PhD Maja Storm Engelstoft – University of Copenhagen

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Food, Fitness & Pharma > Scientists > PhD's from UNIK > PhD Maja Storm Engelstoft

Maja Storm Engelstoft, PhD

Affiliated with UNIK project no. 9 

Thesis title: Peptide hormones and seven transmembrane receptors in enteroendocrine cells

Thesis summary: Enteroendocrine cells produce hormones essential for gastrointestinal functioning as well as energy and glucose homeostasis and may hold the key for coming advances in obesity and diabetes treatment. To be able to characterize the enteroendocrine cells, which are scattered in the gastrointestinal epithelium, transgenic reporter mice were used as tools to study gene expression in enteroendocrine cells as the basis for functional studies focusing on the peptide hormone expression in chromogranin A, ghrelin and CCK cells, as well as the seven transmembrane domain (7TM) G protein-coupled receptor expression in ghrelin, CCK, GLP-1 and GIP cells.

A chromogranin A reporter mouse was established as part of this work which specifically labelled monoaminergic cells in gastric and colonic tissue emphasizing the important function of chromogranin A as a monoamine binding protein in large dense-core secretory vesicles. In a reporter mouse expressing a variant of the fluorescent protein GFP under the control of the ghrelin promoter the ghrelin precursor was expressed at around three orders of magnitude higher levels than any other of the 93 tested precursors for secreted proteins. In contrast in a CCK reporter mouse enteroendocrine cells of the small intestine expressed CCK, GLP-1, GIP, PYY, neurotensin and secretin at varying levels – but not somatostatin or substance P – determined by qPCR, single cell qPCR, immunohistochemistry and LC-MS. A similar co-expression pattern was obtained through a cell ablation study using GLP-1 promoter-driven diphtheria receptor expressing mice. Thus, toxin treatment resulted in a marked reduction in GLP-1, PYY, neurotensin, GIP, CCK and secretin cells, whereas somatostatin and substance P cells were spared. Importantly, this co-expression pattern was confirmed in human tissue. The ability of a lineage of enteroendocrine cells to co-express this group of functionally related peptides is intriguing with respect to the possibilities for manipulations of the expression profile.

The 7TM receptor repertoire of gastric ghrelin cells, and CCK, GLP-1 and GIP cells from the proximal small intestine was identified through qPCR analysis of FACS-purified cells from the corresponding reporter mice. In the ghrelin cell, the function of highly expressed and enriched 7TM receptors was comprehensively examined using mainly primary cultures of gastric mucosal cells. In addition to a series of expected receptors a number of receptors not previously known to be involved in the control of ghrelin secretion were identified including the composite receptor for the sensory, gastro-protective neuropeptide CGRP, the melanocortin 4 receptor and the orphan receptor GPR142, which all efficiently stimulated ghrelin secretion, and the lactate receptor GPR81 and the short chain fatty acid receptor FFAR2 (GPR43), which effectively inhibited ghrelin secretion. Surprisingly, in the ghrelin cell a number of receptors known to couple through Gαq instead inhibited hormone secretion through Gαi/o mechanisms, which is suggested to be caused by the identified high expression and enrichment of the three non-common Gαi/o subunits, GαoA, GαoB and Gαz.

Thus, the work identifies a novel co-expression pattern for a group of functionally related peptide hormones in enteroendocrine cells of the small intestine and identifies a number of novel receptors controlling ghrelin secretion as well as a molecular mechanism for cell specific signalling of metabolite receptors.

Publications

Maja S Engelstoft at PubMed

Links

Maja S Engelstoft at LinkedIn