Gly²]-GLP-2, But Not Glucagon or [D-Ala²]-GLP-1, Controls Collagen Crosslinking in Murine Osteoblast Cultures.

Aleksandra Mieczkowska,Guillaume Mabilleau,Erick Legrand,Beatrice Bouvard

doi:10.3389/fendo.2021.721506

Aleksandra Mieczkowska, Guillaume Mabilleau + Show 2 more

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https://doi.org/10.3389/fendo.2021.721506

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Abstract

Bone tissue is organized at the molecular level to resist fracture with the minimum of bone material. This implies that several modifications of the extracellular matrix, including enzymatic collagen crosslinking, take place. We previously highlighted the role of several gut hormones in enhancing collagen maturity and bone strength. The present study investigated the effect of proglucagon-derived peptides on osteoblast-mediated collagen post-processing. Briefly, MC3T3-E1 murine osteoblasts were cultured in the presence of glucagon (GCG), [D-Ala²]-glucagon-like peptide-1 ([D-Ala²]-GLP-1), and [Gly²]-glucagon-like peptide-2 ([Gly²]-GLP-2). Gut hormone receptor expression at the mRNA and protein levels were investigated by qPCR and Western blot. Extent of collagen postprocessing was examined by Fourier transform infrared microspectroscopy. GCG and GLP-1 receptors were not evidenced in osteoblast cells at the mRNA and protein levels. However, it is not clear whether the known GLP-2 receptor is expressed. Nevertheless, administration of [Gly²]-GLP-2, but not GCG or [D-Ala²]-GLP-1, led to a dose-dependent increase in collagen maturity and an acceleration of collagen post-processing. This mechanism was dependent on adenylyl cyclase activation. In conclusion, the present study highlighted a direct effect of [Gly²]-GLP-2 to enhance collagen post-processing and crosslinking maturation in murine osteoblast cultures. Whether this effect is translatable to human osteoblasts remains to be elucidated.

Highlights

Bone tissue is a two-component material made primarily of poorly crystalline hydroxyapatite tablets and type I collagen-based organic matrices [1]
As a band was present in empty vector-transfected CHO-K1 cells, it is difficult to ascertain whether GLP-2r is expressed by MC3T3-E1 osteoblast cells
In order to verify whether such response was specific to [Gly2]-glucagon-like peptide-2 (GLP-2) or could be extrapolated to native GLP-2 peptide, we investigated the extent of activation in intracellular pathways in MC3T3-E1 cells

Summary

Introduction

Bone tissue is a two-component material made primarily of poorly crystalline hydroxyapatite tablets and type I collagen-based organic matrices [1]. Type I collagen biosynthesis is a complex process and includes several intracellular post-translational modifications in osteoblast including assembly of pro-collagen chains, secretion, extracellular processing, and crosslinking to form a mature functional organic matrix [6]. GLP-2 and Osteoblasts collagen molecules assemble into collagen type I fibrils that are subsequently stabilized as a result of oxidative deamination of e-amino groups of specific lysine and hydroxylysine residues by lysyl oxidase, forming aldehyde moieties [7]. Aldehyde moieties are extremely reactive and their interactions with other aldehydes in the side chain of amino acids present in the primary structure of type I collagen result in the formation of covalent crosslinks, critical for a functional mature collagen. Functional lysyl hydroxylase and lysyl oxidase, two key enzymes involved in post-translational modifications of collagen molecules, are compulsory for normal collagen deposition and fibril organization [11]

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