Distribution of Basement Membrane Molecules, Laminin and Collagen Type IV, in Normal and Degenerated Cartilage Tissues.

Science.gov (United States)

Foldager, Casper Bindzus; Toh, Wei Seong; Gomoll, Andreas H; Olsen, Bjørn Reino; Spector, Myron

2014-04-01

The objective of the present study was to investigate the presence and distribution of 2 basement membrane (BM) molecules, laminin and collagen type IV, in healthy and degenerative cartilage tissues. Normal and degenerated tissues were obtained from goats and humans, including articular knee cartilage, the intervertebral disc, and meniscus. Normal tissue was also obtained from patella-tibial enthesis in goats. Immunohistochemical analysis was performed using anti-laminin and anti-collagen type IV antibodies. Human and goat skin were used as positive controls. The percentage of cells displaying the pericellular presence of the protein was graded semiquantitatively. When present, laminin and collagen type IV were exclusively found in the pericellular matrix, and in a discrete layer on the articulating surface of normal articular cartilage. In normal articular (hyaline) cartilage in the human and goat, the proteins were found co-localized pericellularly. In contrast, in human osteoarthritic articular cartilage, collagen type IV but not laminin was found in the pericellular region. Nonpathological fibrocartilaginous tissues from the goat, including the menisci and the enthesis, were also positive for both laminin and collagen type IV pericellularly. In degenerated fibrocartilage, including intervertebral disc, as in degenerated hyaline cartilage only collagen type IV was found pericellularly around chondrocytes but with less intense staining than in non-degenerated tissue. In calcified cartilage, some cells were positive for laminin but not type IV collagen. We report differences in expression of the BM molecules, laminin and collagen type IV, in normal and degenerative cartilaginous tissues from adult humans and goats. In degenerative tissues laminin is depleted from the pericellular matrix before collagen type IV. The findings may inform future studies of the processes underlying cartilage degeneration and the functional roles of these 2 extracellular matrix proteins

Distribution of Basement Membrane Molecules, Laminin and Collagen Type IV, in Normal and Degenerated Cartilage Tissues

Science.gov (United States)

Toh, Wei Seong; Gomoll, Andreas H.; Olsen, Bjørn Reino; Spector, Myron

2014-01-01

Objective: The objective of the present study was to investigate the presence and distribution of 2 basement membrane (BM) molecules, laminin and collagen type IV, in healthy and degenerative cartilage tissues. Design: Normal and degenerated tissues were obtained from goats and humans, including articular knee cartilage, the intervertebral disc, and meniscus. Normal tissue was also obtained from patella-tibial enthesis in goats. Immunohistochemical analysis was performed using anti-laminin and anti–collagen type IV antibodies. Human and goat skin were used as positive controls. The percentage of cells displaying the pericellular presence of the protein was graded semiquantitatively. Results: When present, laminin and collagen type IV were exclusively found in the pericellular matrix, and in a discrete layer on the articulating surface of normal articular cartilage. In normal articular (hyaline) cartilage in the human and goat, the proteins were found co-localized pericellularly. In contrast, in human osteoarthritic articular cartilage, collagen type IV but not laminin was found in the pericellular region. Nonpathological fibrocartilaginous tissues from the goat, including the menisci and the enthesis, were also positive for both laminin and collagen type IV pericellularly. In degenerated fibrocartilage, including intervertebral disc, as in degenerated hyaline cartilage only collagen type IV was found pericellularly around chondrocytes but with less intense staining than in non-degenerated tissue. In calcified cartilage, some cells were positive for laminin but not type IV collagen. Conclusions: We report differences in expression of the BM molecules, laminin and collagen type IV, in normal and degenerative cartilaginous tissues from adult humans and goats. In degenerative tissues laminin is depleted from the pericellular matrix before collagen type IV. The findings may inform future studies of the processes underlying cartilage degeneration and the functional

Deletion of the basement membrane heparan sulfate proteoglycan type XVIII collagen causes hypertriglyceridemia in mice and humans.

Directory of Open Access Journals (Sweden)

Joseph R Bishop

2010-11-01

Full Text Available Lipoprotein lipase (Lpl acts on triglyceride-rich lipoproteins in the peripheral circulation, liberating free fatty acids for energy metabolism or storage. This essential enzyme is synthesized in parenchymal cells of adipose tissue, heart, and skeletal muscle and migrates to the luminal side of the vascular endothelium where it acts upon circulating lipoproteins. Prior studies suggested that Lpl is immobilized by way of heparan sulfate proteoglycans on the endothelium, but genetically altering endothelial cell heparan sulfate had no effect on Lpl localization or lipolysis. The objective of this study was to determine if extracellular matrix proteoglycans affect Lpl distribution and triglyceride metabolism.We examined mutant mice defective in collagen XVIII (Col18, a heparan sulfate proteoglycan present in vascular basement membranes. Loss of Col18 reduces plasma levels of Lpl enzyme and activity, which results in mild fasting hypertriglyceridemia and diet-induced hyperchylomicronemia. Humans with Knobloch Syndrome caused by a null mutation in the vascular form of Col18 also present lower than normal plasma Lpl mass and activity and exhibit fasting hypertriglyceridemia.This is the first report demonstrating that Lpl presentation on the lumenal side of the endothelium depends on a basement membrane proteoglycan and demonstrates a previously unrecognized phenotype in patients lacking Col18.

Collective cell behavior on basement membranes floating in space

Science.gov (United States)

Ellison, Sarah; Bhattacharjee, Tapomoy; Morley, Cameron; Sawyer, W.; Angelini, Thomas

The basement membrane is an essential part of the polarity of endothelial and epithelial tissues. In tissue culture and organ-on-chip devices, monolayer polarity can be established by coating flat surfaces with extracellular matrix proteins and tuning the trans-substrate permeability. In epithelial 3D culture, spheroids spontaneously establish inside-out polarity, morphing into hollow shell-like structures called acini, generating their own basement membrane on the inner radius of the shell. However, 3D culture approaches generally lack the high degree of control provided by the 2D culture plate or organ-on-chip devices, making it difficult to create more faithful in vitro tissue models with complex surface curvature and morphology. Here we present a method for 3D printing complex basement membranes covered in cells. We 3D print collagen-I and Matrigel into a 3D growth medium made from jammed microgels. This soft, yielding material allows extracellular matrix to be formed as complex surfaces and shapes, floating in space. We then distribute MCF10A epithelial cells across the polymerized surface. We envision employing this strategy to study 3D collective cell behavior in numerous model tissue layers, beyond this simple epithelial model.

Study of the relationship between mononuclear inflammatory infiltrate and Ki-67 and basement membrane and extracellular matrix protein expression in radicular cysts.

Science.gov (United States)

Mourão, R V C; Júnior, E C Pinheiro; Barros Silva, P G; Turatti, E; Mota, M R L; Alves, A P N N

2016-05-01

To evaluate the relationship between mononuclear inflammatory infiltrate and the expression of a proliferative immunomarker (Ki-67) as well as to evaluate basement membrane and extracellular matrix proteins (laminin and collagen type IV) in radicular cysts and dentigerous cysts (DC). Immunohistochemical analyses were performed in heavily inflamed radicular cysts (HIRC), slightly inflamed radicular cysts (SIRC) and DC (n = 20) using Ki-67 (Dako(®) , 1 : 50), anticollagen type IV (DBS(®) , 1 : 40) and antilaminin (DBS(®) , 1 : 20). The data were analysed using anova/Tukey's test (Ki-67) and Kruskal-Wallis/Dunn's test (collagen type IV and laminin) (P collagen type IV in the basement membrane of the SIRC group was significantly more continuous (P = 0.0475) than in the HIRC group. DC had significantly less collagen type IV in extracellular matrix immunoexpression than HIRC and SIRC (P = 0.0246). Laminin was absent in the basement membrane in the SIRC and DC groups, and the extracellular matrix of the HIRC was weak and punctate. The presence of inflammatory factors in the radicular cyst wall modified the expression of proliferation factors in the epithelial lining and the expression of collagen type IV and laminin in the basement membrane, but did not modify extracellular matrix behaviour in radicular cysts. © 2015 International Endodontic Journal. Published by John Wiley & Sons Ltd.

The Acinar Cage: Basement Membranes Determine Molecule Exchange and Mechanical Stability of Human Breast Cell Acini.

Directory of Open Access Journals (Sweden)

Aljona Gaiko-Shcherbak

Full Text Available The biophysical properties of the basement membrane that surrounds human breast glands are poorly understood, but are thought to be decisive for normal organ function and malignancy. Here, we characterize the breast gland basement membrane with a focus on molecule permeation and mechanical stability, both crucial for organ function. We used well-established and nature-mimicking MCF10A acini as 3D cell model for human breast glands, with ether low- or highly-developed basement membrane scaffolds. Semi-quantitative dextran tracer (3 to 40 kDa experiments allowed us to investigate the basement membrane scaffold as a molecule diffusion barrier in human breast acini in vitro. We demonstrated that molecule permeation correlated positively with macromolecule size and intriguingly also with basement membrane development state, revealing a pore size of at least 9 nm. Notably, an intact collagen IV mesh proved to be essential for this permeation function. Furthermore, we performed ultra-sensitive atomic force microscopy to quantify the response of native breast acini and of decellularized basement membrane shells against mechanical indentation. We found a clear correlation between increasing acinar force resistance and basement membrane formation stage. Most important native acini with highly-developed basement membranes as well as cell-free basement membrane shells could both withstand physiologically relevant loads (≤ 20 nN without loss of structural integrity. In contrast, low-developed basement membranes were significantly softer and more fragile. In conclusion, our study emphasizes the key role of the basement membrane as conductor of acinar molecule influx and mechanical stability of human breast glands, which are fundamental for normal organ function.

Nephritogenic antigen determinants in epidermal and renal basement membranes of kindreds with Alport-type familial nephritis.

Science.gov (United States)

Kashtan, C; Fish, A J; Kleppel, M; Yoshioka, K; Michael, A F

1986-10-01

We probed epidermal basement membranes (EBM) of acid-urea denatured skin from members of kindreds with Alport-type familial nephritis (FN) for the presence of antigens reactive with Goodpasture sera (GPS) and serum (FNS) from an Alport patient who developed anti-glomerular basement membrane (GBM) nephritis in a renal allograft. By immunoblotting, GPS reacted primarily with the 28,000 molecular weight (mol wt) monomer but also the 24,000 mol wt and 26,000 mol wt monomers of the noncollagenous globular domain (NC1) of type IV collagen from normal human GBM, while FNS identified only the 26,000-mol wt monomer. FNS reacted with EBM of 12 controls and nine unaffected male kindred members but not EBM of eight affected males. Five affected females exhibited interrupted reactivity of FNS with EBM. GPS showed variable reactivity with EBM and was not discriminating with respect to Alport-type FN. FNS did not stain renal basement members of five affected males. However, the EBM, tubular basement membrane, and Bowman's capsules of affected males contained antigens reactive with GPS. These immunochemical studies suggest that the FNS antigen is distinct from Goodpasture antigen(s). The expression of FNS antigen located on the NC1 domain of type IV collagen is altered in basement membranes of patients with Alport-type FN, and the distribution of this antigenic anomaly within kindreds suggests X-linked dominant transmission of a defective gene.

Basement Membrane Defects in Genetic Kidney Diseases

Directory of Open Access Journals (Sweden)

Christine Chew

2018-01-01

Full Text Available The glomerular basement membrane (GBM is a specialized structure with a significant role in maintaining the glomerular filtration barrier. This GBM is formed from the fusion of two basement membranes during development and its function in the filtration barrier is achieved by key extracellular matrix components including type IV collagen, laminins, nidogens, and heparan sulfate proteoglycans. The characteristics of specific matrix isoforms such as laminin-521 (α5β2γ1 and the α3α4α5 chain of type IV collagen are essential for the formation of a mature GBM and the restricted tissue distribution of these isoforms makes the GBM a unique structure. Detailed investigation of the GBM has been driven by the identification of inherited abnormalities in matrix proteins and the need to understand pathogenic mechanisms causing severe glomerular disease. A well-described hereditary GBM disease is Alport syndrome, associated with a progressive glomerular disease, hearing loss, and lens defects due to mutations in the genes COL4A3, COL4A4, or COL4A5. Other proteins associated with inherited diseases of the GBM include laminin β2 in Pierson syndrome and LMX1B in nail patella syndrome. The knowledge of these genetic mutations associated with GBM defects has enhanced our understanding of cell–matrix signaling pathways affected in glomerular disease. This review will address current knowledge of GBM-associated abnormalities and related signaling pathways, as well as discussing the advances toward disease-targeted therapies for patients with glomerular disease.

ER stress and basement membrane defects combine to cause glomerular and tubular renal disease resulting from Col4a1 mutations in mice

Directory of Open Access Journals (Sweden)

Frances E. Jones

2016-02-01

Full Text Available Collagen IV is a major component of basement membranes, and mutations in COL4A1, which encodes collagen IV alpha chain 1, cause a multisystemic disease encompassing cerebrovascular, eye and kidney defects. However, COL4A1 renal disease remains poorly characterized and its pathomolecular mechanisms are unknown. We show that Col4a1 mutations in mice cause hypotension and renal disease, including proteinuria and defects in Bowman's capsule and the glomerular basement membrane, indicating a role for Col4a1 in glomerular filtration. Impaired sodium reabsorption in the loop of Henle and distal nephron despite elevated aldosterone levels indicates that tubular defects contribute to the hypotension, highlighting a novel role for the basement membrane in vascular homeostasis by modulation of the tubular response to aldosterone. Col4a1 mutations also cause diabetes insipidus, whereby the tubular defects lead to polyuria associated with medullary atrophy and a subsequent reduction in the ability to upregulate aquaporin 2 and concentrate urine. Moreover, haematuria, haemorrhage and vascular basement membrane defects confirm an important vascular component. Interestingly, although structural and compositional basement membrane defects occurred in the glomerulus and Bowman's capsule, no tubular basement membrane defects were detected. By contrast, medullary atrophy was associated with chronic ER stress, providing evidence for cell-type-dependent molecular mechanisms of Col4a1 mutations. These data show that both basement membrane defects and ER stress contribute to Col4a1 renal disease, which has important implications for the development of treatment strategies for collagenopathies.

Insights into early extracellular matrix evolution: spongin short chain collagen-related proteins are homologous to basement membrane type IV collagens and form a novel family widely distributed in invertebrates.

Science.gov (United States)

Aouacheria, Abdel; Geourjon, Christophe; Aghajari, Nushin; Navratil, Vincent; Deléage, Gilbert; Lethias, Claire; Exposito, Jean-Yves

2006-12-01

Collagens are thought to represent one of the most important molecular innovations in the metazoan line. Basement membrane type IV collagen is present in all Eumetazoa and was found in Homoscleromorpha, a sponge group with a well-organized epithelium, which may represent the first stage of tissue differentiation during animal evolution. In contrast, spongin seems to be a demosponge-specific collagenous protein, which can totally substitute an inorganic skeleton, such as in the well-known bath sponge. In the freshwater sponge Ephydatia mülleri, we previously characterized a family of short-chain collagens that are likely to be main components of spongins. Using a combination of sequence- and structure-based methods, we present evidence of remote homology between the carboxyl-terminal noncollagenous NC1 domain of spongin short-chain collagens and type IV collagen. Unexpectedly, spongin short-chain collagen-related proteins were retrieved in nonsponge animals, suggesting that a family related to spongin constitutes an evolutionary sister to the type IV collagen family. Formation of the ancestral NC1 domain and divergence of the spongin short-chain collagen-related and type IV collagen families may have occurred before the parazoan-eumetazoan split, the earliest divergence among extant animal phyla. Molecular phylogenetics based on NC1 domain sequences suggest distinct evolutionary histories for spongin short-chain collagen-related and type IV collagen families that include spongin short-chain collagen-related gene loss in the ancestors of Ecdyzosoa and of vertebrates. The fact that a majority of invertebrates encodes spongin short-chain collagen-related proteins raises the important question to the possible function of its members. Considering the importance of collagens for animal structure and substratum attachment, both families may have played crucial roles in animal diversification.

ROCK1-directed basement membrane positioning coordinates epithelial tissue polarity.

Science.gov (United States)

Daley, William P; Gervais, Elise M; Centanni, Samuel W; Gulfo, Kathryn M; Nelson, Deirdre A; Larsen, Melinda

2012-01-01

The basement membrane is crucial for epithelial tissue organization and function. However, the mechanisms by which basement membrane is restricted to the basal periphery of epithelial tissues and the basement membrane-mediated signals that regulate coordinated tissue organization are not well defined. Here, we report that Rho kinase (ROCK) controls coordinated tissue organization by restricting basement membrane to the epithelial basal periphery in developing mouse submandibular salivary glands, and that ROCK inhibition results in accumulation of ectopic basement membrane throughout the epithelial compartment. ROCK-regulated restriction of PAR-1b (MARK2) localization in the outer basal epithelial cell layer is required for basement membrane positioning at the tissue periphery. PAR-1b is specifically required for basement membrane deposition, as inhibition of PAR-1b kinase activity prevents basement membrane deposition and disrupts overall tissue organization, and suppression of PAR-1b together with ROCK inhibition prevents interior accumulations of basement membrane. Conversely, ectopic overexpression of wild-type PAR-1b results in ectopic interior basement membrane deposition. Significantly, culture of salivary epithelial cells on exogenous basement membrane rescues epithelial organization in the presence of ROCK1 or PAR-1b inhibition, and this basement membrane-mediated rescue requires functional integrin β1 to maintain epithelial cell-cell adhesions. Taken together, these studies indicate that ROCK1/PAR-1b-dependent regulation of basement membrane placement is required for the coordination of tissue polarity and the elaboration of tissue structure in the developing submandibular salivary gland.

Ultrastructural localization of the core protein of a basement membrane-specific chondroitin sulfate proteoglycan in adult rat skin

DEFF Research Database (Denmark)

McCarthy, K J; Horiguchi, Y; Couchman, J R

1990-01-01

Basement membranes are complex extracellular matrices present at epithelial/mesenchymal interfaces of tissues. The dermal-epidermal junction has been shown to contain numerous components, some of the most well known being laminin, types IV and VII collagens, heparan sulfate proteoglycan, fibronec...

MT1-MMP-mediated basement membrane remodeling modulates renal development

International Nuclear Information System (INIS)

Riggins, Karen S.; Mernaugh, Glenda; Su, Yan; Quaranta, Vito; Koshikawa, Naohiko; Seiki, Motoharu; Pozzi, Ambra; Zent, Roy

2010-01-01

Extracellular matrix (ECM) remodeling regulates multiple cellular functions required for normal development and tissue repair. Matrix metalloproteinases (MMPs) are key mediators of this process and membrane targeted MMPs (MT-MMPs) in particular have been shown to be important in normal development of specific organs. In this study we investigated the role of MT1-MMP in kidney development. We demonstrate that loss of MT1-MMP leads to a renal phenotype characterized by a moderate decrease in ureteric bud branching morphogenesis and a severe proliferation defect. The kidneys of MT1-MMP-null mice have increased deposition of collagen IV, laminins, perlecan, and nidogen and the phenotype is independent of the MT-1MMP target, MMP-2. Utilizing in vitro systems we demonstrated that MTI-MMP proteolytic activity is required for renal tubule cells to proliferate in three dimensional matrices and to migrate on collagen IV and laminins. Together these data suggest an important role for MT1-MMP in kidney development, which is mediated by its ability to regulate cell proliferation and migration by proteolytically cleaving kidney basement membrane components.

