Cellular and sub-cellular localisation of oxyntomodulin-like immunoreactivity in enteroendocrine cells of human, mouse, pig and rat
We use a monoclonal antibody against the C-terminal of oxyntomodulin (OXM) to investigate enteroendocrine cells (EEC) in mouse, rat, human and pig. This antibody has cross-reactivity with the OXM precursor, glicentin (Gli) but does not recognise glucagon. The antibody stained EEC in the jejunum and colon of each species. We compared OXM/Gli immunoreactivity with that revealed by antibodies against structurally related peptides, GLP-1 and glucagon and against GIP and PYY that are predicted to be in some EEC that express OXM/Gli. We used super-resolution to locate immunoreactive vesicles. In the pancreas, OXM/Gli was in glucagon cells but was located in separate storage vesicles to glucagon. In jejunal EEC, OXM/Gli and GIP were in many of the same cells but often in separate vesicles, whereas PYY and OXM/Gli were commonly colocalised in the same storage vesicles of colonic EEC. When binding of anti-GLP-1 to the structurally related GIP was removed by absorption with GIP peptide, GLP-1 and OXM/Gli immunoreactivities were contained in the same population of EEC in the intestine. We conclude that anti-OXM/Gli is a more reliable marker than anti-GLP-1 for EEC expressing preproglucagon products. Storage vesicles that were immunoreactive for OXM/Gli were almost always immunoreactive for GLP-1. OXM concentrations, measured by ELISA, were highest in the distal ileum and colon. Lesser concentrations were found in more proximal parts of small intestine and pancreas. Very little was in the stomach. In EEC containing GIP and OXM/Gli, these hormones are packaged in different secretory vesicles. Separate packaging also occurred for OXM and glucagon, whereas OXM/Gli and PYY and OXM/Gli and GLP-1 were commonly contained together in secretory vesicles.
KeywordsGut hormones Vesicular stores Human Oxyntomodulin intestine Pancreas
Insulin and glucagon antibodies were kindly provided by Prof Len Harrison. Confocal imaging was performed at the Biological Optical Microscopy Platform (BOMP), The University of Melbourne (www.microscopy.unimelb.edu.au). Ellie Cho (applications specialist, BOMP) provided valuable advice on experimental design and image analysis.
This work was supported by an Australian Government Research Training Program Scholarship to LJF.
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Conflict of interest
The authors declare that they have no conflict of interest.