scholarly journals A Major Lineage of Enteroendocrine Cells Coexpress CCK, Secretin, GIP, GLP-1, PYY, and Neurotensin but Not Somatostatin

Endocrinology ◽  
2012 ◽  
Vol 153 (12) ◽  
pp. 5782-5795 ◽  
Author(s):  
Kristoffer L. Egerod ◽  
Maja S. Engelstoft ◽  
Kaare V. Grunddal ◽  
Mark K. Nøhr ◽  
Anna Secher ◽  
...  
Keyword(s):  
Quantitative Pcr ◽  
Fluorescent Protein ◽  
Peptide Yy ◽  
Enteroendocrine Cells ◽  
Inhibitory Peptide ◽  
Treatment And Prevention ◽  
Peptide Precursor ◽  
Glucagon Like Peptide 1 ◽  
Green Fluorescent ◽  
Coexpression Pattern

Abstract Enteroendocrine cells such as duodenal cholecystokinin (CCK cells) are generally thought to be confined to certain segments of the gastrointestinal (GI) tract and to store and release peptides derived from only a single peptide precursor. In the current study, however, transgenic mice expressing enhanced green fluorescent protein (eGFP) under the control of the CCK promoter demonstrated a distribution pattern of CCK-eGFP positive cells that extended throughout the intestine. Quantitative PCR and liquid chromatography-mass spectrometry proteomic analyses of isolated, FACS-purified CCK-eGFP-positive cells demonstrated expression of not only CCK but also glucagon-like peptide 1 (GLP-1), gastric inhibitory peptide (GIP), peptide YY (PYY), neurotensin, and secretin, but not somatostatin. Immunohistochemistry confirmed this expression pattern. The broad coexpression phenomenon was observed both in crypts and villi as demonstrated by immunohistochemistry and FACS analysis of separated cell populations. Single-cell quantitative PCR indicated that approximately half of the duodenal CCK-eGFP cells express one peptide precursor in addition to CCK, whereas an additional smaller fraction expresses two peptide precursors in addition to CCK. The coexpression pattern was further confirmed through a cell ablation study based on expression of the human diphtheria toxin receptor under the control of the proglucagon promoter, in which activation of the receptor resulted in a marked reduction not only in GLP-1 cells, but also PYY, neurotensin, GIP, CCK, and secretin cells, whereas somatostatin cells were spared. Key elements of the coexpression pattern were confirmed by immunohistochemical double staining in human small intestine. It is concluded that a lineage of mature enteroendocrine cells have the ability to coexpress members of a group of functionally related peptides: CCK, secretin, GIP, GLP-1, PYY, and neurotensin, suggesting a potential therapeutic target for the treatment and prevention of diabetes and obesity.

scholarly journals Intragranular targeting of syncollin, but not a syncollinGFP chimera, inhibits regulated insulin exocytosis in pancreatic β-cells

2005 ◽  
Vol 185 (1) ◽  
pp. 57-67 ◽  
Author(s):  
L B Hays ◽  
B Wicksteed ◽  
Y Wang ◽  
J F McCuaig ◽  
L H Philipson ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Fluorescent Protein ◽  
Zymogen Granule ◽  
Β Cells ◽  
Rat Islets ◽  
Intracellular Ca ◽  
Specific Localization ◽  
Glucagon Like Peptide 1 ◽  
Green Fluorescent ◽  
Insulin Exocytosis