Halogens are key cofactors in building of collagen IV scaffolds outside the cell.

Science.gov (United States)

Brown, Kyle L; Hudson, Billy G; Voziyan, Paul A

2018-05-01

The purpose of this review is to highlight recent advances in understanding the molecular assembly of basement membranes, as exemplified by the glomerular basement membrane (GBM) of the kidney filtration apparatus. In particular, an essential role of halogens in the basement membrane formation has been discovered. Extracellular chloride triggers a molecular switch within non collagenous domains of collagen IV that induces protomer oligomerization and scaffold assembly outside the cell. Moreover, bromide is an essential cofactor in enzymatic cross-linking that reinforces the stability of scaffolds. Halogenation and halogen-induced oxidation of the collagen IV scaffold in disease states damage scaffold function. Halogens play an essential role in the formation of collagen IV scaffolds of basement membranes. Pathogenic damage of these scaffolds by halogenation and halogen-induced oxidation is a potential target for therapeutic interventions.

Basement membrane abnormalities in human eyes with diabetic retinopathy

DEFF Research Database (Denmark)

Ljubimov, A V; Burgeson, R E; Butkowski, R J

1996-01-01

Vascular and parenchymal basement membranes (BMs) are thickened in diabetes, but alterations in individual BM components in diabetic eyes, especially in diabetic retinopathy (DR), are obscure. To identify abnormalities in the distribution of specific constituents, we analyzed cryostat sections...... of human eyes obtained at autopsy (seven normal, five diabetic without DR, and 13 diabetic with DR) by immunofluorescence with antibodies to 30 BM and extracellular matrix components. In non-DR eyes, no qualitative changes of ocular BM components were seen. In some DR corneas, epithelial BM was stained...... discontinuously for laminin-1, entactin/nidogen, and alpha3-alpha4 Type IV collagen, in contrast to non-DR corneas. Major BM alterations were found in DR retinas compared to normals and non-DR diabetics. The inner limiting membrane (retinal BM) of DR eyes had accumulations of fibronectin (including cellular...

An in vitro model of the glomerular capillary wall using electrospun collagen nanofibres in a bioartificial composite basement membrane.

Directory of Open Access Journals (Sweden)

Sadie C Slater

Full Text Available The filtering unit of the kidney, the glomerulus, contains capillaries whose walls function as a biological sieve, the glomerular filtration barrier. This comprises layers of two specialised cells, glomerular endothelial cells (GEnC and podocytes, separated by a basement membrane. Glomerular filtration barrier function, and dysfunction in disease, remains incompletely understood, partly due to difficulties in studying the relevant cell types in vitro. We have addressed this by generation of unique conditionally immortalised human GEnC and podocytes. However, because the glomerular filtration barrier functions as a whole, it is necessary to develop three dimensional co-culture models to maximise the benefit of the availability of these cells. Here we have developed the first two tri-layer models of the glomerular capillary wall. The first is based on tissue culture inserts and provides evidence of cell-cell interaction via soluble mediators. In the second model the synthetic support of the tissue culture insert is replaced with a novel composite bioartificial membrane. This consists of a nanofibre membrane containing collagen I, electrospun directly onto a micro-photoelectroformed fine nickel supporting mesh. GEnC and podocytes grew in monolayers on either side of the insert support or the novel membrane to form a tri-layer model recapitulating the human glomerular capillary in vitro. These models will advance the study of both the physiology of normal glomerular filtration and of its disruption in glomerular disease.

Perlecan and basement membrane-chondroitin sulfate proteoglycan (bamacan) are two basement membrane chondroitin/dermatan sulfate proteoglycans in the Engelbreth-Holm-Swarm tumor matrix

DEFF Research Database (Denmark)

Couchman, J R; Kapoor, R; Sthanam, M

1996-01-01

heparan sulfate proteoglycan, widespread in many basement membranes and connective tissues. We now identify two distinct proteoglycan species from this tumor source, which are substituted with galactosaminoglycans and which show basement membrane localization by immunohistochemistry. One species......The presence of proteoglycans bearing galactosaminoglycan chains has been reported, but none has been identified previously in the matrix of the Engelbreth-Holm-Swarm tumor, which is a source of several basement membrane components. This tumor matrix contains perlecan, a large, low buoyant density......-CSPG are distinct in core protein structure. Both are, however, basement membrane components, although there are tissue-specific differences in their distribution....

Collagen mRNA levels changes during colorectal cancer carcinogenesis

DEFF Research Database (Denmark)

Skovbjerg, Hanne; Anthonsen, Dorit; Lothe, Inger M B

2009-01-01

BACKGROUND: Invasive growth of epithelial cancers is a complex multi-step process which involves dissolution of the basement membrane. Type IV collagen is a major component in most basement membranes. Type VII collagen is related to anchoring fibrils and is found primarily in the basement membrane...... zone of stratified epithelia. Immunohistochemical studies have previously reported changes in steady-state levels of different alpha(IV) chains in several epithelial cancer types. In the present study we aimed to quantitatively determine the mRNA levels of type IV collagen (alpha1/alpha 4/alpha 6......) and type VII collagen (alpha1) during colorectal cancer carcinogenesis. METHODS: Using quantitative RT-PCR, we have determined the mRNA levels for alpha1(IV), alpha 4(IV), alpha 6(IV), and alpha1(VII) in colorectal cancer tissue (n = 33), adenomas (n = 29) and in normal tissue from the same individuals...

Synthesis and deposition of basement membrane proteins by primary brain capillary endothelial cells in a murine model of the blood-brain barrier.

Science.gov (United States)

Thomsen, Maj Schneider; Birkelund, Svend; Burkhart, Annette; Stensballe, Allan; Moos, Torben

2017-03-01

The brain vascular basement membrane is important for both blood-brain barrier (BBB) development, stability, and barrier integrity and the contribution hereto from brain capillary endothelial cells (BCECs), pericytes, and astrocytes of the BBB is probably significant. The aim of this study was to analyse four different in vitro models of the murine BBB for expression and possible secretion of major basement membrane proteins from murine BCECs (mBCECs). mBCECs, pericytes and glial cells (mainly astrocytes and microglia) were prepared from brains of C57BL/6 mice. The mBCECs were grown as monoculture, in co-culture with pericytes or mixed glial cells, or as a triple-culture with both pericytes and mixed glial cells. The integrity of the BBB models was validated by measures of transendothelial electrical resistance (TEER) and passive permeability to mannitol. The expression of basement membrane proteins was analysed using RT-qPCR, mass spectrometry and immunocytochemistry. Co-culturing mBCECs with pericytes, mixed glial cells, or both significantly increased the TEER compared to the monoculture, and a low passive permeability was correlated with high TEER. The mBCECs expressed all major basement membrane proteins such as laminin-411, laminin-511, collagen [α1(IV)] 2 α2(IV), agrin, perlecan, and nidogen 1 and 2 in vitro. Increased expression of the laminin α5 subunit correlated with the addition of BBB-inducing factors (hydrocortisone, Ro 20-1724, and pCPT-cAMP), whereas increased expression of collagen IV α1 primarily correlated with increased levels of cAMP. In conclusion, BCECs cultured in vitro coherently form a BBB and express basement membrane proteins as a feature of maturation. Cover Image for this issue: doi: 10.1111/jnc.13789. © 2016 International Society for Neurochemistry.

Removal of the basement membrane enhances corneal wound healing.

Science.gov (United States)

Pal-Ghosh, Sonali; Pajoohesh-Ganji, Ahdeah; Tadvalkar, Gauri; Stepp, Mary Ann

2011-12-01

Recurrent corneal erosions are painful and put patients' vision at risk. Treatment typically begins with debridement of the area around the erosion site followed by more aggressive treatments. An in vivo mouse model has been developed that reproducibly induces recurrent epithelial erosions in wild-type mice spontaneously within two weeks after a single 1.5 mm corneal debridement wound created using a dulled-blade. This study was conducted to determine whether 1) inhibiting MMP9 function during healing after dulled-blade wounding impacts erosion development and 2) wounds made with a rotating-burr heal without erosions. Oral or topical inhibition of MMPs after dulled-blade wounding does not improve healing. Wounds made by rotating-burr heal with significantly fewer erosions than dulled-blade wounds. The localization of MMP9, β4 integrin and basement membrane proteins (LN332 and type VII collagen), immune cell influx, and reinnervation of the corneal nerves were compared after both wound types. Rotating-burr wounds remove the anterior basement membrane centrally but not at the periphery near the wound margin, induce more apoptosis of corneal stromal cells, and damage more stromal nerve fibers. Despite the fact that rotating-burr wounds do more damage to the cornea, fewer immune cells are recruited and significantly more wounds resolve completely. Copyright © 2011 Elsevier Ltd. All rights reserved.

Lack of collagen XVIII/endostatin exacerbates immune-mediated glomerulonephritis.

Science.gov (United States)

Hamano, Yuki; Okude, Takashi; Shirai, Ryota; Sato, Ikumi; Kimura, Ryota; Ogawa, Makoto; Ueda, Yoshihiko; Yokosuka, Osamu; Kalluri, Raghu; Ueda, Shiro

2010-09-01

Collagen XVIII is a component of the highly specialized extracellular matrix associated with basement membranes of epithelia and endothelia. In the normal kidney, collagen XVIII is distributed throughout glomerular and tubular basement membranes, mesangial matrix, and Bowman's capsule. Proteolytic cleavage within its C-terminal domain releases the fragment endostatin, which has antiangiogenic properties. Because damage to the glomerular basement membrane (GBM) accompanies immune-mediated renal injury, we investigated the role of collagen XVIII/endostatin in this disorder. We induced anti-GBM glomerulonephritis in collagen XVIII alpha1-null and wild-type mice and compared the resulting matrix accumulation, inflammation, and capillary rarefaction. Anti-GBM disease upregulated collagen XVIII/endostatin expression within the GBM and Bowman's capsule of wild-type mice. Collagen XVIII/endostatin-deficient mice developed more severe glomerular and tubulointerstitial injury than wild-type mice. Collagen XVIII/endostatin deficiency altered matrix remodeling, enhanced the inflammatory response, and promoted capillary rarefaction and vascular endothelial cell damage, but did not affect endothelial proliferation. Supplementing collagen XVIII-deficient mice with exogenous endostatin did not affect the progression of anti-GBM disease. Taken together, these results suggest that collagen XVIII/endostatin preserves the integrity of the extracellular matrix and capillaries in the kidney, protecting against progressive glomerulonephritis.

[Relationship between the changes in ischemia/reperfusion cerebro-microvessel basement membrane injury and gelatinase system in senile rat].

Science.gov (United States)

Li, Jian-sheng; Liu, Ke; Liu, Jing-xia; Wang, Ming-hang; Zhao, Yue-wu; Liu, Zheng-guo

2008-11-01

To study the relationship of cerebro-microvessel basement membrane injury and gelatinase system after cerebral ischemia/reperfusion (I/R) in aged rats. Cerebral I/R injury model was reproduced by intraluminal silk ligature thrombosis of the middle cerebral artery occlusion (MCAO). Rats were divided randomly into sham control and I/R groups in young rats [ischemia 3 hours (I 3 h) and reperfusion 6 hours (I/R 6 h), 12 hours (I/R 12 h), 24 hours (I/R 24 h), 3 days (I/R 3 d), 6 days (I/R 6 d)], and sham control group and I/R group in aged rats (I 3 h and I/R 6 h, I/R 12 h, I/R 24 h , I/R 3 d, I/R 6 d). The change in cerebro-cortex microvessel basement membrane structure, basement membrane type IV collagen (Col IV) and laminin (LN) contents, matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinases (TIMPs) expression in every group were determined with immunohistochemical method and zymogram analysis. With the increase in age, Col IV and LN contents of the microvessel basement membrane were increased, and MMP-2 and MMP-9 expressions were stronger. With prolongation of I/R, the degradation of microvessel basement membrane components (Col IV and LN) was positively correlated with the duration of cerebral I/R. MMP-2 expression was increased gradually, and MMP-9 and TIMP-1 expression increased at the beginning and decreased subsequently. Col IV(I 3 h, I/R 6 h , I/R 12 h), LN (I 3 h, I/R 6-24 h), MMP-2 (I 3 h, I/R 6 h-6 d) and MMP-9 (I 3 h, I/R 6-24 h) expression level in aged rats with I/R injury were higher, and TIMP-1 (I/R 24 h) expression was lower than those in young rats (Pcerebro-microvessel basement membrane in rats is related with MMPs and TIMP. Cerebro-microvessel basement membrane injury is more serious in aged rats than that of young rats. Changes in cerebro-microvessel basement membrane injury in aged rats is related with gelatinase system change.

Coarctation induces alterations in basement membranes in the cardiovascular system

DEFF Research Database (Denmark)

Lipke, D W; McCarthy, K J; Elton, T S

1993-01-01

ventricular hypertrophy was maximal within 5 days. In immunohistochemical studies, fibronectin and laminin were increased and the basement membrane chondroitin sulfate proteoglycan decreased in both the subendothelial space and smooth muscle cell basement membranes of the aorta above the clip compared...... membrane components in the heart and vasculature peaked before maximal cardiac hypertrophy (5 days). These studies indicate that alterations in basement membrane component deposition in the hypertrophied vasculature occur at both transcriptional and translational levels and suggest that the cell attachment...

Anti-glomerular basement membrane disease superimposed on membranous nephropathy: a case report and review of the literature

Directory of Open Access Journals (Sweden)

Nivera Noel

2010-08-01

Full Text Available Abstract Introduction Anti-glomerular basement membrane disease is a rare autoimmune disorder characterized by pulmonary hemorrhage, crescentic glomerulonephritis and the presence of circulating anti-glomerular basement membrane antibodies. The simultaneous occurrence of both anti-glomerular basement membrane disease and membranous nephropathy is rare. Case presentation A 59-year-old Hispanic man presented with acute onset of nausea and vomiting and was found to have renal insufficiency. Work-up included a kidney biopsy, which revealed anti-glomerular basement membrane disease with underlying membranous nephropathy. He was treated with emergent hemodialysis, intravenous corticosteroids, plasmapheresis, and cyclophosphamide without improvement in his renal function. Conclusion Simultaneous anti-glomerular basement membrane disease and membranous nephropathy is very rare. There have been 16 previous case reports in the English language literature that have been associated with a high mortality and morbidity, and a very high rate of renal failure resulting in hemodialysis. Co-existence of membranous nephropathy and anti-glomerular basement membrane disease may be immune-mediated, although the exact mechanism is not clear.

Immunohistochemical expression of basement membrane proteins of verrucous carcinoma of the oral mucosa.

Science.gov (United States)

Arduino, Paolo G; Carrozzo, Marco; Pagano, Marco; Broccoletti, Roberto; Scully, Crispian; Gandolfo, Sergio

2010-06-01

Squamous cell carcinoma (SCC) of the oral cavity is an extremely invasive tumour of stratified squamous epithelium that spreads throughout degradation of the basement membrane (BM) and extra-cellular matrix. Oral verrucous carcinoma (VC) is a rare low-grade variant of oral SCC that penetrates into the subepithelial connective tissue. It also has a different clinical behaviour from classical oral SCC. We investigated the immunohistochemical expression of laminin, laminin-5, collagen IV and fibronectin in VC, severe epithelial dysplasia (SED) and SCC in order to analyse if the pattern of these molecules expression contributes to the differences in the biological behaviour of these diseases. The staining pattern of laminin was less intensive in SCC compared with SED and VC, and collagen IV expression was increased in VC compared with SED. Discontinuities of laminin, collagen IV and fibronectin were more evident in SED than in VC. This study indicates that VC has a biological behaviour different from SED or SCC, observable by immunohistochemistry in the BM zone.

Force-dependent breaching of the basement membrane.

Science.gov (United States)

Chang, Tammy T; Thakar, Dhruv; Weaver, Valerie M

2017-01-01

Clinically, non-invasive carcinomas are confined to the epithelial side of the basement membrane and are classified as benign, whereas invasive cancers invade through the basement membrane and thereby acquire the potential to metastasize. Recent findings suggest that, in addition to protease-mediated degradation and chemotaxis-stimulated migration, basement membrane invasion by malignant cells is significantly influenced by the stiffness of the associated interstitial extracellular matrix and the contractility of the tumor cells that is dictated in part by their oncogenic genotype. In this review, we highlight recent findings that illustrate unifying molecular mechanisms whereby these physical cues contribute to tissue fibrosis and malignancy in three epithelial organs: breast, pancreas, and liver. We also discuss the clinical implications of these findings and the biological properties and clinical challenges linked to the unique biology of each of these organs. Copyright © 2017 International Society of Matrix Biology. Published by Elsevier B.V. All rights reserved.

Basement membrane and interstitial proteoglycans produced by MDCK cells correspond to those expressed in the kidney cortex

DEFF Research Database (Denmark)

Erickson, A C; Couchman, J R

2001-01-01

Multiple proteoglycans (PGs) are present in all basement membranes (BM) and may contribute to their structure and function, but their effects on cell behavior are not well understood. Their postulated functions include: a structural role in maintaining tissue histoarchitecture, or aid in selective...... filtration processes; sequestration of growth factors; and regulation of cellular differentiation. Furthermore, expression PGs has been found to vary in several disease states. In order to elucidate the role of PGs in the BM, a well-characterized model of polarized epithelium, Madin-Darby canine kidney (MDCK...... core proteins or CS stubs generated by cABC treatment, revealed that both basement membrane and interstitial PGs are secreted by MDCK cells. HSPGs expressed by MDCK cells are perlecan, agrin, and collagen XVIII. Various CSPG core proteins are made by MDCK cells and have been identified as biglycan...

Basement membrane proteoglycans and development

DEFF Research Database (Denmark)

Couchman, J R; Abrahamson, D R; McCarthy, K J

1993-01-01

-CSPG was only strongly expressed in the vasculature invading late comma stage glomeruli, and later in presumptive and mature Bowman's capsule. Over the first six to eight weeks, the capillary basement membranes contained BM-CSPG, but in gradually decreasing amounts until it became completely undetectable...

Mouse endometrial stromal cells produce basement-membrane components

DEFF Research Database (Denmark)

Wewer, U M; Damjanov, A; Weiss, J

1986-01-01

During mouse pregnancy, uterine stromal cells transform into morphologically distinct decidual cells under the influence of the implanting embryo and a proper hormonal environment. Mechanical stimulation of hormonally primed uterine stromal cells leads to the same morphologic alterations. The dec......During mouse pregnancy, uterine stromal cells transform into morphologically distinct decidual cells under the influence of the implanting embryo and a proper hormonal environment. Mechanical stimulation of hormonally primed uterine stromal cells leads to the same morphologic alterations....... Mouse decidual cells isolated from 6- to 7-day pregnant uteri explanted in vitro continue to synthesize basement-membrane-like extracellular matrix. Using immunohistochemistry and metabolic labeling followed by immunoprecipitation, SDS-PAGE, and fluorography, it was shown that the decidual cells...... to undergo pseudodecidualization. We thus showed that stromal cells from pregnant and nonpregnant mouse uteri synthesize significant amounts of basement-membrane components in vitro, and hence could serve as a good model for the study of normal basement-membrane components....

