Distribution Of Matrix Components Research Articles

Identification of Extracellular Matrix Components and Biological Factors in Micronized Dehydrated Human Amnion/Chorion Membrane.

Objective: The use of bioactive extracellular matrix (ECM) grafts such as amniotic membranes is an attractive treatment option for enhancing wound repair. In this study, the concentrations, activity, and distribution of matrix components, growth factors, proteases, and inhibitors were evaluated in PURION® Processed, micronized, dehydrated human amnion/chorion membrane (dHACM; MiMedx Group, Inc.).Approach: ECM components in dHACM tissue were assessed by using immunohistochemical staining, and growth factors, cytokines, proteases, and inhibitors were quantified by using single and multiplex ELISAs. The activities of proteases that were native to the tissue were determined via gelatin zymography and EnzChek® activity assay.Results: dHACM tissue contained the ECM components collagens I and IV, hyaluronic acid, heparin sulfate proteoglycans, fibronectin, and laminin. In addition, numerous growth factors, cytokines, chemokines, proteases, and protease inhibitors that are known to play a role in the wound-healing process were quantified in dHACM. Though matrix metalloproteinases (MMPs) were present in dHACM tissues, inhibitors of MMPs overwhelmingly outnumbered the MMP enzymes by an overall molar ratio of 28:1. Protease activity assays revealed that the MMPs in the tissue existed primarily either in their latent form or complexed with inhibitors.Innovation: This is the first study to characterize components that function in wound healing, including inhibitor and protease content and activity, in micronized dHACM.Conclusion: A variety of matrix components and growth factors, as well as proteases and their inhibitors, were identified in micronized dHACM, providing a better understanding of how micronized dHACM tissue can be used to effectively promote wound repair.

Mollusk shell formation: Mapping the distribution of organic matrix components underlying a single aragonitic tablet in nacre

Control over mineral formation in mollusk shells is exerted by the macromolecules of the organic matrix. Using histochemical methods, we mapped the carboxylates and sulfates of proteins and polysaccharides on the surfaces of decalcified interlamellar matrices from the nacreous shell layer of the cephalopod Nautilus pompilius, expanding upon an earlier study by Crenshaw and Ristedt [Crenshaw, M.A., Ristedt, H., 1976. The histochemical localization of reactive groups in septal nacre from Nautilus pompilius. In: Watabe, N., Wilbur, K.M. (Ed.), The Mechanisms of Mineralization in the Invertebrates and Plants. University of South Carolina Press, Colombia, pp. 355–367]. We observed four different zones underlying a single crystal: (1) a central spot rich in carboxylates; (2) a central ring-shaped area rich in sulfates; (3) an area between the central nucleation region and the imprint periphery containing carboxylates, and (4) the intertabular matrix, rich in carboxylates and sulfates. We also mapped matrix functional groups on the nacreous matrix surfaces of the bivalve Atrina rigida, but did not identify well-defined zones. Immuno-mapping of the constituents of the aragonite-nucleating protein fraction from Atrina nacre showed that these macromolecules are located both in the intertabular matrix and in the center of the crystal imprints for both Atrina and Nautilus matrix surfaces. Their presence at the latter location is consistent with their purported role in aragonite nucleation. The observed differentiation in the distribution of matrix components and their functional groups shows that the different stages of single crystal growth are highly controlled by the matrix.

Homogeneity in the distribution of matrix components in the hamster zona pellucida as revealed by backscattered electron imaging fracture‐label