Several proteins play a role in the mechanism of insulin exocytosis. However, these ‘exocytotic proteins’ have yet to account for the regulated aspect of insulin exocytosis, and other factors are involved. In pancreatic exocrine cells, the intralumenal zymogen granule protein, syncollin, is required for efficient regulated exocytosis, but it is not known whether intragranular peptides similarly influence regulated insulin exocytosis. Here, this issue has been addressed using expression of syncollin and a syncollin-green fluorescent protein (syncollinGFP) chimera in rat islet β-cells as experimental tools. Syncollin is not normally expressed in β-cells but adenoviral-mediated expression of both syncollin and syncollinGFP indicated that these were specifically targeted to the lumen of β-granules. Syncollin expression in isolated rat islets had no effect on basal insulin secretion but significantly inhibited regulated insulin secretion stimulated by glucose (16.7 mM), glucagon-like peptide-1 (GLP-1) (10 nM) and glyburide (5μM). Consistent with specific localization of syncollin to β-granules, constitutive secretion was unchanged by syncollin expression in rat islets. Syncollin-mediated inhibition of insulin secretion was not due to inadequate insulin production. Moreover, secretagogue-induced increases in cytosolic intracellular Ca2+, which is a prerequisite for triggering insulin exocytosis, were unaffected in syncollin-expressing islets. Therefore, syncollin was most likely acting downstream of secondary signals at the level of insulin exocytosis. Thus, syncollin expression in β-cells has highlighted the importance of intralumenal β-granule peptide factors playing a role in the control of insulin exocytosis. In contrast to syncollin, syncollinGFP had no effect on insulin secretion, underlining its usefulness as a ‘fluorescent tag’ to track β-granule transport and exocytosis in real time.

scholarly journals Characterization of Leptin-Responsive Neurons in the Caudal Brainstem

Endocrinology ◽  
2006 ◽  
Vol 147 (7) ◽  
pp. 3190-3195 ◽  
Author(s):  
Kate L. J. Ellacott ◽  
Ilia G. Halatchev ◽  
Roger D. Cone
Keyword(s):  
Green Fluorescent Protein ◽  
Energy Homeostasis ◽  
Leptin Receptor ◽  
Fluorescent Protein ◽  
Physiological Role ◽  
Cell Group ◽  
Melanocortin System ◽  
Peptide Precursor ◽  
Green Fluorescent

The central melanocortin system plays a key role in the regulation of energy homeostasis. Neurons containing the peptide precursor proopiomelanocortin (POMC) are found at two sites in the brain, the arcuate nucleus of the hypothalamus (ARC) and the caudal region of the nucleus of the solitary tract (NTS). ARC POMC neurons, which also express cocaine- and amphetamine-regulated transcript (CART), are known to mediate part of the response to factors regulating energy homeostasis, such as leptin and ghrelin. In contrast, the physiological role(s) of the POMC neurons in the caudal brainstem are not well characterized. However, development of a transgenic mouse expressing green fluorescent protein under the control of the POMC promoter [POMC-enhanced green fluorescent protein (EGFP) mouse] has aided the study of these neurons. Indeed, recent studies have shown significant activation of NTS POMC-EGFP cells by the gut released satiety factor cholecystokinin (CCK). Here we show that peripheral leptin administration induces the expression of phospho-signal transducer and activator of transcription 3 immunoreactivity (pSTAT3-IR), a marker of leptin receptor signaling, in more than 50% of NTS POMC-EGFP neurons. Furthermore, these POMC-EGFP neurons comprise 30% of all pSTAT3-IR cells in the NTS. Additionally, we also show that in contrast to the ARC population, NTS POMC-EGFP neurons do not coexpress CART immunoreactivity. These data suggest that NTS POMC neurons may participate with ARC POMC cells in mediating some of the effects of leptin and thus comprise a novel cell group regulated by both long-term adipostatic signals and satiety factors such as CCK.

scholarly journals Glucagon-like peptide 1 and peptide YY are in separate storage organelles in enteroendocrine cells

2014 ◽  
Vol 357 (1) ◽  
pp. 63-69 ◽  
Author(s):  
Hyun-Jung Cho ◽  
Eliza S. Robinson ◽  
Leni R. Rivera ◽  
Paul J. McMillan ◽  
Adam Testro ◽  
...  
Keyword(s):  
Peptide Yy ◽  
Enteroendocrine Cells ◽  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1

scholarly journals The Alphavirus Sindbis Infects Enteroendocrine Cells in the Midgut of Aedes aegypti