Basement membrane chondroitin sulfate proteoglycans: localization in adult rat tissues

DEFF Research Database (Denmark)

McCarthy, K J; Couchman, J R

1990-01-01

Heparan sulfate proteoglycans have been described as the major proteoglycan component of basement membranes. However, previous investigators have also provided evidence for the presence of chondroitin sulfate glycosaminoglycan in these structures. Recently we described the production...... and characterization of core protein-specific monoclonal antibodies (MAb) against a chondroitin sulfate proteoglycan (CSPG) present in Reichert's membrane, a transient extra-embryonic structure of rodents. This CSPG was also demonstrated to be present in adult rat kidney. We report here the tissue distribution...... of epitopes recognized by these MAb. The ubiquitous presence of these epitopes in the basement membranes of nearly all adult rat tissues demonstrates that at least one CSPG is a constituent of most basement membranes, and by virtue of its unique distribution is distinct from other chondroitin and dermatan...

Collagens - structure, function and biosynthesis.

OpenAIRE

Gelse, K; Poschl, E; Aigner, T

2003-01-01

The extracellular matrix represents a complex alloy of variable members of diverse protein families defining structural integrity and various physiological functions. The most abundant family is the collagens with more than 20 different collagen types identified so far. Collagens are centrally involved in the formation of fibrillar and microfibrillar networks of the extracellular matrix, basement membranes as well as other structures of the extracellular matrix. This review focuses on the dis...

Nanoscale protein architecture of the kidney glomerular basement membrane

Science.gov (United States)

Suleiman, Hani; Zhang, Lei; Roth, Robyn; Heuser, John E; Miner, Jeffrey H; Shaw, Andrey S; Dani, Adish

2013-01-01

In multicellular organisms, proteins of the extracellular matrix (ECM) play structural and functional roles in essentially all organs, so understanding ECM protein organization in health and disease remains an important goal. Here, we used sub-diffraction resolution stochastic optical reconstruction microscopy (STORM) to resolve the in situ molecular organization of proteins within the kidney glomerular basement membrane (GBM), an essential mediator of glomerular ultrafiltration. Using multichannel STORM and STORM-electron microscopy correlation, we constructed a molecular reference frame that revealed a laminar organization of ECM proteins within the GBM. Separate analyses of domains near the N- and C-termini of agrin, laminin, and collagen IV in mouse and human GBM revealed a highly oriented macromolecular organization. Our analysis also revealed disruptions in this GBM architecture in a mouse model of Alport syndrome. These results provide the first nanoscopic glimpse into the organization of a complex ECM. DOI: http://dx.doi.org/10.7554/eLife.01149.001 PMID:24137544

[Collagen nephritis].

Science.gov (United States)

Lago, N R; Bulos, M J; Monserrat, A J

1997-01-01

Fibrillar collagen in the glomeruli is considered specific of the nail-patella syndrome. A new nephropathy with diffuse intraglomerular deposition of type III collagen without nail and skeletal abnormalities has been described. We report the case of a 26-year-old woman who presented persistent proteinuria, hematuria, deafness without nail and skeletal abnormalities. The renal biopsy showed focal and segmental glomerulosclerosis by light microscopy. The electron microscopy revealed the presence of massive fibrillar collagen within the mesangial matriz and the basement membrane. This is the first patient reported in our country. We emphasize the usefulness of electron microscopy in the study of glomerular diseases.

Convective influx/glymphatic system: tracers injected into the CSF enter and leave the brain along separate periarterial basement membrane pathways.

Science.gov (United States)

Albargothy, Nazira J; Johnston, David A; MacGregor-Sharp, Matthew; Weller, Roy O; Verma, Ajay; Hawkes, Cheryl A; Carare, Roxana O

2018-05-12

Tracers injected into CSF pass into the brain alongside arteries and out again. This has been recently termed the "glymphatic system" that proposes tracers enter the brain along periarterial "spaces" and leave the brain along the walls of veins. The object of the present study is to test the hypothesis that: (1) tracers from the CSF enter the cerebral cortex along pial-glial basement membranes as there are no perivascular "spaces" around cortical arteries, (2) tracers leave the brain along smooth muscle cell basement membranes that form the Intramural Peri-Arterial Drainage (IPAD) pathways for the elimination of interstitial fluid and solutes from the brain. 2 μL of 100 μM soluble, fluorescent fixable amyloid β (Aβ) were injected into the CSF of the cisterna magna of 6-10 and 24-30 month-old male mice and their brains were examined 5 and 30 min later. At 5 min, immunocytochemistry and confocal microscopy revealed Aβ on the outer aspects of cortical arteries colocalized with α-2 laminin in the pial-glial basement membranes. At 30 min, Aβ was colocalised with collagen IV in smooth muscle cell basement membranes in the walls of cortical arteries corresponding to the IPAD pathways. No evidence for drainage along the walls of veins was found. Measurements of the depth of penetration of tracer were taken from 11 regions of the brain. Maximum depths of penetration of tracer into the brain were achieved in the pons and caudoputamen. Conclusions drawn from the present study are that tracers injected into the CSF enter and leave the brain along separate periarterial basement membrane pathways. The exit route is along IPAD pathways in which Aβ accumulates in cerebral amyloid angiopathy (CAA) in Alzheimer's disease. Results from this study suggest that CSF may be a suitable route for delivery of therapies for neurological diseases, including CAA.

Degradation of type IV collagen by neoplastic human skin fibroblasts

International Nuclear Information System (INIS)

Sheela, S.; Barrett, J.C.

1985-01-01

An assay for the degradation of type IV (basement membrane) collagen was developed as a biochemical marker for neoplastic cells from chemically transformed human skin fibroblasts. Type IV collagen was isolated from basement membrane of Syrian hamster lung and type I collagen was isolated from rat tails; the collagens were radioactively labelled by reductive alkylation. The abilities of normal (KD) and chemically transformed (Hut-11A) human skin fibroblasts to degrade the collagens were studied. A cell-associated assay was performed by growing either normal or transformed cells in the presence of radioactively labelled type IV collagen and measuring the released soluble peptides in the medium. This assay also demonstrated that KD cells failed to synthesize an activity capable of degrading type IV collagen whereas Hut-11A cells degraded type IV collagen in a linear manner for up to 4 h. Human serum at very low concentrations, EDTA and L-cysteine inhibited the enzyme activity, whereas protease inhibitors like phenylmethyl sulfonyl fluoride, N-ethyl maleimide or soybean trypsin inhibitor did not inhibit the enzyme from Hut-11A cells. These results suggest that the ability to degrade specifically type IV collagen may be an important marker for neoplastic human fibroblasts and supports a role for this collagenase in tumor cell invasion

Expression of basement membrane components through morphological changes in the hair growth cycle

DEFF Research Database (Denmark)

Couchman, J R; Gibson, W T

1985-01-01

The amount and distribution of fibronectin associated with hair follicles was found to vary during the hair growth cycle in the rat. Immunocytochemical staining of follicles in mid-late anagen (the growth stage) revealed the presence of fibronectin in the dermal papilla matrix, in the basement...... membrane separating this from the epithelial cells of the hair bulb, and in the basement membrane and connective tissue sheath which underly the cells of the outer root sheath. Early in catagen, the transitional stage, staining of the dermal papilla matrix disappeared. Fibronectin persisted in the basement...

Characterizing nanoscale topography of the aortic heart valve basement membrane for tissue engineering heart valve scaffold design.

Science.gov (United States)

Brody, Sarah; Anilkumar, Thapasimuthu; Liliensiek, Sara; Last, Julie A; Murphy, Christopher J; Pandit, Abhay

2006-02-01

A fully effective prosthetic heart valve has not yet been developed. A successful tissue-engineered valve prosthetic must contain a scaffold that fully supports valve endothelial cell function. Recently, topographic features of scaffolds have been shown to influence the behavior of a variety of cell types and should be considered in rational scaffold design and fabrication. The basement membrane of the aortic valve endothelium provides important parameters for tissue engineering scaffold design. This study presents a quantitative characterization of the topographic features of the native aortic valve endothelial basement membrane; topographical features were measured, and quantitative data were generated using scanning electron microscopy (SEM), atomic force microscopy (AFM), transmission electron microscopy (TEM), and light microscopy. Optimal conditions for basement membrane isolation were established. Histological, immunohistochemical, and TEM analyses following decellularization confirmed basement membrane integrity. SEM and AFM photomicrographs of isolated basement membrane were captured and quantitatively analyzed. The basement membrane of the aortic valve has a rich, felt-like, 3-D nanoscale topography, consisting of pores, fibers, and elevations. All features measured were in the sub-100 nm range. No statistical difference was found between the fibrosal and ventricular surfaces of the cusp. These data provide a rational starting point for the design of extracellular scaffolds with nanoscale topographic features that mimic those found in the native aortic heart valve basement membrane.

Vascular basement membranes as pathways for the passage of fluid into and out of the brain.

Science.gov (United States)

Morris, Alan W J; Sharp, Matthew MacGregor; Albargothy, Nazira J; Fernandes, Rute; Hawkes, Cheryl A; Verma, Ajay; Weller, Roy O; Carare, Roxana O

2016-05-01

In the absence of conventional lymphatics, drainage of interstitial fluid and solutes from the brain parenchyma to cervical lymph nodes is along basement membranes in the walls of cerebral capillaries and tunica media of arteries. Perivascular pathways are also involved in the entry of CSF into the brain by the convective influx/glymphatic system. The objective of this study is to differentiate the cerebral vascular basement membrane pathways by which fluid passes out of the brain from the pathway by which CSF enters the brain. Experiment 1: 0.5 µl of soluble biotinylated or fluorescent Aβ, or 1 µl 15 nm gold nanoparticles was injected into the mouse hippocampus and their distributions determined at 5 min by transmission electron microscopy. Aβ was distributed within the extracellular spaces of the hippocampus and within basement membranes of capillaries and tunica media of arteries. Nanoparticles did not enter capillary basement membranes from the extracellular spaces. Experiment 2: 2 µl of 15 nm nanoparticles were injected into mouse CSF. Within 5 min, groups of nanoparticles were present in the pial-glial basement membrane on the outer aspect of cortical arteries between the investing layer of pia mater and the glia limitans. The results of this study and previous research suggest that cerebral vascular basement membranes form the pathways by which fluid passes into and out of the brain but that different basement membrane layers are involved. The significance of these findings for neuroimmunology, Alzheimer's disease, drug delivery to the brain and the concept of the Virchow-Robin space are discussed.

The central role of vascular extracellular matrix and basement membrane remodeling in metabolic syndrome and type 2 diabetes: the matrix preloaded

Directory of Open Access Journals (Sweden)

Tyagi Suresh C

2005-06-01

Full Text Available Abstract The vascular endothelial basement membrane and extra cellular matrix is a compilation of different macromolecules organized by physical entanglements, opposing ionic charges, chemical covalent bonding, and cross-linking into a biomechanically active polymer. These matrices provide a gel-like form and scaffolding structure with regional tensile strength provided by collagens, elasticity by elastins, adhesiveness by structural glycoproteins, compressibility by proteoglycans – hyaluronans, and communicability by a family of integrins, which exchanges information between cells and between cells and the extracellular matrix of vascular tissues. Each component of the extracellular matrix and specifically the capillary basement membrane possesses unique structural properties and interactions with one another, which determine the separate and combined roles in the multiple diabetic complications or diabetic opathies. Metabolic syndrome, prediabetes, type 2 diabetes mellitus, and their parallel companion (atheroscleropathy are associated with multiple metabolic toxicities and chronic injurious stimuli. The adaptable quality of a matrix or form genetically preloaded with the necessary information to communicate and respond to an ever-changing environment, which supports the interstitium, capillary and arterial vessel wall is individually examined.

The degree of collagen crosslinks in medical collagen membranes determined by water absorption

International Nuclear Information System (INIS)

Braczko, M.; Tederko, A.; Grzybowski, J.

1994-01-01

Collagen membranes were crosslinked by using three agents: glutaraldehyde, hexametylenediisocyanate, and UV irradiation. The increasing concentrations of above chemical agents or longer time of UV exposition resulted in the higher cross-links degree and in the decrease of collagen membranes swelling (measured as water absorption), their elasticity and mechanical resistance. According to American standards, the degree of collagen biomaterial cross-links is determined by measuring of the digestion time by pepsin. However, that method is very time-consuming. In our study, we have that a simple, linear regression between logarithm of digestion time by pepsin exists and it was identical for all three cross-linking agents used. We have concluded that determination of water absorption can be an alternative, simple and fast method for examination of collagen membrane cross-links degree. (author). 16 refs, 7 figs, 1 tab

A Case of Alport Syndrome with Posttransplant Antiglomerular Basement Membrane Disease despite Negative Antiglomerular Basement Membrane Antibodies by EIA Treated with Plasmapheresis and Intravenous Immunoglobulin

Directory of Open Access Journals (Sweden)

Sumiko I. Armstead

2013-01-01

Full Text Available Posttransplant antiglomerular basement membrane (anti-GBM disease occurs in approximately 5% of Alport patients and usually ends in irreversible graft failure. Recent research has focused on characterizing the structure of the anti-GBM alloepitope. Here we present a case of a 22-year-old male with end-stage renal disease secondary to Alport syndrome, with a previously failed renal allograft, who received a second deceased-donor kidney transplant. Six days after transplantation, he developed acute kidney injury. The serum anti-GBM IgG was negative by enzyme immunoassay (EIA. On biopsy, he had crescentic glomerulonephritis with linear GBM fixation of IgG. With further analysis by western blotting, we were able to detect antibodies to an unidentified protein from the basement membrane. This patient was treated with plasmapheresis twice per week and monthly intravenous immunoglobulin (IVIG for a total of five months. At the end of treatment, these unknown antibodies were no longer detected. His renal function improved, and he has not required dialysis. We conclude that anti-GBM disease in patients with Alport Syndrome may be caused by circulating antibodies to other components of the basement membrane that are undetectable by routine anti-GBM EIA and may respond to treatment with plasmapheresis and IVIG.

A Case of Alport Syndrome with Posttransplant Antiglomerular Basement Membrane Disease despite Negative Antiglomerular Basement Membrane Antibodies by EIA Treated with Plasmapheresis and Intravenous Immunoglobulin.

Science.gov (United States)

Armstead, Sumiko I; Hellmark, Thomas; Wieslander, Jorgen; Zhou, Xin J; Saxena, Ramesh; Rajora, Nilum

2013-01-01

Posttransplant antiglomerular basement membrane (anti-GBM) disease occurs in approximately 5% of Alport patients and usually ends in irreversible graft failure. Recent research has focused on characterizing the structure of the anti-GBM alloepitope. Here we present a case of a 22-year-old male with end-stage renal disease secondary to Alport syndrome, with a previously failed renal allograft, who received a second deceased-donor kidney transplant. Six days after transplantation, he developed acute kidney injury. The serum anti-GBM IgG was negative by enzyme immunoassay (EIA). On biopsy, he had crescentic glomerulonephritis with linear GBM fixation of IgG. With further analysis by western blotting, we were able to detect antibodies to an unidentified protein from the basement membrane. This patient was treated with plasmapheresis twice per week and monthly intravenous immunoglobulin (IVIG) for a total of five months. At the end of treatment, these unknown antibodies were no longer detected. His renal function improved, and he has not required dialysis. We conclude that anti-GBM disease in patients with Alport Syndrome may be caused by circulating antibodies to other components of the basement membrane that are undetectable by routine anti-GBM EIA and may respond to treatment with plasmapheresis and IVIG.

Cdc42 expression in keratinocytes is required for the maintenance of the basement membrane in skin

DEFF Research Database (Denmark)

Wu, Xunwei; Quondamatteo, Fabio; Brakebusch, Cord

2006-01-01

, structure and number of hemidesomosomes were not significantly changed in the Cdc42 mutant skin compared with the control mice and no blister formation was observed in mutant skin. These data indicate that Cdc42 in keratinocytes is important for maintenance of the basement membrane of skin....... process, which requires directed secretion, deposition and organization of basement membrane components at the basal side of epithelial cells. In the current study, we analyzed the maintenance of skin basement membrane in mice with a keratinocyte-restricted deletion of the Cdc42 gene. In the absence...

Basement membrane proteoglycans in glomerular morphogenesis: chondroitin sulfate proteoglycan is temporally and spatially restricted during development

DEFF Research Database (Denmark)

McCarthy, K J; Bynum, K; St John, P L

1993-01-01

We previously reported the presence of a basement membrane-specific chondroitin sulfate proteoglycan (BM-CSPG) in basement membranes of almost all adult tissues. However, an exception to this ubiquitous distribution was found in the kidney, where BM-CSPG was absent from the glomerular capillary......, the present study used light and electron microscopic immunohistochemistry to examine the distribution of BM-CSPG and basement membrane heparan sulfate proteoglycan (BM-HSPG) during prenatal and postnatal renal development in the rat. Our results show that the temporal and spatial pattern of expression of BM...

Collagens--structure, function, and biosynthesis.

Science.gov (United States)

Gelse, K; Pöschl, E; Aigner, T

2003-11-28

The extracellular matrix represents a complex alloy of variable members of diverse protein families defining structural integrity and various physiological functions. The most abundant family is the collagens with more than 20 different collagen types identified so far. Collagens are centrally involved in the formation of fibrillar and microfibrillar networks of the extracellular matrix, basement membranes as well as other structures of the extracellular matrix. This review focuses on the distribution and function of various collagen types in different tissues. It introduces their basic structural subunits and points out major steps in the biosynthesis and supramolecular processing of fibrillar collagens as prototypical members of this protein family. A final outlook indicates the importance of different collagen types not only for the understanding of collagen-related diseases, but also as a basis for the therapeutical use of members of this protein family discussed in other chapters of this issue.

Evaluation of nanohydroxyapaptite (nano-HA) coated epigallocatechin-3-gallate (EGCG) cross-linked collagen membranes.