Backscattered electron imaging fracture-label (BEI-FL), an adaptation of the fracture-label method for scanning electron microscopy, offers the advantage of providing information about the distribution of antigenic and receptor sites with respect to the three-dimensional organization of tissues and cells over relatively large surfaces. Recently, using post-embedding cytochemistry on thin-sections of Lowicryl-embedded oocytes, a homogenous distribution of glycoproteins in the zona pellucida (ZP) was demonstrated (Kan et al., 1989. Biol. Reprod., 40:585-598, Anat. Rec., 226:37-47; Roux and Kan, 1991. Anat. Rec., 230:347-360). However, it can be argued that the chemical nature of resins and the physical conditions of tissue processing required for post-embedding cytochemistry may introduce changes in the tissue components and result in altered distribution of components. On the other hand, freeze-fracture exposes constituents in a minimally denaturing manner and, since no embedding media are used, binding sites are sterically available to the probe. We have, therefore, applied BEI-FL to examine the distribution of matrix glycoproteins in the ZP of hamster oocytes. Ovaries and cumulus masses obtained from superovulated female golden hamsters were fixed by immersion in 2.5% glutaraldehyde and processed for fracture-label. Tissues were labeled, respectively, with Wheat germ agglutinin (WGA) followed by ovomucoid-colloidal gold, Ricinus communis agglutinin I (RCA I)-colloidal gold or a monoclonal antibody against Hamster Oviductin-1 followed by protein A-gold, and then examined in the scanning electron microscope. Backscattered electron imaging revealed a homogenous distribution of WGA and RCA I binding sites throughout the cross-fractured matrix of the ZP of ovarian and postovulatory oocytes. Hamster Oviductin-1, an oviductal glycoprotein which is transferred to the ZP of oocytes during oviductal transit, was also found to be uniformly distributed throughout the ZP of postovulatory oocytes. Our results indicate that BEI-FL can be advantageously used to examine extracellular matrices and are consistent with the concept that glycoproteins are uniformly distributed throughout the ZP of the hamster oocyte.

Rat chromosome 5 (q22–23) contains elements that control cell morphology and interactions with the extracellular matrix: A study of normal fibroblast X malignant hepatoma cell hybrids

Cell interactions with the extracellular matrix are consistently modified in neoplasia. Malignant transformation has been correlated with modifications in the synthesis and distribution of matrix components and with alterations of cell adhesive properties to these components. A particular class of genes, able to suppress the transformed phenotype in normal cells, may be involved in those phenotypic changes. By studying somatic cell hybrids between mouse hepatoma (BWTG3) cells and normal rat skin fibroblasts (RSF), Islam and co-workers were able to localize a gene or a group of genes controlling anchorage dependence and cell growth in vitro. This (or these) gene(s) was (were) assigned to the q22–23 fragment of rat chromosome 5. In the present study, we compare the morphology and the interactions with the extracellular matrix proteins (laminin, fibronectin, and collagen IV) and the synthesis of these proteins by RSF X BWTG3 hybrid cells that had either retained (BS181p 10) or lost (BS181a 5) the q22–23 region of rat chromosome 5. Our results suggest that the rat 5q22–23 fragment controls a part of the cell differentiation program including morphology, attachment to extracellular matrix, and synthesis of some matrix proteins, particularly α1 and α2 chains of collagen IV.

Laminin-like glycoproteins in extracellular matrix of endodermal cells

Extracellular matrix from a mouse endodermal cell line consisted mainly of two polypeptides with molecular weights of about 200,000 (200K) and 400,000 (400K). Both poly-peptides incorporated radioactivity from [ 3H]proline and [ 3H]glucosamine and were solubilized from the matrix by treatment with bacterial collagenase or 0.5 m sodium chloride. These polypeptides appeared similar to those of laminin ( R. Timpl, H. Rohde, P. G. Robey, S. I. Rennard, J.-M. Foidart, and G. R. Martin, 1979, J. Biol. Chem., 254, 9933–9937) in polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate but the laminin polypeptides seemed slightly larger than the 200K and 400K polypeptides, respectively. The amino acid compositions of the isolated 200K and 400K polypeptides resembled one another and the previously published amino acid composition of laminin. Antibodies prepared against the solubilized extracellular matrix protein (mixture of 200K and 400K components) as well as those against the isolated 400K component precipitated both the 400K component and the 200K component from culture media. These antisera and antisera to laminin showed identical reactivities in immunodiffusion and in immunofluorescence of tissue sections where they stained basement membranes. The immunofluorescent staining pattern was similar to that obtained with antifibronectin except in the liver where antifibronectin stained the biliary ducts and the liver sinusoids, while laminin-like immunoreactivity was not present in the sinusoidal areas. Such differences in distribution of matrix components could be involved in generation of signals for differentiation and growth of the adjacent cells.