Viruses ◽  
2020 ◽  
Vol 12 (8) ◽  
pp. 848
Author(s):  
Yani P. Ahearn ◽  
Jason J. Saredy ◽  
Doria F. Bowers
Keyword(s):  
Fluorescent Protein ◽  
Secretory Granules ◽  
Enteroendocrine Cells ◽  
Laser Confocal Microscopy ◽  
Gfp Expression ◽  
Unique Role ◽  
Posterior Midgut ◽  
Arbovirus Infection ◽  
Green Fluorescent ◽  
Infection Pathway

Transit of the arthropod-borne-virus (arbovirus) Sindbis (SINV) throughout adult female mosquitoes initiates with its attachment to the gut lumen, entry and amplification in midgut cells, followed by dissemination into the hemolymph. Free-mated adult females, aged day 5–7, were proffered a viremic blood suspension via sausage casings containing SINV-TaV-Green Fluorescent Protein (GFP) at a final titer of 106 PFU/mL. Midguts (MGs) from fully engorged mosquitoes were resected on days 5 and 7 post-bloodmeal, and immunolabeled using FMRFamide antibody against enteroendocrine cells (ECs) with a TX-Red secondary antibody. Following immunolabeling, the organs were investigated via laser confocal microscopy to identify the distribution of GFP and TX-Red. Infection using this reporter virus was observed as multiple GFP expression foci along the posterior midgut (PMG) epithelium and ECs were observed as TX-Red labeled cells scattered along the entire length of the MG. Our results demonstrated that SINVGFP did infect ECs, as indicated by the overlapping GFP and TX-Red channels shown as yellow in merged images. We propose that ECs may be involved in the SINV infection pathway in the mosquito MG. Due to the unique role that ECs have in the exocytosis of secretory granules from the MG and the apical-basolateral position of ECs in the PMG monolayer, we speculate that these cells may assist as a mechanism for arboviruses to cross the gut barriers. These findings suggest that MG ECs are involved in arbovirus infection of the invertebrate host.

scholarly journals A stem cell marker-expressing subset of enteroendocrine cells resides at the crypt base in the small intestine

2011 ◽  
Vol 300 (2) ◽  
pp. G345-G356 ◽  
Author(s):  
Yoshitatsu Sei ◽  
Xinping Lu ◽  
Alice Liou ◽  
Xilin Zhao ◽  
Stephen A. Wank
Keyword(s):  
Stem Cell ◽  
Fluorescent Protein ◽  
Ex Vivo ◽  
Enteroendocrine Cells ◽  
Stem Cell Marker ◽  
Stem Cell Markers ◽  
Ki 67 ◽  
Green Fluorescent ◽  
G Protein Coupled ◽  
Crypt Base

The spatial orientation of the enteroendocrine cells along the crypt-villus axis is closely associated with their differentiation in the intestine. Here we studied this relationship using primary duodenal crypts and an ex vivo organoid system established from cholecystokinin-green fluorescent protein (CCK-GFP) transgenic mice. In the primary duodenal crypts, GFP+ cells were found not only in the upper crypt but also at the crypt base, where the stem cells reside. Many GFP+ cells below +4 position were positive for the putative intestinal stem cell markers, leucine-rich repeat-containing G protein-coupled receptor 5, CD133, and doublecortin and CaM kinase-like-1, and also for the neuroendocrine transcription factor neurogenin 3. However, these cells were neither stem nor transient amplifying precursor cells because they were negative for both Ki-67 and phospho-Histone H3 and positive for the mature endocrine marker chromogranin A. Furthermore, these cells expressed multiple endocrine hormones. Tracking of GFP+ cells in the organoids from CCK-GFP mice indicated that GFP+ cells were first observed around the +4 position, some of which localized to the crypt base later in the culture period. These results suggest that a subset of enteroendocrine cells migrates down to the crypt base or stays localized at the crypt base, where they express stem and postmitotic endocrine markers. Further investigation of the function of this subset may provide novel insights into the genesis and development of enteroendocrine cells as well as enteroendocrine tumorigenesis.