Science.gov (United States)

Chu, Chenyu; Deng, Jia; Man, Yi; Qu, Yili

2017-09-01

Collagen is the main component of extracellular matrix (ECM) with desirable biological activities and low antigenicity. Collagen materials have been widely utilized in guided bone regeneration (GBR) surgery due to its abilities to maintain space for hard tissue growth. However, pure collagen lacks optimal mechanical properties. In our previous study, epigallocatechin-3-gallate (EGCG) cross-linked collagen membranes, with better biological activities and enhanced mechanical properties, may promote osteoblast proliferation, but their effect on osteoblast differentiation is not very significant. Nanohydroxyapatite (nano-HA) is the main component of mineral bone, which possesses exceptional bioactivity properties including good biocompatibility, high osteoconductivity and osteoinductivity, non-immunogenicity and non-inflammatory behavior. Herein, by analyzing the physical and chemical properties as well as the effects on promoting bone regeneration, we have attempted to present a novel EGCG-modified collagen membrane with nano-HA coating, and have found evidence that the novel collagen membrane may promote bone regeneration with a better surface morphology, without destroying collagen backbone. To evaluate the surface morphologies, chemical and mechanical properties of pure collagen membranes, epigallocatechin-3-gallate (EGCG) cross-linked collagen membranes, nano-HA coated collagen membranes, nano-HA coated EGCG-collagen membranes, (ii) to evaluate the bone regeneration promoted by theses membranes. In the present study, collagen membranes were divided into 4 groups: (1) untreated collagen membranes (2) EGCG cross-linked collagen membranes (3) nano-HA modified collagen membranes (4) nano-HA modified EGCG-collagen membranes. Scanning electron microscope (SEM) and Fourier transform infrared spectroscopy (FTIR) were used to evaluate surface morphologies and chemical properties, respectively. Mechanical properties were determined by differential scanning calorimeter (DSC

Regulation of the basement membrane by epithelia generated forces

Science.gov (United States)

Tanner, Kandice

2012-12-01

Tumor metastasis involves a progressive loss of tissue architecture and dissolution of structural boundaries between the epithelium and connective tissue. The basement membrane (BM), a specialized network of extracellular matrix proteins forms a barrier that physically restricts pre-invasive lesions such that they remain as local insults. The BM is not a static structure, but one that is constantly regenerated and remodeled in the adult organism. Matrix organization also regulates cell function. Thus alterations in the balance of synthesis, remodeling and proteolytic degradation of the extracellular matrix proteins may contribute to a loss of structural integrity. However, the de novo assembly and maintenance of the complex structural properties of in vivo basement membranes remain elusive. Here, this paper highlights the current understanding on the structural properties and the establishment of the BM, and discusses the potential role of self-generated forces in adult tissue remodeling and the maintenance of the BM as a malignancy suppressor.

Regulation of the basement membrane by epithelia generated forces

International Nuclear Information System (INIS)

Tanner, Kandice

2012-01-01

Tumor metastasis involves a progressive loss of tissue architecture and dissolution of structural boundaries between the epithelium and connective tissue. The basement membrane (BM), a specialized network of extracellular matrix proteins forms a barrier that physically restricts pre-invasive lesions such that they remain as local insults. The BM is not a static structure, but one that is constantly regenerated and remodeled in the adult organism. Matrix organization also regulates cell function. Thus alterations in the balance of synthesis, remodeling and proteolytic degradation of the extracellular matrix proteins may contribute to a loss of structural integrity. However, the de novo assembly and maintenance of the complex structural properties of in vivo basement membranes remain elusive. Here, this paper highlights the current understanding on the structural properties and the establishment of the BM, and discusses the potential role of self-generated forces in adult tissue remodeling and the maintenance of the BM as a malignancy suppressor. (paper)

The collagen receptor uPARAP/Endo180 in tissue degradation and cancer (Review)

DEFF Research Database (Denmark)

Carlsen Melander, Eva Maria; Jürgensen, Henrik J; Madsen, Daniel H

2015-01-01

The collagen receptor uPARAP/Endo180, the product of the MRC2 gene, is a central component in the collagen turnover process governed by various mesenchymal cells. Through the endocytosis of collagen or large collagen fragments, this recycling receptor serves to direct basement membrane collagen...... as well as interstitial collagen to lysosomal degradation. This capacity, shared only with the mannose receptor from the same protein family, endows uPARAP/Endo180 with a critical role in development and homeostasis, as well as in pathological disruptions of the extracellular matrix structure. Important...

A novel functional role of collagen glycosylation

DEFF Research Database (Denmark)

Jürgensen, Henrik J; Madsen, Daniel H; Ingvarsen, Signe

2011-01-01

Collagens make up the most abundant component of interstitial extracellular matrices and basement membranes. Collagen remodeling is a crucial process in many normal physiological events and in several pathological conditions. Some collagen subtypes contain specific carbohydrate side chains......, the function of which is poorly known. The endocytic collagen receptor urokinase plasminogen activator receptor-associated protein (uPARAP)/Endo180 plays an important role in matrix remodeling through its ability to internalize collagen for lysosomal degradation. uPARAP/Endo180 is a member of the mannose...... receptor protein family. These proteins all include a fibronectin type II domain and a series of C-type lectin-like domains, of which only a minor part possess carbohydrate recognition activity. At least two of the family members, uPARAP/Endo180 and the mannose receptor, interact with collagens...

Regeneration of the epidermis and basement membrane of the planarian Dugesia japonica after total-body x irradiation

Energy Technology Data Exchange (ETDEWEB)

Hori, I.

1979-03-01

Fresh-water planarians were studied to examine effects of x rays on regeneration of the epidermis and basement membrane. During early stages of regeneration, free rhabdite-forming cells were associated with the wound epidermis and recruited it. In later stages, however, a gradual degeneration occurred in the epidermis and cells undergoing epithelization decreased in number. Eventually epidermal cells on the wound surface appeared necrotic as evidenced by pyknotic nuclei and vacuolized dense cytoplasm. The entire basement membrane could not be reconstituted in any stage after wounding though its precursor-like material was secreted in the interspace between epidermis and parenchyma. Morphological changes in extracellular products and in the cells surrounding the products suggest that epidermal cells which have covered the wound surface synthesize precursors of the basement membrane. Possible factors of a characteristic perturbation in epithelization and basement membrane formation after total-body irradiation are discussed.

Regeneration of the epidermis and basement membrane of the planarian Dugesia japonica after total-body x irradiation

International Nuclear Information System (INIS)

Hori, I.

1979-01-01

Fresh-water planarians were studied to examine effects of x rays on regeneration of the epidermis and basement membrane. During early stages of regeneration, free rhabdite-forming cells were associated with the wound epidermis and recruited it. In later stages, however, a gradual degeneration occurred in the epidermis and cells undergoing epithelization decreased in number. Eventually epidermal cells on the wound surface appeared necrotic as evidenced by pyknotic nuclei and vacuolized dense cytoplasm. The entire basement membrane could not be reconstituted in any stage after wounding though its precursor-like material was secreted in the interspace between epidermis and parenchyma. Morphological changes in extracellular products and in the cells surrounding the products suggest that epidermal cells which have covered the wound surface synthesize precursors of the basement membrane. Possible factors of a characteristic perturbation in epithelization and basement membrane formation after total-body irradiation are discussed

Chitosan Cross-linked Reconstituted Amniotic Collagen Membrane ...

Indian Academy of Sciences (India)

First page Back Continue Last page Overview Graphics. Chitosan Cross-linked Reconstituted Amniotic Collagen Membrane – An Excellent Cell Substratum. The KERATINOCYTE proliferation and Differentiation into multiple layers is due to the presence of type - IV collagen in the amnion. Cultured FIBROBLASTS had good ...

Dynamic interplay between the collagen scaffold and tumor evolution

DEFF Research Database (Denmark)

Egeblad, Mikala; Rasch, Morten G; Weaver, Valerie M

2010-01-01

and remodeling of the ECM network regulate tissue tension, generate pathways for migration, and release ECM protein fragments to direct normal developmental processes such as branching morphogenesis. Collagens are major components of the ECM of which basement membrane type IV and interstitial matrix type I...... are the most prevalent. Here we discuss how abnormal expression, proteolysis and structure of these collagens influence cellular functions to elicit multiple effects on tumors, including proliferation, initiation, invasion, metastasis, and therapy response....

Human skin basement membrane-associated heparan sulphate proteoglycan: distinctive differences in ultrastructural localization as a function of developmental age

DEFF Research Database (Denmark)

Horiguchi, Y; Fine, J D; Couchman, J R

1991-01-01

was identical to that observed in neonatal and adult human skin. These findings demonstrate that active remodelling of the dermo-epidermal junction occurs during at least the first two trimesters, and affects not only basement membrane-associated structures but also specific antigens.......Recent studies have demonstrated that skin basement membrane components are expressed within the dermo-epidermal junction in an orderly sequence during human foetal development. We have investigated the ultrastructural localization of basement membrane-related antigens in human foetal skin...... at different developmental ages using two monoclonal antibodies to a well-characterized basement membrane-associated heparan sulphate proteoglycan. A series of foetal skin specimens (range, 54-142 gestational days) were examined using an immunoperoxidase immunoelectron microscopic technique. In specimens...

Basement membrane proteoglycans are of epithelial origin in rodent skin

DEFF Research Database (Denmark)

Yamane, Y; Yaoita, H; Couchman, J R

1996-01-01

. For in vivo experiments, pieces of newborn rat epidermis obtained by dispase treatment were grafted onto athymic nude mice. Three and six weeks after grafting, immunofluorescence analysis of the grafted skin was carried out, using monoclonal antibodies specific for rat basement membrane chondroitin sulfate...

Retinoid inhibition of in vitro invasion of human amnion basement membrane by human tumor cells

International Nuclear Information System (INIS)

Fazely, F.

1988-01-01

The effects measured were the inhibition of tumor cell migration through the basement membrane (BM) and tumor cell degradative enzyme activity on 3 H-proline labeled collagenous and non collagenous components of the BM. The human lung carcinoma A549 or the human Ewing's sarcoma TC-106 cell lines treated with retinoids for two days were incubated on the BM in the absence of retinoids. A dose-dependent inhibition of cell invasion was produced by retinoids. Among the retinoids tested the most powerful was retinol acetate which inhibited invasion by 50% of A549 cells at a concentration of 0.09 μg/ml, and TC-106 cells at 0.08 μg/ml. Retinol acetate inhibited A549 and TC-106 cell growth by approximately 50% at levels almost 100-fold higher than those needed for antiinvasive activity. Retinol acetate was about 20 times more potent than retinoic acid and 30 times more than retinol palmitate. Furthermore, A549 cells treated with retinol acetate, under conditions whereby an anti-invasive state was induced,showed an increase in the number of cellular retinoic acid binding proteins (CRABP), a decrease in the activity of type IV collagenase and ectosialyltransferase, and no change in the activity of transglutaminase

Quantitative Proteome Analysis Reveals Increased Content of Basement Membrane Proteins in Arteries from Patients with Type 2 Diabetes and Lower Levels among Metformin Users

DEFF Research Database (Denmark)

Rørdam Preil, Simone; Kristensen, Lars P; Beck, Hans C

2015-01-01

hypothesized that metformin intake influences the protein composition. METHODS AND RESULTS: -We analyzed non-atherosclerotic repair arteries gathered at coronary by-pass operations from 30 patients with type 2 diabetes, as well as from 30 age- and gender-matched non-diabetic individuals. Quantitative proteome......BACKGROUND: -The increased risk of cardiovascular diseases (CVD) in type 2 diabetes has been extensively documented, but the origins of the association remain largely unknown. We sought to determine changes in protein expressions in arterial tissue from patients with type 2 diabetes and moreover...... analysis was done by iTRAQ-labelling and LC-MS/MS analysis on individual arterial samples. The amounts of the basement membrane (BM) components, alpha-1- and alpha-2- type IV collagen, gamma-1- and beta-2-laminin were significantly increased in patients with diabetes. Moreover, the expressions of basement...

Evidence for the existence of multiple heparan sulfate proteoglycans in the human glomerular basement membrane and mesangial matrix

NARCIS (Netherlands)

Groffen, Alexander J A; Hop, Frank W H; Tryggvason, Karl; Dijkman, Henri; Assmann, Karel J M; Veerkamp, Jacques H.; Monnens, Leo A H; Van Den Heuvel, Lambert P W J

1997-01-01

Heparan sulfate proteoglycans (HSPGs) are essential components of the glomerular basement membrane (GBM) carrying a strong anionic charge. A well- characterized extracellular HSPG is perlecan, ubiquitously expressed in basement membranes. A cDNA construct encoding domains I and II of human perlecan

Rac1 is essential for basement membrane-dependent epiblast survival

DEFF Research Database (Denmark)

He, Xiaowen; Liu, Jie; Qi, Yanmei

2010-01-01

During murine peri-implantation development, the egg cylinder forms from a solid cell mass by the apoptotic removal of inner cells that do not contact the basement membrane (BM) and the selective survival of the epiblast epithelium, which does. The signaling pathways that mediate this fundamental...

Tissue-engineered cartilaginous constructs for the treatment of caprine cartilage defects, including distribution of laminin and type IV collagen.

Science.gov (United States)

Jeng, Lily; Hsu, Hu-Ping; Spector, Myron

2013-10-01

The purpose of this study was the immunohistochemical evaluation of (1) cartilage tissue-engineered constructs; and (2) the tissue filling cartilage defects in a goat model into which the constructs were implanted, particularly for the presence of the basement membrane molecules, laminin and type IV collagen. Basement membrane molecules are localized to the pericellular matrix in normal adult articular cartilage, but have not been examined in tissue-engineered constructs cultured in vitro or in tissue filling cartilage defects into which the constructs were implanted. Cartilaginous constructs were engineered in vitro using caprine chondrocyte-seeded type II collagen scaffolds. Autologous constructs were implanted into 4-mm-diameter defects created to the tidemark in the trochlear groove in the knee joints of skeletally mature goats. Eight weeks after implantation, the animals were sacrificed. Constructs underwent immunohistochemical and histomorphometric evaluation. Widespread staining for the two basement membrane molecules was observed throughout the extracellular matrix of in vitro and in vivo samples in a distribution unlike that previously reported for cartilage. At sacrifice, 70% of the defect site was filled with reparative tissue, which consisted largely of fibrous tissue and some fibrocartilage, with over 70% of the reparative tissue bonded to the adjacent host tissue. A novel finding of this study was the observation of laminin and type IV collagen in in vitro engineered cartilaginous constructs and in vivo cartilage repair samples from defects into which the constructs were implanted, as well as in normal caprine articular cartilage. Future work is needed to elucidate the role of basement membrane molecules during cartilage repair and regeneration.

Expression and organization of basement membranes and focal adhesion proteins in pregnant myometrium is regulated by uterine stretch.

Science.gov (United States)

Shynlova, Oksana; Chow, Michelle; Lye, Stephen J

2009-10-01

The mechanisms underlying the preparation of the uterus for labor are not fully understood. We have previously found a significant increase in the expression of messenger RNA (mRNAs) encoding extracellular basement membrane (BM) proteins of the smooth muscle cells (SMCs) in late pregnant rat myometrium. At term, the myometrium is stretched by growing fetuses and these mechanical signals are transmitted from extracellular matrix into SMCs through focal adhesions (FA). The aim of this study was to investigate the effect of gravidity on the expression and spatiotemporal distribution of major BM proteins, laminin-gamma2 and collagen IV, as well as typical FA constituents, vinculin and paxillin, in the myometrium during gestation and parturition, using a unilaterally pregnant rat model. We found that the expression of laminin-gamma2 and collagen IV proteins increased significantly with gestational age (P proteins were not affected. Near term, BM proteins from gravid horn myometrium demonstrated increased extracellular immunostaining and major rearrangement from sporadic protein distribution to organized, continuous, and regular structures surrounding the plasma membrane of each myocyte. Examination of FA proteins revealed that paxillin was translocated from the cytoplasm to the cell periphery, while vinculin was sequestered specifically to FAs. At labor, BM and FA proteins, organized in similar bead-like structures, were localized on opposing sides of SMC plasma membrane into 2 different compartments. We suggest that these stretch-induced changes facilitate formation of stable cell-matrix adhesions and provide the molecular basis for optimal force transduction during labor contractions.

Evaluation of epigallocatechin-3-gallate (EGCG) cross-linked collagen membranes and concerns on osteoblasts

Energy Technology Data Exchange (ETDEWEB)

Chu, Chenyu; Deng, Jia [State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu 610041 (China); Xiang, Lin; Wu, Yingying [State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu 610041 (China); Department of Oral Implantology, West China Hospital of Stomatology, Sichuan University, Chengdu 610041 (China); Wei, Xiawei [State Key Laboratory of Biotherapy and Laboratory for Aging Research, West China Hospital, Sichuan University, and Collaborative Innovation Center for Biotherapy, Chengdu, Sichuan 610041 (China); Qu, Yili [State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu 610041 (China); Department of Oral Implantology, West China Hospital of Stomatology, Sichuan University, Chengdu 610041 (China); Man, Yi, E-mail: manyi780203@126.com [State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu 610041 (China); Department of Oral Implantology, West China Hospital of Stomatology, Sichuan University, Chengdu 610041 (China)

2016-10-01

Collagen membranes have ideal biological and mechanical properties for supporting infiltration and proliferation of osteoblasts and play a vital role in guided bone regeneration (GBR). However, pure collagen can lead to inflammation, resulting in progressive bone resorption. Therefore, a method for regulating the level of inflammatory cytokines at surgical sites is paramount for the healing process. Epigallocatechin-3-gallate (EGCG) is a component extracted from green tea with numerous biological activities including an anti-inflammatory effect. Herein, we present a novel cross-linked collagen membrane containing different concentrations of EGCG (0.0064%, 0.064%, and 0.64%) to regulate the level of inflammatory factors secreted by pre-osteoblast cells; improve cell proliferation; and increase the tensile strength, wettability, and thermal stability of collagen membranes. Scanning electron microscope images show that the surfaces of collagen membranes became smoother and the collagen fiber diameters became larger with EGCG treatment. Measurement of the water contact angle demonstrated that introducing EGCG improved membrane wettability. Fourier transform infrared spectroscopy analyses indicated that the backbone of collagen was intact, and the thermal stability was significant improved in differential scanning calorimetry. The mechanical properties of 0.064% and 0.64% EGCG-treated collagen membranes were 1.5-fold greater than those of the control. The extent of cross-linking was significantly increased, as determined by a 2,4,6-trinitrobenzenesulfonic acid solution assay. The Cell Counting Kit-8 (CCK-8) and live/dead assays revealed that collagen membrane cross-linked by 0.0064% EGCG induced greater cell proliferation than pure collagen membranes. Additionally, real-time polymerase chain reaction and enzyme-linked immunosorbent assay results showed that EGCG significantly affected the production of inflammatory factors secreted by MC3T3-E1 cells. Taken together, our

[Research on ultrasonic permeability of low intensity pulsed ultrasound through PTFE membrane and Bio-Gide collagen membrane].

Science.gov (United States)

Chai, Zhaowu; Zhao, Chunliang; Song, Jinlin; Deng, Feng; Yang, Ji; Gao, Xiang; Liu, Minyi

2013-12-01

The aim of the present study was to detect the transmission rate of ultrasonic low intensity pulsed ultrasound (LIPUS) through polytetrafluoroethylene (PTFE) membrane (Thickness: 0.01 mm) and Bio-Gide collagen membrane, and to provide the basis for the barrier membrane selection on the study of LIPUS combined with guided tissue regeneration (GTR). The ultrasonic (LIPUS, frequency 1.5 MHz, pulse width 200 micros, repetition rate 1.0 kHz) transmission coefficient of the two kinds of barrier membrane were detected respectively through setting ten groups from 10 to 100mW/cm2 every other 10 mW/cm2. We found in the study that the ultrasonic transmission coefficient through 0.01 mm PTFE membrane was 78.1% to 92.%, and the ultrasonic transmission coefficient through Bio-Gide collagen membrane was 43.9% to 55.8%. The ultrasonic transmission coefficient through PTFE membrane was obviously higher than that through Bio-Gide collagen membrane. The transmission coefficient of the same barrier membrane of the ultrasonic ion was statistically different under different powers (P PTFE membrane and Bio-Gide collagen membrane were relatively high. We should select barrier membranes based on different experimental needs, and exercise ultrasonic transmission coefficient experiments to ensure effective power.

