-secreting D cells with fluorescent proteins facilitated fluorescence activated cell sorting enrichment of D cells, which in turn permitted semiquantitative expression analyses of the hormones, receptors and other expressed transcripts within the D cells. Although Adriaenssens et al (11) undertook a broad RNA sequencing approach followed by a quantitative PCR approach on genes of interest, Egerod et al (10), focused on characterizing peptides and GPCR transcripts. The roles of subclasses of the identified coexpressed proteins were then tested pharmacologically using cell culture models containing mixed populations of gastric mucosal cells. The studies not only confirm expression of receptors for ligands with known function on somatostatin cells, but also report novel data, including some surprising findings, on transcript expression for several receptors and peptides.

One of the most interesting findings from this set of papers was the identification of other secreted proteins coexpressed with somatostatin. Indeed, transcripts of propancreatic polypeptide (ppy), peptide YY (pyy), proamylin (iapp), gip, cck, gastrin, and ghrelin were all found in somatostatin(+) cells, with the first 3 peptides enriched about 100-fold (11) to more than 1000-fold (10) compared with somatostatin(−) cells. This finding lends substantial credence to the view of coproduction of multiple regulatory peptides within the same enteroendocrine cell types, as opposed to the older dogma of one cell type-one hormone (13–15). The variation in the fold enrichment of somatostatin and coexpressed proteins between the 2 studies could be due to higher efficiency of reporter labeling (∼90%) with the knock-in approach (10) compared with the moderate labeling (60%–80%) with the transgenic approach (11), or could be due to differences in methods used for selecting the somatostatin(+) and somatostatin(−) pools. Although somatostatin was by far the most highly enriched transcript among the peptides in the labeled somatostatin(+) cells, the finding of multiple other regulatory peptides highlights the potential heterogeneity of the gastric D-cell population and thus a potential nomenclature problem in identifying these cells exclusively as “somatostatin cells.” It remains to be seen whether these enriched and/or highly expressed other peptides within the gastric D cells are synthesized and secreted alongside somatostatin from the same secretory vesicles and what their specific function could be when secreted from the inhibitory somatostatin cells. Transcriptional profiling of coexpressed proteins and transcription factors in gastric endocrine cells can help in lineage tracing and potentially form the basis of future work investigating the developmental biology of normal gastric endocrine cells as well as that of endocrine tumors.

Also of interest, the transcript analysis studies on somatostatin(+) cells identified expression of several receptors that also occur in other gastric exocrine and endocrine cells. For example, let us consider the metabolite GPCR calcium-sensing receptor (CaSR), the function of which is allosterically regulated by aromatic amino acids and pH within the stomach lumen. CaSRs expressed in gastrin secreting G cells in the stomach antrum have been postulated to serve as the putative acid/nutrient sensors inducing gastrin secretion, based on results from genetic deletion of CaSR receptors in mice (16). The present studies (10, 11) suggest that Casr is also highly enriched in D cells, as was recently shown to furthermore be the case for ghrelin cells (17). Thus, besides inducing gastrin secretion from gastric G cells (16), inducing acid secretion from gastric parietal cells (18), and inhibiting ghrelin secretion from gastric ghrelin cells (17), direct activation of CaSR also induces somatostatin secretion from D cells (10, 11). Somatostatin, on the other hand, is proposed to inhibit secretion of acid, gastrin and ghrelin by acting on all these cell types in paracrine fashion, providing a potential counterregulatory, “checks-and-balances”-type response to most of the same stimuli (17, 19, 20). Somatostatin cells also express other nutrient-sensing receptors that include, GPR43 (Ffar2), GPR120 (Ffar4), and GPR81, suggesting that somatostatin cells can respond to fatty acids, carbohydrates as well as amino acids emanating from the digesta or circulation. Somatostatin-secreting gastric D cells could thus be viewed as a local “brain,” integrating both nutrient signals and neuronal signals to enhance or release the brake on the function of gastric exocrine and endocrine cells.