scholarly journals Intermittent Hypoxia Up-Regulates Gene Expressions of Peptide YY (PYY), Glucagon-like Peptide-1 (GLP-1), and Neurotensin (NTS) in Enteroendocrine Cells

2019 ◽  
Vol 20 (8) ◽  
pp. 1849 ◽  
Author(s):  
Ryogo Shobatake ◽  
Asako Itaya-Hironaka ◽  
Akiyo Yamauchi ◽  
Mai Makino ◽  
Sumiyo Sakuramoto-Tsuchida ◽  
...  
Keyword(s):  
Nervous System ◽  
Intermittent Hypoxia ◽  
Peptide Yy ◽  
Trichostatin A ◽  
Sleep Apnea Syndrome ◽  
Enteroendocrine Cells ◽  
Mrna Levels ◽  
Gene Expressions ◽  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1

The patients with sleep apnea syndrome are exposed to intermittent hypoxia (IH) during sleep. We previously demonstrated the IH-induced up-regulation of the mRNA levels of anorexigenic peptides proopiomelanocortin (POMC), and cocaine- and amphetamine-regulated transcript (CART) in human neuronal cells. Appetite is regulated not only by the central nervous system but also by the peptides from gastrointestinal tract. Here, we investigated the effects of IH on the gene expression(s) of appetite-inhibiting gut hormones. Human enteroendocrine Caco-2 and mouse STC-1 cells were exposed to IH [64 cycles of 5 min hypoxia (1% O2) and 10 min normoxia (21% O2)] or normoxia for 24 h. Real-time RT-PCR revealed that IH significantly increased the mRNA levels of peptide YY (PYY), glucagon-like peptide-1 (GLP-1), and neurotensin (NTS) in Caco-2 and STC-1 cells. ELISA showed that the concentrations of PYY, GLP-1, and NTS in the culture medium were significantly increased by IH. The mRNA levels of PYY, GLP-1, and NTS were significantly up-regulated even in normoxia by Trichostatin A (TSA) and were significantly decreased even in IH by 5-azacytidine (5AZC), suggesting that IH increases PYY, GLP-1, and NTS mRNAs via alterations in the chromatin structure in enteroendocrine cells. IH might have an anorexigenic influence on the enteric nervous system.

scholarly journals Artificial 64-Residue HIV-1 Enhancer-Binding Peptide Is a Potent Inhibitor of Viral Replication in HIV-1-Infected Cells

2011 ◽  
Vol 2011 ◽  
pp. 1-13 ◽  
Author(s):  
Mouhssin Oufir ◽  
Leslie R. Bisset ◽  
Stefan R. K. Hoffmann ◽  
Gongda Xue ◽  
Stephan Klauser ◽  
...  
Keyword(s):  
Fluorescent Protein ◽  
T Antigen ◽  
Inhibitory Peptide ◽  
Sv40 Large T Antigen ◽  
Binding Peptide ◽  
C Terminus ◽  
Arginine Residues ◽  
Green Fluorescent ◽  
Hiv 1

An artificial HIV-1 enhancer-binding peptide was extended by nine consecutive arginine residues at the C-terminus and by the nuclear localization signal of SV40 large T antigen at the N-terminus. The resulting synthetic 64-residue peptide was found to bind to the two enhancers of the HIV-1 long terminal repeat, cross the plasma membrane and the nuclear envelope of human cells, and suppress the HIV-1 enhancer-controlled expression of a green fluorescent protein reporter gene. Moreover, HIV-1 replication is inhibited by this peptide in HIV-1-infected CEM-GFP cells as revealed by HIV-1 p24 ELISA and real-time RT-PCR of HIV-1 RNA. Rapid uptake of this intracellular stable and inhibitory peptide into the cells implies that this peptide may have the potential to attenuate HIV-1 replicationin vivo.

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