Basement membrane reconstruction in human skin equivalents is regulated by fibroblasts and/or exogenously activated keratinocytes.

Science.gov (United States)

El Ghalbzouri, Abdoelwaheb; Jonkman, Marcel F; Dijkman, Remco; Ponec, Maria

2005-01-01

This study was undertaken to examine the role fibroblasts play in the formation of the basement membrane (BM) in human skin equivalents. For this purpose, keratinocytes were seeded on top of fibroblast-free or fibroblast-populated collagen matrix or de-epidermized dermis and cultured in the absence of serum and exogenous growth factors. The expression of various BM components was analyzed on the protein and mRNA level. Irrespective of the presence or absence of fibroblasts, keratin 14, hemidesmosomal proteins plectin, BP230 and BP180, and integrins alpha1beta1, alpha2beta1, alpha3beta1, and alpha6beta4 were expressed but laminin 1 was absent. Only in the presence of fibroblasts or of various growth factors, laminin 5 and laminin 10/11, nidogen, uncein, type IV and type VII collagen were decorating the dermal/epidermal junction. These findings indicate that the attachment of basal keratinocytes to the dermal matrix is most likely mediated by integrins alpha1beta1 and alpha2beta1, and not by laminins that bind to integrin alpha6beta4 and that the epithelial-mesenchymal cross-talk plays an important role in synthesis and deposition of various BM components.

Chitosan-Coated Collagen Membranes Promote Chondrocyte Adhesion, Growth, and Interleukin-6 Secretion

Directory of Open Access Journals (Sweden)

Nabila Mighri

2015-11-01

Full Text Available Designing scaffolds made from natural polymers may be highly attractive for tissue engineering strategies. We sought to produce and characterize chitosan-coated collagen membranes and to assess their efficacy in promoting chondrocyte adhesion, growth, and cytokine secretion. Porous collagen membranes were placed in chitosan solutions then crosslinked with glutaraldehyde vapor. Fourier transform infrared (FTIR analyses showed elevated absorption at 1655 cm-1 of the carbon–nitrogen (N=C bonds formed by the reaction between the (NH2 of the chitosan and the (C=O of the glutaraldehyde. A significant peak in the amide II region revealed a significant deacetylation of the chitosan. Scanning electron microscopy (SEM images of the chitosan-coated membranes exhibited surface variations, with pore size ranging from 20 to 50 µm. X-ray photoelectron spectroscopy (XPS revealed a decreased C–C groups and an increased C–N/C–O groups due to the reaction between the carbon from the collagen and the NH2 from the chitosan. Increased rigidity of these membranes was also observed when comparing the chitosan-coated and uncoated membranes at dried conditions. However, under wet conditions, the chitosan coated collagen membranes showed lower rigidity as compared to dried conditions. Of great interest, the glutaraldehyde-crosslinked chitosan-coated collagen membranes promoted chondrocyte adhesion, growth, and interleukin (IL-6 secretion. Overall results confirm the feasibility of using designed chitosan-coated collagen membranes in future applications, such as cartilage repair.

Rituximab for the treatment of refractory simultaneous anti-glomerular basement membrane (anti-GBM) and membranous nephropathy.

Science.gov (United States)

Bandak, Ghassan; Jones, Bruce A; Li, Jian; Yee, Jerry; Umanath, Kausik

2014-02-01

Antibody-mediated anti-glomerular basement membrane (anti-GBM) disease occurs rarely in the presence of another B-cell disorder, membranous nephropathy. The coexistence of these two autoimmune disorders would be anticipated to require differing, specific therapies targeted to each disease process. We describe a case of concomitant membranous nephropathy and anti-GBM disease in which conventional therapy, including steroids, plasmapheresis and cyclophosphamide, failed to attenuate the anti-GBM disease, yet responded to an alternative treatment of rituximab. This B-cell directed, monoclonal, chimeric antibody treatment substantially reduced anti-GBM antibody titers and led to discontinuation of plasmapheresis, while maintaining the remission of membranous nephropathy and anti-GBM disease.

Nephritogenic antigen determinants in epidermal and renal basement membranes of kindreds with Alport-type familial nephritis.

OpenAIRE

Kashtan, C; Fish, A J; Kleppel, M; Yoshioka, K; Michael, A F

1986-01-01

We probed epidermal basement membranes (EBM) of acid-urea denatured skin from members of kindreds with Alport-type familial nephritis (FN) for the presence of antigens reactive with Goodpasture sera (GPS) and serum (FNS) from an Alport patient who developed anti-glomerular basement membrane (GBM) nephritis in a renal allograft. By immunoblotting, GPS reacted primarily with the 28,000 molecular weight (mol wt) monomer but also the 24,000 mol wt and 26,000 mol wt monomers of the noncollagenous ...

Basement membrane changes in breast cancer detected by immunohistochemical staining for laminin

DEFF Research Database (Denmark)

Albrechtsen, R; Nielsen, M; Wewer, U

1981-01-01

The distribution of the basement membrane glycoprotein laminin was studied by the immunoperoxidase technique in benign and malignant human breast tissue and in axillary lymph nodes from patients with breast cancer. An antiserum prepared against rat laminin was used. The specificity...

The basement membrane constituents in the mouse embryo's tooth. An autoradiographic study

International Nuclear Information System (INIS)

Osman, M.

1987-01-01

Enamel organs isolated from the lower first teeth of 18-days old white mouse embryo by trypsin treatment were used in this study. The organs were cultured during periods of increasing time on a semi-solid medium containing cock serum. In another chase experiments, the organs were cultured on a liquid medium containing proline- 3 H, leucine- 3 H, and glucosamine- 3 H, were studied by autoradiography using both light and electron microscopes. It has been shown that the nature of the culture medium does not apparently interfere with the ability of the enamel to reconstitute the basement membrane. On the other hand, it have been found obvious differences concerning the kinetic of the used isotopes. The results indicate that the turn-over of the basement membrane constituents represents a continuous and homogenous process which continues to take place during, before and after reconstitution. 42 refs. (author)

Agar/collagen membrane as skin dressing for wounds

Energy Technology Data Exchange (ETDEWEB)

Bao Lei; Yang Wei; Mao Xuan; Mou Shansong; Tang Shunqing [Biomedical Engineering Institute, Jinan University, Guangzhou (China)], E-mail: tshunqt@jnu.edu.cn, E-mail: tmuss@jnu.edu.cn

2008-12-15

Agar, a highly hydrophilic polymer, has a special gel property and favorable biocompatibility, but moderate intension strength in an aqueous condition and a low degradation rate. In order to tailor both properties of mechanical intension and degradation, type I collagen was composited with agar in a certain ratio by drying at 50 {sup 0}C or by a freeze-dry process. Glutaraldehyde was chosen as a crosslinking agent, and the most favorable condition for crosslinking was that the weight ratio of agar to glutaraldehyde was 66.7 and the pH value about 5. Dynamic mechanical analysis results showed that the single agar membrane had a modulus value between 640 MPa and 1064 MPa, but it was between 340 MPa and 819 MPa after being composited with type I collagen. It was discovered under an optical microscope that the pores were interconnected in the composite scaffolds instead of the honeycomb-like pores in a single type I collagen scaffold or the laminated gaps in a single agar scaffold. The results of an acute toxicity test disclosed that the composites were not toxic to mice although the composites were crosslinked with a certain concentration of glutaraldehyde. The results of gross examinations showed that when the composite membranes or scaffolds were applied to a repair rabbit skin lesion, the composites had a good repair effect without infection, liquid exudation or visible scar in the lesion covered with them. But in the control group, the autologous skin showed necrosis and there were a lot of scar tissues in the lesion site. H and E staining results showed that the repair tissue was similar to the normal one and very few scaffolds or membranes were left without degradation after 2 or 3 weeks. In conclusion, it is proved that type I collagen increases the toughness of the agar membrane, and the agar/type I collagen composites are promising biomaterials as wound dressings for healing burns or ulcers.

Agar/collagen membrane as skin dressing for wounds

International Nuclear Information System (INIS)

Bao Lei; Yang Wei; Mao Xuan; Mou Shansong; Tang Shunqing

2008-01-01

Agar, a highly hydrophilic polymer, has a special gel property and favorable biocompatibility, but moderate intension strength in an aqueous condition and a low degradation rate. In order to tailor both properties of mechanical intension and degradation, type I collagen was composited with agar in a certain ratio by drying at 50 0 C or by a freeze-dry process. Glutaraldehyde was chosen as a crosslinking agent, and the most favorable condition for crosslinking was that the weight ratio of agar to glutaraldehyde was 66.7 and the pH value about 5. Dynamic mechanical analysis results showed that the single agar membrane had a modulus value between 640 MPa and 1064 MPa, but it was between 340 MPa and 819 MPa after being composited with type I collagen. It was discovered under an optical microscope that the pores were interconnected in the composite scaffolds instead of the honeycomb-like pores in a single type I collagen scaffold or the laminated gaps in a single agar scaffold. The results of an acute toxicity test disclosed that the composites were not toxic to mice although the composites were crosslinked with a certain concentration of glutaraldehyde. The results of gross examinations showed that when the composite membranes or scaffolds were applied to a repair rabbit skin lesion, the composites had a good repair effect without infection, liquid exudation or visible scar in the lesion covered with them. But in the control group, the autologous skin showed necrosis and there were a lot of scar tissues in the lesion site. H and E staining results showed that the repair tissue was similar to the normal one and very few scaffolds or membranes were left without degradation after 2 or 3 weeks. In conclusion, it is proved that type I collagen increases the toughness of the agar membrane, and the agar/type I collagen composites are promising biomaterials as wound dressings for healing burns or ulcers.

Concerted regulation of retinal pigment epithelium basement membrane and barrier function by angiocrine factors.

Science.gov (United States)

Benedicto, Ignacio; Lehmann, Guillermo L; Ginsberg, Michael; Nolan, Daniel J; Bareja, Rohan; Elemento, Olivier; Salfati, Zelda; Alam, Nazia M; Prusky, Glen T; Llanos, Pierre; Rabbany, Sina Y; Maminishkis, Arvydas; Miller, Sheldon S; Rafii, Shahin; Rodriguez-Boulan, Enrique

2017-05-19

The outer blood-retina barrier is established through the coordinated terminal maturation of the retinal pigment epithelium (RPE), fenestrated choroid endothelial cells (ECs) and Bruch's membrane, a highly organized basement membrane that lies between both cell types. Here we study the contribution of choroid ECs to this process by comparing their gene expression profile before (P5) and after (P30) the critical postnatal period when mice acquire mature visual function. Transcriptome analyses show that expression of extracellular matrix-related genes changes dramatically over this period. Co-culture experiments support the existence of a novel regulatory pathway: ECs secrete factors that remodel RPE basement membrane, and integrin receptors sense these changes triggering Rho GTPase signals that modulate RPE tight junctions and enhance RPE barrier function. We anticipate our results will spawn a search for additional roles of choroid ECs in RPE physiology and disease.

Coexistence of anti-glomerular basement membrane antibodies and myeloperoxidase-ANCAs in crescentic glomerulonephritis

NARCIS (Netherlands)

Rutgers, Abraham; Slot, Marjan; van Paassen, Pieter; van Breda Vriesman, Peter; Heeringa, Peter; Tervaert, Jan Willem Cohen

BACKGROUND: In a substantial proportion of patients with crescentic glomerulonephritis (CGN), both anti-glomerular basement membrane (GBM) antibodies and antineutrophil cytoplasmic antibodies (ANCAs) with specificity for myeloperoxidase (MPO-ANCA) are detected. In the present study, we questioned

WY14,643, a PPARα ligand, attenuates expression of anti-glomerular basement membrane disease

Science.gov (United States)

Archer, D C; Frkanec, J T; Cromwell, J; Clopton, P; Cunard, R

2007-01-01

Peroxisome proliferator-activated receptor alpha (PPARα) ligands are medications used to treat hyperlipidaemia and atherosclerosis. Increasing evidence suggests that these agents are immunosuppressive. In the following studies we demonstrate that WY14,643, a PPARα ligand, attenuates expression of anti-glomerular basement membrane disease (AGBMD). C57BL/6 mice were fed 0·05% WY14,643 or control food and immunized with the non-collagenous domain of the α3 chain of Type IV collagen [α3(IV) NC1] in complete Freund's adjuvant (CFA). WY14,643 reduced proteinuria and greatly improved glomerular and tubulo-interstitial lesions. However, the PPARα ligand did not alter the extent of IgG-binding to the GBM. Immunohistochemical studies revealed that the prominent tubulo-interstitial infiltrates in the control-fed mice consisted predominately of F4/80+ macrophages and WY14,643-feeding decreased significantly the number of renal macrophages. The synthetic PPARα ligand also reduced significantly expression of the chemokine, monocyte chemoattractant protein (MCP)-1/CCL2. Sera from mice immunized with AGBMD were also evaluated for antigen-specific IgGs. There was a significant increase in the IgG1 : IgG2c ratio and a decline in the intrarenal and splenocyte interferon (IFN)-γ mRNA expression in the WY14,643-fed mice, suggesting that the PPARα ligand could skew the immune response to a less inflammatory T helper 2-type of response. These studies suggest that PPARα ligands may be a novel treatment for inflammatory renal disease. PMID:17888025

WY14,643, a PPARalpha ligand, attenuates expression of anti-glomerular basement membrane disease.

Science.gov (United States)

Archer, D C; Frkanec, J T; Cromwell, J; Clopton, P; Cunard, R

2007-11-01

Peroxisome proliferator-activated receptor alpha (PPARalpha) ligands are medications used to treat hyperlipidaemia and atherosclerosis. Increasing evidence suggests that these agents are immunosuppressive. In the following studies we demonstrate that WY14,643, a PPARalpha ligand, attenuates expression of anti-glomerular basement membrane disease (AGBMD). C57BL/6 mice were fed 0.05% WY14,643 or control food and immunized with the non-collagenous domain of the alpha3 chain of Type IV collagen [alpha3(IV) NC1] in complete Freund's adjuvant (CFA). WY14,643 reduced proteinuria and greatly improved glomerular and tubulo-interstitial lesions. However, the PPARalpha ligand did not alter the extent of IgG-binding to the GBM. Immunohistochemical studies revealed that the prominent tubulo-interstitial infiltrates in the control-fed mice consisted predominately of F4/80(+) macrophages and WY14,643-feeding decreased significantly the number of renal macrophages. The synthetic PPARalpha ligand also reduced significantly expression of the chemokine, monocyte chemoattractant protein (MCP)-1/CCL2. Sera from mice immunized with AGBMD were also evaluated for antigen-specific IgGs. There was a significant increase in the IgG1 : IgG2c ratio and a decline in the intrarenal and splenocyte interferon (IFN)-gamma mRNA expression in the WY14,643-fed mice, suggesting that the PPARalpha ligand could skew the immune response to a less inflammatory T helper 2-type of response. These studies suggest that PPARalpha ligands may be a novel treatment for inflammatory renal disease.

The extracellular matrix of Gadus morhua muscle contains types III, V, VI and IV collagens in addition to type I

DEFF Research Database (Denmark)

Brüggemann, Dagmar Adeline; Lawson, M.A.

2005-01-01

Confocal microscopy and immuno‐histochemistry were used to examine collagens in the extracellular matrix of cod Gadus morhua swimming muscle. In addition to the well known presence of type I fibrous collagen, types III and VI were also found in the myocommata and the endomysium. The beaded collagen......, type VI, was found in the endomysium and the network forming collagen, type IV, was found in the basement membrane. This is the first report of type V collagen in cod muscle and of types II, IV and VI in the muscle of a teleost....

Expression of type IV collagen in different histological grades of oral squamous cell carcinoma: An immunohistochemical study

Directory of Open Access Journals (Sweden)

Pankaj Agarwal

2013-01-01

Conclusion: The results indicated that there was a direct relationship between the presence of type IV collagen and the differentiation degree of SCC cells and thus that SCC cells loose their capability to form the basement membrane as they become less differentiated.

Normal and tumor-derived myoepithelial cells differ in their ability to interact with luminal breast epithelial cells for polarity and basement membrane deposition

Energy Technology Data Exchange (ETDEWEB)

Gudjonsson, Thorarinn; Ronnov-Jessen, Lone; Villadsen, Rene; Rank, Fritz; Bissell, Mina J.; Petersen, Ole William

2001-10-04

The signals that determine the correct polarity of breast epithelial structures in vivo are not understood. We have shown previously that luminal epithelial cells can be polarized when cultured within a reconstituted basement membrane gel. We reasoned that such cues in vivo may be given by myoepithelial cells. Accordingly, we used an assay where luminal epithelial cells are incorrectly polarized to test this hypothesis. We show that culturing human primary luminal epithelial cells within collagen-I gels leads to formation of structures with no lumina and with reverse polarity as judged by dual stainings for sialomucin, epithelial specific antigen or occludin. No basement membrane is deposited, and {beta}4-integrin staining is negative. Addition of purified human myoepithelial cells isolated from normal glands corrects the inverse polarity, and leads to formation of double-layered acini with central lumina. Among the laminins present in the human breast basement membrane (laminin-1, -5 and -10/11), laminin-1 was unique in its ability to substitute for myoepithelial cells in polarity reversal. Myoepithelial cells were purified also from four different breast cancer sources including a biphasic cell line. Three out of four samples either totally lacked the ability to interact with luminal epithelial cells, or conveyed only correction of polarity in a fraction of acini. This behavior was directly related to the ability of the tumor myoepithelial cells to produce {alpha}-1 chain of laminin. In vivo, breast carcinomas were either negative for laminin-1 (7/12 biopsies) or showed a focal, fragmented deposition of a less intensely stained basement membrane (5/12 biopsies). Dual staining with myoepithelial markers revealed that tumorassociated myoepithelial cells were either negative or weakly positive for expression of laminin-1, establishing a strong correlation between loss of laminin-1 and breast cancer. We conclude that the double-layered breast acinus may be

Nonlinear optical response of the collagen triple helix and second harmonic microscopy of collagen liquid crystals

Science.gov (United States)

Deniset-Besseau, A.; De Sa Peixoto, P.; Duboisset, J.; Loison, C.; Hache, F.; Benichou, E.; Brevet, P.-F.; Mosser, G.; Schanne-Klein, M.-C.

2010-02-01

Collagen is characterized by triple helical domains and plays a central role in the formation of fibrillar and microfibrillar networks, basement membranes, as well as other structures of the connective tissue. Remarkably, fibrillar collagen exhibits efficient Second Harmonic Generation (SHG) and SHG microscopy proved to be a sensitive tool to score fibrotic pathologies. However, the nonlinear optical response of fibrillar collagen is not fully characterized yet and quantitative data are required to further process SHG images. We therefore performed Hyper-Rayleigh Scattering (HRS) experiments and measured a second order hyperpolarisability of 1.25 10-27 esu for rat-tail type I collagen. This value is surprisingly large considering that collagen presents no strong harmonophore in its amino-acid sequence. In order to get insight into the physical origin of this nonlinear process, we performed HRS measurements after denaturation of the collagen triple helix and for a collagen-like short model peptide [(Pro-Pro-Gly)10]3. It showed that the collagen large nonlinear response originates in the tight alignment of a large number of weakly efficient harmonophores, presumably the peptide bonds, resulting in a coherent amplification of the nonlinear signal along the triple helix. To illustrate this mechanism, we successfully recorded SHG images in collagen liquid solutions by achieving liquid crystalline ordering of the collagen triple helices.

