DP2, a Carbohydrate Derivative, Enhances In Vitro Osteoblast Mineralisation.

Bone fracture healing is a complex biological process involving four phases coordinated over time: hematoma formation, granulation tissue formation, bony callus formation, and bone remodelling. Bone fractures represent a significant health problem, particularly among the elderly population and patients with comorbidities. Therapeutic strategies proposed to treat such fractures include the use of autografts, allografts, and tissue engineering strategies. It has been shown that bone morphogenetic protein 2 (BMP-2) has a therapeutic potential to enhance fracture healing. Despite the clinical efficacy of BMP-2 in osteoinduction and bone repair, adverse side effects and complications have been reported. Therefore, in this in vitro study, we propose the use of a disaccharide compound (DP2) to improve the mineralisation process. We first evaluated the effect of DP2 on primary human osteoblasts (HOb), and then investigated the mechanisms involved. Our findings showed that (i) DP2 improved osteoblast differentiation by inducing alkaline phosphatase activity, osteopontin, and osteocalcin expression; (ii) DP2 induced earlier in vitro mineralisation in HOb cells compared to BMP-2 mainly by earlier activation of Runx2; and (iii) DP2 is internalized in HOb cells and activates the protein kinase C signalling pathway. Consequently, DP2 is a potential therapeutical candidate molecule for bone fracture repair.

Oligogalacturonic Acid Inhibits Vascular Calcification by Two Mechanisms: Inhibition of Vascular Smooth Muscle Cell Osteogenic Conversion and Interaction With Collagen.

OBJECTIVE: Cardiovascular diseases constitute the leading cause of mortality worldwide. Calcification of the vessel wall is associated with cardiovascular morbidity and mortality in patients having many diseases, including diabetes mellitus, atherosclerosis, and chronic kidney disease. Vascular calcification is actively regulated by inductive and inhibitory mechanisms (including vascular smooth muscle cell adaptation) and results from an active osteogenic process. During the calcification process, extracellular vesicles (also known as matrix vesicles) released by vascular smooth muscle cells interact with type I collagen and then act as nucleating foci for calcium crystallization. Our primary objective was to identify new, natural molecules that inhibit the vascular calcification process. APPROACH AND RESULTS: We have found that oligogalacturonic acids (obtained by the acid hydrolysis of polygalacturonic acid) reduce in vitro inorganic phosphate-induced calcification of vascular smooth muscle cells by 80% and inorganic phosphate-induced calcification of isolated rat aortic rings by 50%. A specific oligogalacturonic acid with a degree of polymerization of 8 (DP8) was found to inhibit the expression of osteogenic markers and, thus, prevent the conversion of vascular smooth muscle cells into osteoblast-like cells. We also evidenced in biochemical and immunofluorescence assays a direct interaction between matrix vesicles and type I collagen via the GFOGER sequence (where single letter amino acid nomenclature is used, O=hydroxyproline) thought to be involved in interactions with several pairs of integrins. CONCLUSIONS: DP8 inhibits vascular calcification development mainly by inhibition of osteogenic marker expression but also partly by masking the GFOGER sequence-thereby, preventing matrix vesicles from binding to type I collagen.

Efficient and stereocontrolled synthesis of chondroitin mono- and disaccharide linked to variously sulfated biotinylated trisaccharides of the linkage region of proteoglycans.

Efficient and stereocontrolled preparation of a library of variously sulfated biotinylated tetra- and pentasaccharides possessing the backbone of the partial linkage region plus the first chondroitin sulfate mono- or disaccharide unit (d-GlcA)n-ß-d-(1,3)-GalNAc-ß-d-(1,4)-GlcA-ß-d-(1,3)-Gal-ß-d-(1,3)-Gal (n = 0 or 1) is reported herein for the first time. The synthesis of these compounds was achieved using common key intermediates and a disaccharide building block obtained by semisynthesis. Stereoselective glycosylation, selective protection/deprotection steps, efficient reduction of the N-trichloroacetyl group into the corresponding N-acetyl group, efficient sulfation strategy, deprotection and biotinylation afforded target oligomers in good yield with high purity.

Synthesis of variously sulfated biotinylated oligosaccharides from the linkage region of proteoglycans.

The synthesis of a collection, as biotinylated conjugates, of various sulfoforms of the trisaccharide ß-D-GlcpA-(1→3)-ß-D-Galp-(1→3)-ß-D-Galp, structures encountered in the linkage region of proteoglycans, is reported herein for the first time. An efficient and stereocontrolled preparation was achieved using common key intermediates in a divergent manner. These molecules should be useful probes to study the substrate specificity of the glycosyltransferases involved at the bifurcation point in the biosynthesis of proteoglycans.