Matrix (Stuttgart, Germany)最新文献

A porous hydroxyapatite was used as a morphogenetic matrix to study early tissue formation preceding the morphogenesis of bone in extraskeletal sites of the baboon (Papio ursinus). Porous hydroxyapatites, obtained by hydrothermal conversion of the calcium carbonate exoskeleton of coral, were implanted extraskeletally in 16 baboons. Specimens were harvested at days 30, 60 and 90, and processed to obtain decalcified sections for histomorphometry, and undecalcified sections for enzyme histochemical demonstration of alkaline phosphatase, immunohistochemical demonstration of laminin and type I collagen, and for comparative histologic analysis. At day 30, the tissue that invaded the porous spaces showed mesenchymal condensations at the hydroxyapatite interface, and prominent vascular penetration. Collagen type I staining was localized within mesenchymal condensations. Bone had not formed in any specimen harvested at day 30. At days 30 and 60, alkaline phosphatase staining was initially localized in the invading vasculature, and subsequently found in cellular condensations prior to their transformation into bone, and in capillaries close to cellular condensations. Laminin staining was localized around invading capillaries adjacent to and within mesenchymal condensations, and in capillaries in direct contact with the hydroxyapatite. Bone had formed by day 60; cartilage, however, was never observed. By day 90, bone formation within the porous spaces was often extensive. Goldner's trichrome stain and fluorescence microscopy of tetracycline-labeled specimens demonstrated nascent mineralization within condensations during initial bone morphogenesis. Coating the hydroxyapatite with collagen type I prepared from baboon bone did not increase the amount of bone formation. In this hydroxyapatite-induced osteogenesis model in primates, vascular invasion and bone differentiation appear to be accompanied by a specific temporal sequence of alkaline phosphatase expression. The differentiation of osteogenic cells in direct apposition to the hydroxyapatite suggests that this substratum may act as a solid state matrix for adsorption and controlled release of endogenously-produced bone morphogenetic proteins. The porous hydroxyapatite, as used in this bioassay in primates, may be an appropriate delivery system for bone morphogenetic proteins for the controlled inititiation of therapeutic osteogenesis.

Extracellular matrix proteins are a diverse family of secreted proteins and glycoproteins that are responsible for a variety of critical functions in different tissues. A large number of multiexon genes encode these proteins of the extracellular matrix. Over the last few years, it has become evident that the processing of the pre-mRNA from several of these genes involves alternative splicing. This review summarizes the known examples of alternative splicing in genes coding for the extracellular matrix and attempts to relate the increase in coding diversity generated by alternate exon usage to the function(s) of individual extracellular matrix proteins.

Bone sialoprotein (BSP) is a highly glycosylated and sulphated phosphoprotein that is a major non-collagenous protein of bone. To further characterize the porcine protein and to study its expression during bone formation BSP cDNA clones were isolated from a porcine bone cDNA library. The primary sequence of the protein was derived from the nucleotide sequence of the largest cDNA insert and from the amino-terminal amino acid sequence determined by the automated Edman degradation procedure. When compared with sequences obtained from the human and rat BSPs 74% and 64% of the amino acids, respectively, were identical and a further 11 % and 17%, respectively, were conservative replacements. Moreover, 60% of the amino acids in a concensus sequence derived from the primary sequences of mammalian BSPs were conserved with 16% conservative replacements. The two stretches of polyglutamic acid, through which the protein is capable of binding to hydroxyapatite, and an RGD motif that mediates cell attachment are retained in conserved sequences as are a number of potential sites of serine, threonine and tyrosine phosphorylation, glycosylation and tyrosine sulphation. Secondary structure prediction and hydrophilicity analysis indicate that the nascent BSP has an open flexible structure with the potential to form significant amounts of α-helix and some β-sheet. In situ hybridization of fetal porcine bone with cRNA probes to porcine BSP mRNA shows that BSP is specifically expressed in differentiated osteoblasts on the surface of newly-forming bone trabeculae with especially high levels of hybridization at sites of de novo bone formation. The highly conserved features of BSP and its restricted distribution indicate an important role for this sialoprotein in the formation of bone.

Biochemical determinations of the collagen and elastin content in 50 mm³ samples of human lung are presented in relation to morphometric measurements of lung structure, as the amount of alveolar wall surface area per unit volume (A WUV), on adjacent slices. There were no differences in A WUV values, collagen content (determined as hydroxyproline) or elastin content (determined as isodesmosine) between upper and lower lobes within a single lung. In a study of 102 samples from 9 smokers lungs with no evidence of macro- or microscopic emphysema (as estimated by A WUV measurement), there was a negative correlation between A WUV and the amounts of collagen or elastin per unit volume of inflated lung. The correlation was stronger when collagen and elastin content were expressed per unit area of alveolar wall. The negative correlation is interpreted as representing either the anatomical variation within the complex hierarchy of normal lung structure or possibly low levels of fibrosis in response to cigarette smoking.

We used a rat model to correlate age, matrix gene expression and lysyl oxidase activity in three connective tissues, skin, aorta and lung. By in situ hybridization, we showed that intense collagen type I and elastin mRNA expression were limited to a brief postnatal period. Although there were some organ-specific differences, the mRNA abundance for these two scleroproteins drastically diminished with time. Thus, the majority of mesenchymal cells in young (60days) and old (720 days) animals, appeared to be in a quiescent state, consistent with the slow turnover of these two scleroproteins. We also measured the activity of lysyl oxidase, an enzyme which plays a crucial role in the formation of crosslinks in both pro collagen and tropoelastin molecules. In all the organs investigated, we observed a tissue-dependent pattern of activity. Moreover in this study we focused on the importance of gene matrix expression in evaluating lysyl oxidase activity of aging tissues.

The expression of pro-_α1 (III) and pro-_α1 (I) collagen mRNAs in mouse and human dental tissues during tooth development and after its completion was analyzed by in situ hybridization, with use of [³⁵S]-labeled RNA probes. The expression of pro-_α1 (III) mRNA was also compared to that of the protein product, as localized by immunostaining with polyclonal antibodies to type III collagen and the N-terminal propeptide of type III procollagen. Contrary to many previous reports, our results suggest that odontoblasts express type III collagen. While proal (III) transcripts were less intensely expressed in odontoblasts than pro-_α1 (I) transcripts, the amounts of both mRNAs increased in odontoblasts with progressing dentin formation, and decreased toward its completion. In contrast to pro-_α1 (III) mRNA, pro-_α1 (I) mRNA was still detectable in odontoblasts of fully developed teeth. Type III collagen immunoreactivity was observed in the early predentin, and again in predentin toward the completion of dentinogenesis, when mRNA was no longer detected. Also in the pulp, the protein product, unlike pro-_α1 (III) mRNA, was relatively strongly expressed. Hence, these immunostaining patterns were inversely related to the expression of pro-_α1 (III) mRNA, suggesting accumulation of the protein. The mesenchymal cells, when condensed in the region of the future mandibular bone, expressed proal (III) mRNA intensely, whereas osteoblasts expressed pro-_α1 (I) but not pro_α1 (III) transcripts strongly. Cell type- and developmental stage-related differences in the expression of the two mRNAs suggest that type I1type III collagen ratio influences the structure of dental tissues.