Embryoid body attachment to reconstituted basement membrane induces a genetic program of epithelial differentiation via jun N-terminal kinase signaling.

Science.gov (United States)

Ho, Hoang-Yen; Moffat, Ryan C; Patel, Rupal V; Awah, Franklin N; Baloue, Kaitrin; Crowe, David L

2010-09-01

Embryonic stem (ES) cells are derived from early stage mammalian embryos and have broad developmental potential. These cells can be manipulated experimentally to generate cells of multiple tissue types which could be important in treating human diseases. The ability to produce relevant amounts of these differentiated cell populations creates the basis for clinical interventions in tissue regeneration and repair. Understanding how embryonic stem cells differentiate also can reveal important insights into cell biology. A previously reported mouse embryonic stem cell model demonstrated that differentiated epithelial cells migrated out of embryoid bodies attached to reconstituted basement membrane. We used genomic technology to profile ES cell populations in order to understand the molecular mechanisms leading to epithelial differentiation. Cells with characteristics of cultured epithelium migrated from embryoid bodies attached to reconstituted basement membrane. However, cells that comprised embryoid bodies also rapidly lost ES cell-specific gene expression and expressed proteins characteristic of stratified epithelia within hours of attachment to basement membrane. Gene expression profiling of sorted cell populations revealed upregulation of the BMP/TGFbeta signaling pathway, which was not sufficient for epithelial differentiation in the absence of basement membrane attachment. Activation of c-jun N-terminal kinase 1 (JNK1) and increased expression of Jun family transcription factors was observed during epithelial differentiation of ES cells. Inhibition of JNK signaling completely blocked epithelial differentiation in this model, revealing a key mechanism by which ES cells adopt epithelial characteristics via basement membrane attachment. Copyright (c) 2010 Elsevier B.V. All rights reserved.

In Vitro and In Vivo Study of a Novel Porcine Collagen Membrane for Guided Bone Regeneration

Directory of Open Access Journals (Sweden)

Eisner Salamanca

2016-11-01

Full Text Available For years, in order to improve bone regeneration and prevent the need of a second stage surgery to remove non-resorbable membranes, biological absorbable membranes have gradually been developed and applied in guided tissue regeneration (GTR. The present study’s main objective was to achieve space maintenance and bone regeneration using a new freeze-dried developed porcine collagen membrane, and compare it with an already commercial collagen membrane, when both were used with a bovine xenograft in prepared alveolar ridge bone defects. Prior to surgery, the membrane’s vitality analysis showed statistically significant higher cell proliferation in the test membrane over the commercial one. In six beagle dogs, commercial bone xenograft was packed in lateral ridge bone defects prepared in the left and right side and then covered with test porcine collagen membrane or commercial collagen membrane. Alveolar height changes were measured. Histomorphometric results, in vitro and in vivo properties indicated that the new porcine collagen membrane is biocompatible, enhances bone xenograft osteoconduction, and reduces the alveolar ridge height reabsorption rate.

Agrin is a major heparan sulfate proteoglycan in the human glomerular basement membrane

NARCIS (Netherlands)

Groffen, Alexander J.; Ruegg, Markus A.; Dijkman, Henri; Van De Velden, Thea J.; Buskens, Carin A.; Van Den Born, Jacob; Assmann, Karel J.; Monnens, Leo A.; Veerkamp, Jacques H.; Van Den Heuvel, Lambert P.

Agrin is a heparan sulfate proteoglycan (HSPG) that is highly concentrated in the synaptic basal lamina at the neuromuscular junction (NMJ). Agrin-like immunoreactivity is also detected outside the NMJ. Here we show that agrin is a major HSPG component of the human glomerular basement membrane

Basement membrane-specific chondroitin sulfate proteoglycan is abnormally associated with the glomerular capillary basement membrane of diabetic rats

DEFF Research Database (Denmark)

McCarthy, K J; Abrahamson, D R; Bynum, K R

1994-01-01

exception being the normal glomerular capillary basement membrane (GBM), where it is absent. In the present study of mature kidneys we examined the distribution of BM-CSPG in streptozocin-induced diabetes mellitus in rats. We found BM-CSPG atypically associated with the GBM of diabetic animals as early as 1...... month after induction of diabetes mellitus. Immunoelectron microscopy (IEM) of affected capillary loops showed BM-CSPG present in the subendothelial matrix in areas of GBM thickening and absent in areas where the GBM appears to be of normal thickness. Moreover, the association of BM-CSPG with regions...... of the pericapillary GBM affects the morphology of the capillary endothelial cells within these areas, directly displacing the cell body from the GBM proper and causing loss of fenestrae. These new data on BM-CSPG distribution reflect abnormal glomerular extracellular matrix protein biosynthesis/turnover in diabetes...

Glucuronylated core 1 glycans are required for precise localization of neuromuscular junctions and normal formation of basement membranes on Drosophila muscles.

Science.gov (United States)

Itoh, Kazuyoshi; Akimoto, Yoshihiro; Kondo, Shu; Ichimiya, Tomomi; Aoki, Kazuhiro; Tiemeyer, Michael; Nishihara, Shoko

2018-04-15

T antigen (Galβ1-3GalNAcα1-Ser/Thr) is an evolutionary-conserved mucin-type core 1 glycan structure in animals synthesized by core 1 β1,3-galactosyltransferase 1 (C1GalT1). Previous studies showed that T antigen produced by Drosophila C1GalT1 (dC1GalT1) was expressed in various tissues and dC1GalT1 loss in larvae led to various defects, including decreased number of circulating hemocytes, hyper-differentiation of hematopoietic stem cells in lymph glands, malformation of the central nervous system, mislocalization of neuromuscular junction (NMJ) boutons, and ultrastructural abnormalities in NMJs and muscle cells. Although glucuronylated T antigen (GlcAβ1-3Galβ1-3GalNAcα1-Ser/Thr) has been identified in Drosophila, the physiological function of this structure has not yet been clarified. In this study, for the first time, we unraveled biological roles of glucuronylated T antigen. Our data show that in Drosophila, glucuronylation of T antigen is predominantly carried out by Drosophila β1,3-glucuronyltransferase-P (dGlcAT-P). We created dGlcAT-P null mutants and found that mutant larvae showed lower expression of glucuronylated T antigen on the muscles and at NMJs. Furthermore, mislocalization of NMJ boutons and a partial loss of the basement membrane components collagen IV (Col IV) and nidogen (Ndg) at the muscle 6/7 boundary were observed. Those two phenotypes were correlated and identical to previously described phenotypes in dC1GalT1 mutant larvae. In addition, dGlcAT-P null mutants exhibited fewer NMJ branches on muscles 6/7. Moreover, ultrastructural analysis revealed that basement membranes that lacked Col IV and Ndg were significantly deformed. We also found that the loss of dGlcAT-P expression caused ultrastructural defects in NMJ boutons. Finally, we showed a genetic interaction between dGlcAT-P and dC1GalT1. Therefore, these results demonstrate that glucuronylated core 1 glycans synthesized by dGlcAT-P are key modulators of NMJ bouton localization

Expression of collagen in ovular membranes of pregnant smokers and non-smokers: a pilot study.

Science.gov (United States)

Negrini, Romulo; Araujo Júnior, Edward; Piato, Sebastião; Chade, Milca Cezar; Rios, Adriana Ribeiro Santos; Silva, Maria Antonieta Galvão; Aldrighi, José Mendes

2015-09-01

Our study compared the amount of total collagen and type I collagen in ovular membranes of pregnant smokers and non-smokers. The study group consisted of 14 pregnant smokers at 24-36 weeks of gestation; 39 pregnant non-smokers between 24-36 weeks of gestation comprised the control group. The expressions of total collagen and type I collagen were analyzed using two histological sections of the fetal membranes. The assessment of total collagen was performed using the Picro-Cirius red stain, and type I collagen expression was determined by means of immunohistochemistry The Mann-Whitney test was applied to verify possible differences between the groups. The average area covered by total collagen was lower in smokers (20630.45 microm2) as compared to non-smokers (24058.61 microm2), although the difference was not statistically significant (p = 0.454). Comparison involving collagen type I deemed similar results (20001.33 microm2 vs. 25328.29 microm2, p = 0.158). The amount of total collagen and type I collagen was lower in ovular membranes of pregnant smokers as compared to non-smokers, although the difference was not statistically significant.

Antibody response against the glomerular basement membrane protein agrin in patients with transplant glomerulopathy.

NARCIS (Netherlands)

Joosten, S.A.; Sijpkens, Y.W.; Ham, V. van; Trouw, L.A.; Vlag, J. van der; Heuvel, L.P.W.J. van den; Kooten, C. van; Paul, L.C.

2005-01-01

Chronic allograft nephropathy (CAN) of renal allografts is still the most important cause of graft loss. A subset of these patients have transplant glomerulopathy (TGP), characterized by glomerular basement membrane (GBM) duplications, but of unknown etiology. Recently, a role for the immune system

ULTRASTRUCTURAL-CHANGES OF THE BASEMENT-MEMBRANE ZONE IN BENIGN LESIONS OF THE VOCAL FOLDS

NARCIS (Netherlands)

DIKKERS, FG; HULSTAERT, CE; OOSTERBAAN, JA; CERVERAPAZ, FJ

The basement membrane zone (BMZ) of the epithelium of the vocal folds was investigated electron microscopically in 10 patients suffering from various benign lesions and in 3 controls. Various defects were observed: a thickening by deposition of electron dense material, a loss of normal architecture,

Targeted Expression of Stromelysin-1 in Mammary Gland Provides Evidence for a Role of Proteinases in Branching Morphogenesis and the Requirement for an Intact Basement Membrane for Tissue-specific Gene Expression

Energy Technology Data Exchange (ETDEWEB)

Sympson, Carolyn J; Talhouk, Rabih S; Alexander, Caroline M; Chin, Jennie R; Cliff, Shirley M; Bissell, Mina J; Werb, Zena

1994-05-01

The extracellular matrix (ECM) is an important regulator of the differentiated phenotype of mammary epithelial cells in culture. Despite the fact that ECM-degrading enzymes have been implicated in morphogenesis and tissue remodeling, there is little evidence for a direct role for such regulation in vivo. We generated transgenic mice that express autoactivated isoforms of the matrix metalloproteinase stromelysin-1, under the control of the whey acidic protein gene promoter, to examine the effect of inappropriate expression of this enzyme. Stromelysin-1 is implicated as the primary player in the loss of basement membrane and loss of function in the mammary gland during involution. The transgene was expressed at low levels in mammary glands of virgin female mice, leading to an unexpected phenotype: The primary ducts had supernumerary branches and showed precocious development of alveoli that expressed beta-casein at levels similar to that of an early- to mid-pregnant gland. Lactating glands showed high levels of transgene expression, with accumulation at the basement membrane, and a decrease in laminin and collagen IV, resulting in a loss of basement membrane integrity; this was accompanied by a dramatic alteration of alveolar morphology, with decreased size and shrunken lumina containing little beta-casein. During pregnancy, expression of endogenous whey acidic protein and beta-casein was reduced in transgenic glands, confirming the observed dependence of milk protein transcription of ECM in mammary epithelial cells in culture. These data provide direct evidence that stromelysin-1 activity can be morphogenic for mammary epithelial cells, inducing hyperproliferation and differentiation in virgin animals, and that its lytic activity can, indeed, disrupt membrane integrity and reduce mammary-specific function. We conclude that the balance of ECM-degrading enzymes with their inhibitors, and the associated regulation of ECM structure, is crucial for tissue-specific gene

De novo deposition of laminin-positive basement membrane in vitro by normal hepatocytes and during hepatocarcinogenesis

DEFF Research Database (Denmark)

Albrechtsen, R; Wewer, U M; Thorgeirsson, S S

1988-01-01

De novo formation of laminin-positive basement membranes was found to be a distinct morphologic feature of diethylnitrosamine/phenobarbital-induced hepatocellular carcinomas of the rat. The first appearance of extracellularly located laminin occurred in the preneoplastic liver lesions...... (corresponding to neoplastic nodules), and this feature became successively more prominent during the course of hepatocellular carcinoma development. Most groups of tumor cells were surrounded by laminin-positive basement membrane material. The laminin-positive material was also deposited along the sinusoids......, a location where no laminin was seen in normal rat liver. The amount of extractable laminin from hepatocellular carcinomas was significantly higher (approximately 100 ng per mg tissue) than that of normal liver tissue (less than 20 ng per mg). In vitro experiments demonstrated that normal and preneoplastic...

Immunohistochmical Study of Glomerular Mesengial Collagen IV Expression in Diabetic Balb/c Mice

Directory of Open Access Journals (Sweden)

M. Jalali

2006-04-01

Full Text Available Introduction & Objective: Extra-cellular matrix and basement membrane play important roles in many developmental phenamenon during development and after birth. Among the components of the basement membrane, collagen fibers specially type IV, are the most important part of this area. As kidney is one of the target organs in diabetes mellitus and diabetic nephropathy is a major cause of end stage-renal disease and result an increase in morbidity and mortality of effected individuals, therefore early diagnosis leads to better treatment. The aim of this investigation was to study the primary diagnostic parameters by special regards to collagen IV fibers.Materials & Methods: In this study, 24 male balb/c mice were divided into experimental and control groups. In experimental group, the beta cells of Langerhance were chemically destroyed by an injection of 160 mg/kg alloxan and subdivided in experimental groups 1 and 2. Controls were kept untreated. Experimental group 1and 2 were sacrified 8 and 16 weeks after treatment with alloxan respectively. The same procedure was performed for control group. Immunohistochmical studies were carried out using monocolonal antibody against collagen type IV in Glomeruli. In addition, using morphometrical and stereological methods the volume of the glumerles was compared in all groups.Results: Our finding showed that in experimental groups with special regards in 16 weeks diabetic mice, the rate of collagen type IV in basement membrane around the parietal layer of Bowman capsule, mesangial cells and endothelium of capillary in glomerules increased significantly compared to controls and experimental group 1 (p<0.05, while there was not a significant difference among experimental group 2 and controls. Our data also revealed that the number of mesangial cells as well as glomerular volume increased significantly in experimental 2 (4.635±0.289×106µm3 compared to experimental 1 (3.504±0.189×106µm3 and controls (3.422�

Electron microscopic study of the myelinated nerve fibres and the perineurial cell basement membrane in the diabetic human peripheral nerves

International Nuclear Information System (INIS)

ElBarrany, Wagih G.; Hamdy, Raid M.; AlHayani, Abdulmonem A.; Jalalah, Sawsan M.

2009-01-01

To study the quantitative and ultrastructural changes in myelinated nerve fibers and the basement membranes of the perineurial cells in diabetic nerves. The study was performed at the Department of Anatomy, Faculty of Medicine, King Abdul-Aziz University, Jeddah, Saudi Arabia from 2003 to 2005. Human sural nerves were obtained from 15 lower limbs and 5 diabetic nerve biopsies. The total mean and density of myelinated nerve fibers per fascicle were calculated, with density of microtubules and mitochondria in the axoplasm. The number of the perineurial cell basement membrane layers was counted, and thickness of the basement membrane was measured. Among the 15 diabetic and 5 normal human sural nerves, the average diameters, number and surface area of myelinated nerve fibers and axonal microtubules density were found to be less in diabetic nerves. Mitochondrial density was higher in diabetic axons. Thickness of the perineurial cell basement membrane had a greater mean, but the number of perineurial cell layers was less than that of the diabetic group. The inner cellular layer of the perineurium of the diabetic nerves contained large vacuoles containing electron-dense degenerated myelin. A few specimens showed degenerated myelinated nerve fibers, while others showed recovering ones. Retracted axoplasms were encountered with albumin extravasation. Diabetes caused an increase in perineurial permeability. The diabetic sural nerve showed marked decrease in the myelinated nerve fibres, increase degenerated mitochondria, and decreased microtubules. (author)

Effect of diabetes on in vivo metabolism of [35S]-labeled glomerular basement membrane

International Nuclear Information System (INIS)

Cohen, M.P.; Surma, M.L.

1984-01-01

Glomerular basement membrane (GBM) was labeled in vivo by the injection of tracer amounts of [ 35 S]-sulfate into normal and streptozotocin-diabetic rats. The biosynthesis and turnover of sulfated glycosaminoglycans in the GBM was determined from the specific activity of [ 35 S] after pronase digestion of basement membranes purified from glomeruli isolated 1-7 days after injection. Peak radiolabeling of both normal and diabetic GBM occurred 24 h after injection and, when corrected for differences in serum sulfate specific activities, was less in diabetic than in normal samples. The specific activity of GBM sulfate, expressed as cpm/microgram uronic acid, progressively diminished over the ensuing period of study in both normal and diabetic samples. The rate of decrease in specific activity of [ 35 S]-labeled GBM was not significantly different in diabetic preparations compared with that in normal controls. The findings are compatible with diminished sulfation and/or production but normal turnover of glycosaminoglycans in the renal GBM in experimental diabetes

Physiochemical properties and resorption progress of porcine skin-derived collagen membranes: In vitro and in vivo analysis.

Science.gov (United States)

An, Yin-Zhe; Kim, You-Kyoung; Lim, Su-Min; Heo, Yeong-Ku; Kwon, Mi-Kyung; Cha, Jae-Kook; Lee, Jung-Seok; Jung, Ui-Won; Choi, Seong-Ho

2018-03-30

The aim of the present study was to evaluate the physiochemical properties and resorption progress of two cross-linked, porcine skin-derived collagen membranes and compare their features with those of a membrane without cross-linking (Bio-Gide ® [BG], Geistlich Biomaterials, Wolhusen, Switzerland). Three porcine skin-derived collagen membranes, dehydrothermally (DHT) cross-linked (experimental), DHT and 1-ethyl-3(3-dimethylaminopropyl)-carbodiimide (DHT/EDC) cross-linked (experimental) and BG were investigated for their morphology, enzyme resistance, and tensile strength in vitro and biodegradation in vivo. DHT and DHT/EDC membranes exhibited irregular, interconnected macro- and micropores that formed a 3D mesh, whereas BG exhibited individual collagen fibrils interlaced to form coarse collagen strands. In enzyme resistance and tensile strength tests, DHT and DHT/EDC membranes demonstrated good resistance and mechanical properties compared with BG. In vivo, all three membranes were well integrated into the surrounding connective tissue. Thus, the DHT membrane exhibited its potential as a barrier membrane for guided bone and tissue regeneration.

Experimental autoimmune glomerulonephritis induced by anti-glomerular basement membrane antibody. II. Effects of injecting heterologous, homologous, or autologous glomerular basement membranes and complete Freund's adjuvant into sheep.

OpenAIRE

Steblay, R. W.; Rudofsky, U. H.

1983-01-01

The effects of injecting human, rabbit, rat, or single-kidney homologous glomerular basement membrane (GBM) or autologous GBM, each in complete Freund's adjuvant (CFA), into 15- to 18-month-old sheep are compared. All sheep receiving heterologous GBM and 3 of 6 sheep receiving homologous GBM had anti-GBM nephritis, but such sheep did not bind autoantibodies or have Goodpasturelike lesions in their lungs. Sheep given injections of human GBM had autoantibodies to antigenic determinants shared b...

