Decorin modulates collagen matrix assembly and mineralization

Abstract

Decorin (DCN) is one of the major matrix proteoglycans in bone. To investigate the role of DCN in matrix mineralization, the expression of DCN in MC3T3-E1 (MC) cell cultures and the phenotypes of MC-derived clones expressing higher (sense; S-DCN) or lower (antisense; AS-DCN) levels of DCN were characterized. DCN expression was significantly decreased as the mineralized nodules were formed and expanded in vitro. In S-DCN clones, in vitro matrix mineralization was inhibited, whereas in AS-DCN clones, mineralization was accelerated. At the microscopic level, collagen fibers in S-DCN clones were thinner while those of AS-DCN clones were thicker and lacked directionality compared to the controls. At the ultrastructural level, the collagen fibrils in S-DCN clones were markedly thinner, whereas those of AS-DCN clones were larger and irregular in shape. The results from Fourier transform infrared spectroscopy analysis demonstrated that in AS-DCN cultures the mineral content was greater but the crystallinity of mineral was poorer than that of the controls at early stage of mineralization. The in vivo transplantation assay demonstrated that no mineralized matrices were formed in S-DCN transplants, whereas they were readily detected in AS-DCN transplants at 3 weeks of transplantation. The areas of bone-like matrices in AS-DCN transplants were significantly greater than the controls at 3 weeks but became comparable at 5 weeks. The bonelike matrices in AS-DCN transplants exhibited woven bone-like non-lamellar structure while the lamellar bone-like structure was evident in the control transplants. These results suggest that DCN regulates matrix mineralization by modulating collagen assembly.

Lysyl Hydroxylase 3-mediated Glucosylation in Type I Collagen

“…Although the values of the cross-links at week 2 showed some variation within the groups, those at week 4 were similar to one another (12). The slight phenotypic difference between EV and MC, possibly due to the transfection effect, has also been reported in other studies (52,54). In all of the Sh clones, the mean fibril diameters and their ranges were significantly larger when compared with those of the controls, MC and EV (p Ͻ 0.05).…”

“…It is not clear what causes the delay. This could be due to an altered interaction between collagen and non-collagenous proteins important for the initiation of mineralization (54,76,77), which could be caused by lower levels of GG-Hyl. Based on the molecular loci of glycosylation identified by MS, the sites are located in close proximity to or in the gap zone, which is the putative site for the initiation of mineralization (78,79).…”

“…Upon confluence, cells were maintained in the mineralization medium containing 50 g/ml ascorbic acid and 2 mM ␤-glycerophosphate and cultured for up to 4 weeks. At the end of each week, the cell/matrix layer from each sample were washed with PBS, fixed with 100% methanol, and stained with 1% Alizarin Red S (Sigma-Aldrich), which binds to calcium in the mineralized nodules deposited (52)(53)(54). The rate and the extent of mineral deposition among the clones with varying levels of LH3 enzyme were compared.…”

TLR signalling can modify the mineralization of tooth germ

“…Several studies have investigated the role of TLR4 signalling in relation to dentin mineralization of the mature tooth. [40,41] According to the literature, LPS treatment can decrease the mineralization of the hard tissues through the modification of non-tissue-specific alkaline phosphatase activity. [13,24] Lower incisors of E16.5 NMRI mice have been used in in vitro culturing to quantitatively analyse the ameloblastin expression level at the end of 5 days long culturing, in order the screen the effects of LPS on function of the secretory ameloblasts.…”

“…We propose two possible reasons: first, the downstream signalling pathway of TLR4 may decrease the ALP activity of dental cells and osteoblasts; second, LPS may bind Ca 2+ that inhibits the incorporation of the mineral phase into the dentin and enamel matrix. [40,41] The absence of free Ca 2+ ions has been reported to inhibit ameloblast maturation. [43] The absence of free Ca 2+ ions contradicts the result of in situ hybridization and RTqPCR, which show the acceleration of ameloblast maturation.…”