Other notable neuropeptide receptors that were found to be enriched and functional in somatostatin(+) cells include those for calcitonin gene-related peptide (Calcrl and Ramp1 subunits) and vasoactive intestinal peptide (Vipr1); notable hormonal receptors include those for glucagon-like peptide 1 (Glp1r), gastric inhibitory peptide (Gipr), and CCK. Regarding CCK, Adriaenssens et al (11) found higher expression and enrichment of CCKA receptor (Cckar) and barely detectable expression of CCKB receptor (Cckbr). Their expression data was complemented by functional studies in which CCK (a preferential agonist for CCKA) induced somatostatin secretion, whereas gastrin (a preferential agonist for CCKB) did not significantly alter somatostatin secretion. These results differed from those of Egerod et al (10), in which high expression and enrichment of CCKB receptors but not CCKA receptors was observed. Their functional studies showed response to both CCK and gastrin. The reason for the contradicting results is not apparent but again could be due to slight differences in methodologies.

The transcriptional profiling also brought to light some unexpected findings, such as the enrichment of adrenergic receptors and the distribution of muscarinic receptor subtypes in somatostatin(+) cells (10). The finding of adrenergic receptors within somatostatin(+) cells was similar to the recent revelation of highly expressed and enriched adrenergic receptors within gastric ghrelin cells (10, 21). These findings draw emphasis on the putative role of adrenergic innervation onto these gastric endocrine cells, contrasting with the more traditionally recognized innervation by and function of the cholinergic system in regulating somatostatin secretion (10, 11) and GI function in general (22). Regarding cholinergic regulation of somatostatin release, the well-understood vagal inhibition of somatostatin secretion upon food intake was believed to be mediated through inhibitory G_i-coupled M₂ and M₄ muscarinic receptors on D cells, in contrast to other gastric endocrine cells which express stimulatory G_q-coupled M₁, M₃ and/or M₅ muscarinic receptors (23). Although both present studies confirm high enrichment of M₄ receptors (chrm4) on gastric D cells, M₃ (chrm3) receptors also were highly expressed compared with M₂ (chrm2) receptors. These results may explain the inconsistent somatostatin secretion results with acetylcholine treatment observed previously. It is also worth noting here that high expression of and/or enrichment of a particular receptor in somatostatin(+) cells does not necessarily mean that the receptor is a critical regulator of D-cell function. Indeed, although the approach of studying in detail those receptors that are both highly expressed and highly enriched in D cells or in any other particular cell type of interest seems like a very reasonable starting point to identify regulatory machinery elements of physiologic significance, this strategy also has potential for concentrating effort on pathways that eventually will be found to be of little consequence or alternatively, for overlooking some important regulators.

In conclusion, the studies by Egerod et al (10) and Adriaenssens et al (11) present highly complementary results and tools that will serve as a curtain raiser to further investigate cellular aspects of somatostatin secretion from D cells as well as to examine as-of-yet understudied facets of somatostatin paracrine and endocrine functions in the stomach, pancreas and elsewhere. Of interest, although a fair amount of literature has described the role of intestinal hormones and changed intestinal function in the pathophysiology of various metabolic diseases and after various metabolic manipulations (24, 25), and although many of these gut hormones are now the targets of marketed or developing drugs to treat conditions such as diabetes and obesity, studies on gastric-derived hormones have lagged behind. Studies such as these on the physiology of somatostatin-secreting gastric D cells, as well as others such as those focused on gastric ghrelin cells (26–28), gastric enterochromaffin-like cells (29), and hopefully future studies making use of similar types of mouse models and methods to study gastric enterochromaffin cells and gastrin cells, are thus of intense interest. Determining how these various gastric endocrine cell types work alone, in concert and coordinately with downstream intestinal endocrine cell types and determining the alterations to their function, number, and anatomic distribution upon exposure to various beneficial (eg, prebiotic fiber therapy) or detrimental (eg, high fat) dietary manipulations, in the settings of diabetes, obesity, or gastric esophageal reflux disease, or after procedures such as bariatric surgery (30), have the promise of revealing novel therapeutic strategies for a wide variety of metabolic and other GI disorders.

Acknowledgments

This work was supported by the National Institutes of Health Grant R01DK103884 and by an International Research Alliance with the Novo Nordisk Foundation Center for Basic Metabolic Research at the University of Copenhagen, Denmark (J.M.Z.) and the Hilda and Preston Davis Foundation Postdoctoral Fellowship Program in Eating Disorders Research (B.K.M.).

Disclosure Summary: The authors have nothing to disclose.

Abbreviations

 
• CaSR

  calcium-sensing receptor

 
• CCK

  cholecystokinin

 
• GI

  gastrointestinal

 
• GPCR

  G protein-coupled receptor.

References