Improving the mechanical properties of collagen-based membranes using silk fibroin for corneal tissue engineering.

Science.gov (United States)

Long, Kai; Liu, Yang; Li, Weichang; Wang, Lin; Liu, Sa; Wang, Yingjun; Wang, Zhichong; Ren, Li

2015-03-01

Although collagen with outstanding biocompatibility has promising application in corneal tissue engineering, the mechanical properties of collagen-based scaffolds, especially suture retention strength, must be further improved to satisfy the requirements of clinical applications. This article describes a toughness reinforced collagen-based membrane using silk fibroin. The collagen-silk fibroin membranes based on collagen [silk fibroin (w/w) ratios of 100:5, 100:10, and 100:20] were prepared by using silk fibroin and cross-linking by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide. These membranes were analyzed by scanning electron microscopy and their optical property, and NaCl and tryptophan diffusivity had been tested. The water content was found to be dependent on the content of silk fibroin, and CS10 membrane (loading 10 wt % of silk fibroin) performed the optimal mechanical properties. Also the suture experiments have proved CS10 has high suture retention strength, which can be sutured in rabbit eyes integrally. Moreover, the composite membrane proved good biocompatibility for the proliferation of human corneal epithelial cells in vitro. Lamellar keratoplasty shows that CS10 membrane promoted complete epithelialization in 35 ± 5 days, and their transparency is restored quickly in the first month. Corneal rejection reaction, neovascularization, and keratoconus are not observed. The composite films show potential for use in the field of corneal tissue engineering. © 2014 Wiley Periodicals, Inc.

Immunochemical and ultrastructural assessment of the nature of the pericellular basement membrane of human decidual cells

DEFF Research Database (Denmark)

Wewer, U M; Faber, M; Liotta, L A

1985-01-01

Human decidual cells of early and late pregnancy were studied immunochemically and ultrastructurally with respect to the presence and nature of pericellular basement membrane material. The most prominent cell type in decidual tissue of both early and late pregnancy were large, mature epithelioid......-linked immunosorbent assay. Biosynthesis of laminin was shown by [35S]methionine labeling of short term organ cultures of decidual tissue followed by immunoprecipation, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and fluorography. The laminin chains migrated with the apparent molecular weights of 300...... and 200 kilodaltons under reducing conditions. Two other separate populations of cells were apparent in the decidual tissue of early pregnancy. A smaller group of rounded intermediate sized (15 to 25 micron) decidual cells had focal deposits basement membrane immunoreactive material scattered at the cell...

Aluminum-containing dense deposits of the glomerular basement membrane: identification by energy dispersive X-ray analysis

International Nuclear Information System (INIS)

Smith, D.M. Jr.; Pitcock, J.A.; Murphy, W.M.

1982-01-01

Heavy metals, including gold, mercury, lead, bismuth, and cadmium, have the potential to cause renal disease. With the development of X-ray microanalysis, these heavy metals can now be identified in tissue deposits. This report describes a case of renal failure, probably related to dysproteinemia, in which granular, electron-opaque dense deposits were present in the glomerular basement membranes. Energy dispersive X-ray analysis demonstrated that these dense deposits contained aluminum. An analysis of this patient's history in relation to the current knowledge of aluminum metabolism suggests that the aluminum deposition occurred secondary to previous glomerular injury. This case emphasizes the need to utilize heavy metal identification technology whenever granular, electron-opaque dense deposits are identified and represents, to our knowledge, the first study to document aluminum deposits within the glomerular basement membrane of humans

Retinoid inhibition of in vitro invasion of human amnion basement membrane by human tumor cells

International Nuclear Information System (INIS)

Fazely, F.; Ledinko, N.; Smith, D.J.

1986-01-01

The biological activity of retinoids was assayed in an in vitro quantitative assay of human tumor cell invasion using human amnion basement membrane (BM). The effects measured were the inhibition of tumor cell migration through the BM and tumor cell degradative enzyme activity on 14 C-proline labeled collagenous and noncollagenous components of the BM. The human lung carcinoma A549 or the human Ewing's sarcoma TC-106 cell lines treated with retinoids for two days were incubated on the BM in the absence of retinoids. A dose-dependent inhibition of cell invasion was produced by retinoids. Among the retinoids tested, the most powerful was retinol acetate which inhibited invasion by 50% of A549 cells at a concentration of 0.009 μg/mL, and of TC-106 cells at 0.07 μg/mL. Retinol acetate inhibited A549 and TC-106 cell growth by approximately 50% at levels over 100-fold higher than those needed for antiinvasive activity. Retinol acetate was about 20 times more potent than retinoic acid and 30 times more potent than retinol palmitate. The model system will be useful for investigating antiinvasive activity of other retinoids as well as other compounds

In vitro sealing of iatrogenic fetal membrane defects by a collagen plug imbued with fibrinogen and plasma.

Science.gov (United States)

Engels, A C; Hoylaerts, M F; Endo, M; Loyen, S; Verbist, G; Manodoro, S; DeKoninck, P; Richter, J; Deprest, J A

2013-02-01

We aimed to demonstrate local thrombin generation by fetal membranes, as well as its ability to generate fibrin from fibrinogen concentrate. Furthermore, we aimed to investigate the efficacy of collagen plugs, soaked with plasma and fibrinogen, to seal iatrogenic fetal membrane defects. Thrombin generation by homogenized fetal membranes was measured by calibrated automated thrombography. To identify the coagulation caused by an iatrogenic membrane defect, we analyzed fibrin formation by optical densitometry, upon various concentrations of fibrinogen. The ability of a collagen plug soaked with fibrinogen and plasma was tested in an ex vivo model for its ability to seal an iatrogenic fetal membrane defect. Fetal membrane homogenates potently induced thrombin generation in amniotic fluid and diluted plasma. Upon the addition of fibrinogen concentrate, potent fibrin formation was triggered. Measured by densiometry, fibrin formation was optimal at 1250 µg/mL fibrinogen in combination with 4% plasma. A collagen plug soaked with fibrinogen and plasma sealed an iatrogenic membrane defect about 35% better than collagen plugs without these additives (P = 0.037). These in vitro experiments suggest that the addition of fibrinogen and plasma may enhance the sealing efficacy of collagen plugs in closing iatrogenic fetal membrane defects. © 2013 John Wiley & Sons, Ltd.

Influence of a reconstituted basement membrane and its components on casein gene expression and secretion in mouse mammary epithelial cells

International Nuclear Information System (INIS)

Li, M.L.; Aggeler, J.; Farson, D.A.; Hatier, C.; Hassell, J.; Bissell, M.J.

1987-01-01

When primary mouse mammary epithelial cells are cultured on plastic, they rapidly lose their ability to synthesize and secrete most milk proteins even in the presence of lactogenic hormones, whereas cells cultured on release type I collagen gels show greatly enhanced mRNA levels and secretion rates of β-casein and of some other milk proteins. The authors show here that culture on a reconstituted basement membrane from Engelbreth-Holm-Swarm tumor (EHS) allows > 90% of cells to produce high levels of β-casein. By comparison, 30-40% of cells on released type 1 gels and only 2-10% of cells on plastic express β-casein after 6 days in culture. Because only 40% of cells from late pregnant gland produced β-casein before culture, the EHS matrix can both induce and maintain an increased level of casein gene expression. Individual basal lamina components were also evaluated. Type IV collagen and fibronectin had little effect on morphology and β-casein mRNA levels. In contrast, both laminin and heparan sulfate proteoglycan increased β-casein mRNA levels. Profound morphological differences were evident between cells cultured on plastic and on EHS matrix, the latter cells forming ducts, ductules, and lumina and resembling secretory alveoli. These results emphasize the vital role of the extracellular matrix in receiving and integrating structural and functional signals that can direct specific gene expression in differentiated tissues

Dynamic interplay between the collagen scaffold and tumor evolution

DEFF Research Database (Denmark)

Egeblad, Mikala; Rasch, Morten G; Weaver, Valerie M

2010-01-01

The extracellular matrix (ECM) is a key regulator of cell and tissue function. Traditionally, the ECM has been thought of primarily as a physical scaffold that binds cells and tissues together. However, the ECM also elicits biochemical and biophysical signaling. Controlled proteolysis...... and remodeling of the ECM network regulate tissue tension, generate pathways for migration, and release ECM protein fragments to direct normal developmental processes such as branching morphogenesis. Collagens are major components of the ECM of which basement membrane type IV and interstitial matrix type I...

A role for collagen IV in cardiovascular disease?

DEFF Research Database (Denmark)

Steffensen, Lasse Bach; Rasmussen, Lars M

2018-01-01

Over the past decade, studies have repeatedly found single nucleotide polymorphisms located in the COL4A1 and COL4A2 genes to be associated with cardiovascular disease (CVD), and the 13q34 locus harboring these genes is one of approximately 160 genome-wide significant risk loci for coronary artery...... disease. COL4A1 and COL4A2 encode the ⍺1- and ⍺2-chains of collagen IV, a major component of basement membranes in various tissues including arteries. In spite of the growing body of evidence indicating a role for collagen IV in CVD, remarkably few studies aim at directly investigating such a role....... The purpose of this review is to summarize the clinical reports linking 13q34 to coronary artery disease, atherosclerosis and artery stiffening and to assemble the scattered pieces of evidence from experimental studies based on vascular cells and -tissue collectively supporting a role for collagen IV...

Circulating Vascular Basement Membrane Fragments are Associated with the Diameter of the Abdominal Aorta and Their Expression Pattern is Altered in AAA Tissue.

Science.gov (United States)

Holsti, Mari; Wanhainen, Anders; Lundin, Christina; Björck, Martin; Tegler, Gustaf; Svensson, Johan; Sund, Malin

2018-04-12

Abdominal aortic aneurysm (AAA) is characterised by enhanced proteolytic activity, and extracellular matrix (ECM) remodelling in the vascular wall. Type IV and XVIII collagen/endostatin are structural proteins in vascular basement membrane (VBM), a specialised ECM structure. Here the association between plasma levels of these collagens with the aortic diameter and expansion rate is studied, and their expression in aortic tissue characterised. This was a retrospective population based cohort study. Type IV and XVIII collagen/endostatin were analysed in plasma by ELISA assay in 615 men, divided into three groups based on the aortic diameter: 1) normal aorta ≤ 25 mm, 2) sub-aneurysmal aorta (SAA) 26-29 mm, and 3) AAA ≥ 30 mm. Follow up data were available for 159 men. The association between collagen levels and aortic diameter at baseline, and with the expansion rate at follow up were analysed in ordinal logistic regression and linear regression models, controlling for common confounding factors. Tissue expression of the collagens was analysed in normal aorta (n = 6) and AAA (n = 6) by immunofluorescence. Plasma levels of type XVIII collagen/endostatin (136 ng/mL [SD 29] in individuals with a normal aorta diameter, 154 ng/ml [SD 45] in SAA, and 162 ng/ml [SD 46] in AAA; p = .001) and type IV collagen (105 ng/mL [SD 42] normal aorta, 124 ng/ml [SD 46] SAA, and 127 ng/ml [SD 47] AAA; p = .037) were associated with a larger aortic diameter. A significant association was found between the baseline levels of type XVIII/endostatin and the aortic expansion rate (p = .035), but in the multivariable model, only the initial aortic diameter remained significantly associated with expansion (p = .005). Altered expression patterns of both collagens were observed in AAA tissue. Plasma levels of circulating type IV and XVIII collagen/endostatin increase with AAA diameter. The expression pattern of VBM proteins is altered in the aneurysm wall. Copyright �

Perlecan (basement membrane heparan sulfate proteoglycan and its role in oral malignancies: An overview

Directory of Open Access Journals (Sweden)

Mithilesh Mishra

2011-01-01

Full Text Available Perlecan means pearl-like structures. Perlecan is a large proteoglycan (400-500 kDa present in virtually all vascularized tissues with a distribution that is primarily confined to basement membranes including those of oral mucosa. It is a basement membrane-type heparan sulfate proteoglycan. Perlecan is synthesized by basal cells and fibroblasts adjacent to the basal lamina . Perlecan is also synthesized by vascular endothelial and smooth muscle cells present in the extracellular matrix. It has been demonstrated in recent years that perlecan is distributed in the stromal space of various pathophysiological conditions. The complex pleiotropy of perlecan suggests that this gene product is involved in several developmental processes, at both early and late stages of embryogenesis, as well as in cancer and diabetes. In the oral cavity, perlecan expression is reported to basal cells in normal mucosa and its expression increases in precancer and cancerous conditions. It is also expressed in various odontogenic tumors such as ameloblastoma, keratocyst odontogenic tumor, and also salivary gland tumors such as adenoid cystic carcinoma, mucoepidermoid carcinoma, etc.

Measurement of the quadratic hyperpolarizability of the collagen triple helix and application to second harmonic imaging of natural and biomimetic collagenous tissues

Science.gov (United States)

Deniset-Besseau, A.; Strupler, M.; Duboisset, J.; De Sa Peixoto, P.; Benichou, E.; Fligny, C.; Tharaux, P.-L.; Mosser, G.; Brevet, P.-F.; Schanne-Klein, M.-C.

2009-09-01

Collagen is a major protein of the extracellular matrix that is characterized by triple helical domains. It plays a central role in the formation of fibrillar and microfibrillar networks, basement membranes, as well as other structures of the connective tissue. Remarkably, fibrillar collagen exhibits efficient Second Harmonic Generation (SHG) so that SHG microscopy proved to be a sensitive tool to probe the three-dimensional architecture of fibrillar collagen and to assess the progression of fibrotic pathologies. We obtained sensitive and reproducible measurements of the fibrosis extent, but we needed quantitative data at the molecular level to further process SHG images. We therefore performed Hyper- Rayleigh Scattering (HRS) experiments and measured a second order hyperpolarisability of 1.25 10-27 esu for rat-tail type I collagen. This value is surprisingly large considering that collagen presents no strong harmonophore in its aminoacid sequence. In order to get insight into the physical origin of this nonlinear process, we performed HRS measurements after denaturation of the collagen triple helix and for a collagen-like short model peptide [(Pro-Pro- Gly)10]3. It showed that the collagen large nonlinear response originates in the tight alignment of a large number of weakly efficient harmonophores, presumably the peptide bonds, resulting in a coherent amplification of the nonlinear signal along the triple helix. To illustrate this mechanism, we successfully recorded SHG images in collagenous biomimetic matrices.

Elevated expression of type VII collagen in the skin of patients with systemic sclerosis. Regulation by transforming growth factor-beta.

OpenAIRE

Rudnicka, L; Varga, J; Christiano, A M; Iozzo, R V; Jimenez, S A; Uitto, J

1994-01-01

A hallmark of systemic sclerosis (SSc) is the development of tissue fibrosis. Excessive production of several connective tissue components normally present in the dermis, including type I, III, V, and VI collagens as well as fibronectin and proteoglycans, is a consistent finding in the skin of SSc patients. Type VII collagen is a major constituent of anchoring fibrils, present in the skin at the dermal-epidermal basement membrane zone. TGF-beta has been shown to upregulate the expression of t...

Graphene oxide enrichment of collagen membranes improves DPSCs differentiation and controls inflammation occurrence.

Science.gov (United States)

Radunovic, Milena; De Colli, Marianna; De Marco, Patrizia; Di Nisio, Chiara; Fontana, Antonella; Piattelli, Adriano; Cataldi, Amelia; Zara, Susi

2017-08-01

Collagen membranes are used in oral surgery for bone defects treatment acting as a barrier that does not allow the invasion of soft tissue into the growing bone. To improve biocompatibility collagen membranes were coated with graphene oxide (GO), a graphene derivative. The aim of this study was to investigate the biocompatibility of GO coated collagen membranes on human dental pulp stem cells (DPSCs) focusing on biomaterial cytotoxicity, ability to promote DPSCs differentiation process and to control inflammation event induction. DPSCs were cultured on uncoated membranes and on both 2 and 10 μg mL -1 GO coated membranes up to 28 days. Alamar blue and LDH cytotocicity assay, PGE2 ELISA assay, real time RT-PCR for RUNX2, BMP2, SP7, TNFα and COX2 genes expression were performed. Proliferation is higher on GO coated membranes at days 14 and 28. LDH assay evidences no cytotoxicity. BMP2 and RUNX2 expression is higher on coated membranes, BMP2 at early and RUNX2 and SP7 at late experimental times. PGE2 levels are lower on GO coated membranes at days 14 and 28, both TNFα and COX2 expression is significantly decreased when GO is applied. GO coated membranes are not toxic for DPSCs, induce a faster DPSCs differentiation into odontoblasts/osteoblasts and may represent good alternative to conventional membranes thus ensuring more efficient bone formation and improving the clinical performance. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2312-2320, 2017. © 2017 Wiley Periodicals, Inc.

cDNA cloning of the basement membrane chondroitin sulfate proteoglycan core protein, bamacan: a five domain structure including coiled-coil motifs

DEFF Research Database (Denmark)

Wu, R R; Couchman, J R

1997-01-01

Basement membranes contain several proteoglycans, and those bearing heparan sulfate glycosaminoglycans such as perlecan and agrin usually predominate. Most mammalian basement membranes also contain chondroitin sulfate, and a core protein, bamacan, has been partially characterized. We have now....... The protein sequence has low overall homology, apart from very small NH2- and COOH-terminal motifs. At the junctions between the distal globular domains and the coiled-coil regions lie glycosylation sites, with up to three N-linked oligosaccharides and probably three chondroitin chains. Three other Ser...

viking: identification and characterization of a second type IV collagen in Drosophila.

Science.gov (United States)

Yasothornsrikul, S; Davis, W J; Cramer, G; Kimbrell, D A; Dearolf, C R

1997-10-01

We have taken an enhancer trap approach to identify genes that are expressed in hematopoietic cells and tissues of Drosophila. We conducted a molecular analysis of two P-element insertion strains that have reporter gene expression in embryonic hemocytes, strain 197 and vikingICO. This analysis has determined that viking encodes a collagen type IV gene, alpha2(IV). The viking locus is located adjacent to the previously described DCg1, which encodes collagen alpha1(IV), and in the opposite orientation. The alpha2(IV) and alpha1(IV) collagens are structurally very similar to one another, and to vertebrate type IV collagens. In early development, viking and DCg1 are transcribed in the same tissue-specific pattern, primarily in the hemocytes and fat body cells. Our results suggest that both the alpha1 and alpha2 collagen IV chains may contribute to basement membranes in Drosophila. This work also provides the foundation for a more complete genetic dissection of collagen type IV molecules and their developmental function in Drosophila.

A novel urinary biomarker of type VI collagen formation and endotrophin is associated with loss of kidney function in patients with diabetic nephropathy

DEFF Research Database (Denmark)

Genovese, Federica; Rasmussen, Daniel; Nielsen, Signe Holm

2017-01-01

-stage renal disease. Fibrosis is characterized by a dysregulated remodeling of the extracellular matrix (ECM). Collagen type VI (COL VI) is a crucial ECM molecule for the control of tissue organization. It is present at the interface of the glomerular basement membrane and interstitial matrix and its levels...

Deposition of nucleosomal antigens (histones and DNA) in the epidermal basement membrane in human lupus nephritis.

NARCIS (Netherlands)

Grootscholten, C.; Bruggen, M.C.J. van; Pijl, J.W. van der; Jong, E.M.G.J. de; Ligtenberg, G.; Derksen, R.H.W.M.; Berden, J.H.M.

2003-01-01

OBJECTIVE: Antinuclear autoantibodies complexed to nucleosomes can bind to heparan sulfate (HS) in the glomerular basement membrane. This binding is due to the binding of the positively charged histones to the strongly anionic HS. Nucleosomes and histones have been identified in glomerular deposits

The chest X-ray in antiglomerular basement membrane antibody disease (Goodpasture's syndrome)

International Nuclear Information System (INIS)

Bowley, N.B.; Steiner, R.E.; Chin, W.S.

1979-01-01

The chest radiographs of 25 patients with proven antiglomerular basement membrane antibody disease (Goodpasture's syndrome) were analysed. All except two of the patients had pulmonary haemorrhage at some stage of their disease. Altogether there were 39 episodes of pulmonary haemorrhage, 25 being relapses. During seven episodes the chest radiograph was normal. Relapses of pulmonary haemorrhage never occurred in isolation but were usually associated with infection (not necessarily a chest infection) or occasionally fluid overload. Conversely fluid overload or infection were always associated with pulmonary haemorrhage provided there were high or rising titres of circulating antibodies at the time. Therefore in a patient with antiglomerular basement membrane antibody disease, the presence of shadowing in the lung fields on the chest radiograph almost invariably means the patient has pulmonary haemorrhage whether or not pulmonary oedema or a chest infection are present. Limitation of shadowing by a fissure, loss of major portions of the diaphragmatic or cardiac silhouette, involvement of the lung apex or costophrenic angles suggest an underlying chest infection. Septal lines suggest fluid overload. Pleural effusions are seen with chest infections and fluid overload. The carbon monoxide uptake (KCO) was invariably high in the presence of pulmonary haemorrhage even if the chest radiograph was normal. A combined use of KCO and chest radiographs is the best method of monitoring lung disease in these patients. (author)

Anti-glomerular basement membrane: A rare cause of renal failure in children

Directory of Open Access Journals (Sweden)

Indira Agarwal

2017-01-01

Full Text Available Anti-glomerular basement membrane (GBM disease is a rare cause of acute renal failure and known to have bad prognosis regarding renal functions recovery and patient survival specially when diagnosed late and presents with severe renal failure that requires dialysis. We report a case of 11-year-old child with acute renal failure secondary to anti-GBM disease and associated with antineutrophil cytoplasmic antibody-positive vasculitis. He was treated with plasmapheresis, steroids, and cyclophosphamide with recovery of his kidney functions.

Regeneration of defective epithelial basement membrane and restoration of corneal transparency

Science.gov (United States)

Marino, Gustavo K.; Santhiago, Marcony R.; Santhanam, Abirami; Torricelli, Andre A. M.; Wilson, Steven E.

2018-01-01

PURPOSE To study regeneration of the normal ultrastructure of the epithelial basement membrane (EBM) in rabbit corneas that had -9D photorefractive keratectomy (PRK) and developed late haze (fibrosis) with restoration of transparency over one to four months after surgery and in corneas that had incisional wounds. METHODS Twenty-four rabbits had one of their eyes included into one of the two procedure groups (-9D PRK or nearly full-thickness incisional wounds), while the opposite eye serving as unwounded controls. All corneas were evaluated with slit lamp photos, transmission electron microscopy and immunohistochemistry for the myofibroblast marker alpha-smooth muscle actin and collagen type III. RESULTS In the ‘-9D PRK group’, corneas at one month after surgery had dense corneal haze and no evidence of regenerated EBM ultrastructure. By two months after surgery, however, small areas of stromal clearing began to appear within the confluent opacity (lacunae), and these corresponded to small islands of normally-regenerated EBM detected within larger area of the excimer laser-ablated zone with no evidence of normal EBM. By four months after surgery, the EBM was fully-regenerated and the corneal transparency was completely restored to the ablated zone. In the ‘Incisional wound group’, the two dense, linear corneal opacities were observed at one month after surgery and progressively faded by two and three months after surgery. The EBM ultrastructure was fully regenerated at the site of the incisions, including around epithelial plugs that extended into the stroma, by one month after surgery in all eyes. CONCLUSIONS In the rabbit model, spontaneous resolution of corneal fibrosis (haze) after high correction PRK is triggered by regeneration of EBM with normal ultrastructure in the excimer laser- ablated zone. Conversely, incisional wounds heal in rabbit corneas without the development of myofibroblasts because the EBM regenerates normally by one month after surgery

Comparative effectiveness of using resorbable membranes of polylactic acid and collagen in regeneration of bone defects in patients with periimplantitis

Directory of Open Access Journals (Sweden)

Gudaryan A.A.

2014-03-01

Full Text Available The article presents the results of comparative study of effectiveness of usage of separation membranes from polylactic acid (PLA and collagen in carrying out targeted regeneration of bone tissue in 22 patients with periimplantitis. Purpose: To conduct a comparative clinico-radiological efficiency of PLA membrane and collagen membranes in removing bone defects of the alveolar bone in patients with periimplantitis in clinic. It was found that depending on the type of membrane, bone tissue growth occurs not in the same way. Surgery in treatment of periimplantitis using osteo-inducing agent «Bio-Oss» and PLA membranes allows to reach full recovery of bone in bone defects in 90.9 % of patients versus 63.63 % of cases with collagen membranes. Thus, reconstitution of bone in bone defects in periimplantitis is more of full value in using PLA membranes than with membranes from collagen.

Distribution of two basement membrane proteoglycans through hair follicle development and the hair growth cycle in the rat

DEFF Research Database (Denmark)

Couchman, J R; King, J L; McCarthy, K J

1990-01-01

The distribution of two distinct populations of basement membrane proteoglycans has been monitored through hair growth development in the rat embryo and subsequent hair growth cycle. An antiserum against a small heparan sulfate proteoglycan uniformly stained the dermal-epidermal junction...... of embryonic rats throughout the period of hair follicle formation. On the other hand, monoclonal antibodies recognizing a basement membrane-specific chondroitin sulfate proteoglycan only weakly stained 16-d embryo dermal-epidermal junction, but strong staining was associated with hair follicle buds...... as they developed. Through the hair growth cycle, it was found that the heparan sulfate proteoglycan persisted around the follicles, while the chondroitin sulfate proteoglycan decreased in amount through catagen until it was undetectable at the base and dermal papilla of the telogen follicle. As anagen commenced...

Ridge preservation of extraction sockets with chronic pathology using Bio-Oss® Collagen with or without collagen membrane: an experimental study in dogs.

Science.gov (United States)

Kim, Jung-Ju; Schwarz, Frank; Song, Hyun Young; Choi, YoonMi; Kang, Kyung-Rim; Koo, Ki-Tae

2017-06-01

This study aimed to evaluate the dynamics of newly bone formation and dimensional change in diseased extraction sockets using Bio-Oss ® Collagen with or without a collagen membrane. In six beagle dogs, right and left 3rd and 4th mandibular premolars were hemisected and the distal roots were removed. Combined endodontic-periodontic lesions were induced in all sites using black silk, collagen sponge, endodontic files, and application of Porphyromonas gingivalis. After 4 months, among 4 premolars, three teeth were randomly selected per dog and allocated to the following experimental groups: Control group (no treatment but debridement), Test 1 group (only Bio-Oss ® Collagen graft), and Test 2 group (Bio-Oss ® Collagen graft with a collagen membrane). After 7 months from the baseline, the beagle dogs were sacrificed for histomorphometric and Micro-CT analysis. The vertical distance between buccal and lingual crests in the Control group (2.22 ± 0.26 mm) and Test 2 group (1.80 ± 0.16 mm) was significantly different. The socket of the Test 2 group (27.04 ± 5.25%) was occupied by a greater quantity of bone graft compared to the Test 1 group (18.49 ± 2.11%). Ridge preservation in diseased extraction sockets could compensate for buccal bone resorption by contact osteogenesis surrounding the bone graft particles at the bucco-coronal area during socket healing, and the application of a collagen membrane at the entrance of the socket is useful for preserving graft material at the coronal part of the socket. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Evaluating adhesion reduction efficacy of type I/III collagen membrane and collagen-GAG resorbable matrix in primary flexor tendon repair in a chicken model.

Science.gov (United States)

Turner, John B; Corazzini, Rubina L; Butler, Timothy J; Garlick, David S; Rinker, Brian D

2015-09-01

Reduction of peritendinous adhesions after injury and repair has been the subject of extensive prior investigation. The application of a circumferential barrier at the repair site may limit the quantity of peritendinous adhesions while preserving the tendon's innate ability to heal. The authors compare the effectiveness of a type I/III collagen membrane and a collagen-glycosaminoglycan (GAG) resorbable matrix in reducing tendon adhesions in an experimental chicken model of a "zone II" tendon laceration and repair. In Leghorn chickens, flexor tendons were sharply divided using a scalpel and underwent repair in a standard fashion (54 total repairs). The sites were treated with a type I/III collagen membrane, collagen-GAG resorbable matrix, or saline in a randomized fashion. After 3 weeks, qualitative and semiquantitative histological analysis was performed to evaluate the "extent of peritendinous adhesions" and "nature of tendon healing." The data was evaluated with chi-square analysis and unpaired Student's t test. For both collagen materials, there was a statistically significant improvement in the degree of both extent of peritendinous adhesions and nature of tendon healing relative to the control group. There was no significant difference seen between the two materials. There was one tendon rupture observed in each treatment group. Surgical handling characteristics were subjectively favored for type I/III collagen membrane over the collagen-GAG resorbable matrix. The ideal method of reducing clinically significant tendon adhesions after injury remains elusive. Both materials in this study demonstrate promise in reducing tendon adhesions after flexor tendon repair without impeding tendon healing in this model.

Bone conditioned media (BCM) improves osteoblast adhesion and differentiation on collagen barrier membranes.

Science.gov (United States)

Fujioka-Kobayashi, Masako; Caballé-Serrano, Jordi; Bosshardt, Dieter D; Gruber, Reinhard; Buser, Daniel; Miron, Richard J

2016-07-04

The use of autogenous bone chips during guided bone regeneration procedures has remained the gold standard for bone grafting due to its excellent combination of osteoconduction, osteoinduction and osteogenesis. Recent protocols established by our group have characterized specific growth factors and cytokines released from autogenous bone that have the potential to be harvested and isolated into bone conditioned media (BCM). Due to the advantageous osteo-promotive properties of BCM, the aims of the present study was to pre-coat collagen barrier membranes with BCM and investigate its effect on osteoblast adhesion, proliferation and differentiation for possible future clinical use. Scanning electron microscopy (SEM) was first used to qualitative assess BCM protein accumulation on the surface of collagen membranes. Thereafter, undifferentiated mouse ST2 stromal bone marrow cells were seeded onto BioGide porcine derived collagen barrier membranes (control) or barrier membranes pre-coated with BCM (test group). Control and BCM samples were compared for cell adhesion at 8 h, cell proliferation at 1, 3 and 5 days and real-time PCR at 5 days for osteoblast differentiation markers including Runx2, alkaline phosphatase (ALP), osteocalcin (OCN) and bone sialoprotein (BSP). Mineralization was further assessed with alizarin red staining at 14 days post seeding. SEM images demonstrated evidence of accumulated proteins found on the surface of collagen membranes following coating with BCM. Analysis of total cell numbers revealed that the additional pre-coating with BCM markedly increased cell attachment over 4 fold when compared to cells seeded on barrier membranes alone. No significant difference could be observed for cell proliferation at all time points. BCM significantly increased mRNA levels of osteoblast differentiation markers including ALP, OCN and BSP at 5 days post seeding. Furthermore, barrier membranes pre-coated with BCM demonstrated a 5-fold increase in alizarin

Pentoxifylline Ameliorates Glomerular Basement Membrane Ultrastructural Changes Caused by Gentamicin Administration in Rats

Directory of Open Access Journals (Sweden)

Nenad Stojiljković

2009-08-01

Full Text Available Gentamicin is commonly used for the treatment of severe gram negative bacterial infections but inevi-tably cause renal failure during prolonged use. The aim of our study was to emphasize protective effects of pentoxifylline on glomerular basement membrane (GBM alterations induced by gentamicin in rats. Experiments were done on 40 male Wistar rats divided in three experimental groups. GM-group was treated daily with gentamicin in dose of 100 mg/kg during 8 days. PTX-group was treated daily with pentoxifylline in dose of 45 mg/kg and the same dose of gentamicin as in GM-group during 8 days. The control group received 1 ml/day saline intraperitoneally. Morphometric parameter measured during the analysis was glomerular basement membrane thickness. In GM-group of animals glomeruli were en-larged and GMB was diffusely and unequally thickened with neutrophil cells infiltration. In proximal tu-bules epithelial cells, vacuolization of cytoplasm with coagulation-type necrosis were observed. In PTX-group of animals glomeruli were somewhat enlarged and GBM was thickened only in some segments. Coagulation-type necrosis was not found. Blood urea and serum creatinine concentration in GM-group were significantly elevated in comparison with PTX-group while potassium level was decreased. Our results suggest that PTX has protective effects on GBM and proximal tubules in GM-treated rats.

Tumstatin, a Matrikine Derived from Collagen Type IVα3, is Elevated in Serum from Patients with Non-Small Cell Lung Cancer

DEFF Research Database (Denmark)

Nielsen, Signe Holm; Willumsen, Nicholas; Brix, Susanne

2018-01-01

of patients with idiopathic pulmonary fibrosis (IPF), chronic obstructive pulmonary disease (COPD), and non–small cell lung cancer (NSCLC) belonging to two cohorts. RESULTS: The developed TUM enzyme-linked immunosorbent assay (ELISA) was technically robust. In cohort 1, levels of TUM were significantly higher......OBJECTIVES: Fibrosis and cancer are characterized by extracellular matrix (ECM) remodeling. The basement membrane is mainly composed by collagen type IV and laminin. Tumstatin is a matrix metalloproteinase-9 (MMP-9) generated matrikine of collagen type IV α3 chain. We evaluated the potential...

Gas Transfer in Cellularized Collagen-Membrane Gas Exchange Devices.

Science.gov (United States)

Lo, Justin H; Bassett, Erik K; Penson, Elliot J N; Hoganson, David M; Vacanti, Joseph P

2015-08-01

Chronic lower respiratory disease is highly prevalent in the United States, and there remains a need for alternatives to lung transplant for patients who progress to end-stage lung disease. Portable or implantable gas oxygenators based on microfluidic technologies can address this need, provided they operate both efficiently and biocompatibly. Incorporating biomimetic materials into such devices can help replicate native gas exchange function and additionally support cellular components. In this work, we have developed microfluidic devices that enable blood gas exchange across ultra-thin collagen membranes (as thin as 2 μm). Endothelial, stromal, and parenchymal cells readily adhere to these membranes, and long-term culture with cellular components results in remodeling, reflected by reduced membrane thickness. Functionally, acellular collagen-membrane lung devices can mediate effective gas exchange up to ∼288 mL/min/m(2) of oxygen and ∼685 mL/min/m(2) of carbon dioxide, approaching the gas exchange efficiency noted in the native lung. Testing several configurations of lung devices to explore various physical parameters of the device design, we concluded that thinner membranes and longer gas exchange distances result in improved hemoglobin saturation and increases in pO2. However, in the design space tested, these effects are relatively small compared to the improvement in overall oxygen and carbon dioxide transfer by increasing the blood flow rate. Finally, devices cultured with endothelial and parenchymal cells achieved similar gas exchange rates compared with acellular devices. Biomimetic blood oxygenator design opens the possibility of creating portable or implantable microfluidic devices that achieve efficient gas transfer while also maintaining physiologic conditions.

Peroxidasin-mediated crosslinking of collagen IV is independent of NADPH oxidases

Directory of Open Access Journals (Sweden)

Gábor Sirokmány

2018-06-01

Full Text Available Collagen IV is a major component of the basement membrane in epithelial tissues. The NC1 domains of collagen IV protomers are covalently linked together through sulfilimine bonds, the formation of which is catalyzed by peroxidasin. Although hydrogen peroxide is essential for this reaction, the exact source of the oxidant remains elusive. Members of the NOX/DUOX NADPH oxidase family are specifically devoted to the production of superoxide and hydrogen peroxide. Our aim in this study was to find out if NADPH oxidases contribute in vivo to the formation of collagen IV sulfilimine crosslinks. We used multiple genetically modified in vivo model systems to provide a detailed assessment of this question. Our data indicate that in various peroxidasin-expressing tissues sulfilimine crosslinks between the NC1 domains of collagen IV can be readily detected in the absence of functioning NADPH oxidases. We also analyzed how subatmospheric oxygen levels influence the collagen IV network in collagen-producing cultured cells with rapid matrix turnover. We showed that collagen IV crosslinks remain intact even under strongly hypoxic conditions. Our hypothesis is that during collagen IV network formation PXDN cooperates with a NOX/DUOX-independent H2O2 source that is functional also at very low ambient oxygen levels. Keywords: Peroxidasin, NADPH oxidase, Hydrogen peroxide, Collagen IV, Sulfilimine

Immunological characterization of a basement membrane-specific chondroitin sulfate proteoglycan

DEFF Research Database (Denmark)

McCarthy, K J; Accavitti, M A; Couchman, J R

1989-01-01

with the proteoglycan preparation and four mAbs recognizing the core protein of a high-density, buoyant chondroitin sulfate proteoglycan were raised. Confirmation of antibody specificity was carried out by the preparation of affinity columns made from each of the mAbs. Chondroitin sulfate proteoglycans (CSPGs) were...... (Mr = 5-6 x 10(5)), with a core protein of Mr = approximately 1.5-1.6 x 10(5) and composed exclusively of chondroitin sulfate chains with an average Mr = 1.6-1.8 x 10(4). In addition, a CSPG was purified from adult rat kidney, whose core protein was also Mr = 1.6 x 10(5). The proteoglycan and its core...... sulfate proteoglycans, it therefore appears that at least one CSPG is a widespread basement membrane component....

Glycosaminoglycans and fibrillar collagen in Priapulida: a histo- and cytochemical study.

Science.gov (United States)

Welsch, U; Erlinger, R; Storch, V

1992-12-01

The distribution of glycosaminoglycans and fibrillar collagen was studied in various tissues of priapulids, which represent an ancient group of marine metazoa. Sulphated glycosaminoglycans, as demonstrated at the electron microscopical level by Cupromeronic blue, were predominantly found in the cuticle, in basement membranes and also in the narrow connective tissue space below epidermis and anterior intestine. On the basis of their morphology the Cupromeronic blue precipitates could be divided into several groups. Fibrillar collagen occurred in the connective tissue under the epidermis and the epithelium of the anterior intestine. The spatial interrelationship between fibrillar collagen and glycosaminoglycans lacked with some exceptions, the high regularity found in connective tissues of other invertebrates and of vertebrates. This might be related to the special skeletal system of priapulids, consisting mainly of a strong extracellular cuticle and the turgor of the fluid-filled body cavity. In such a system the usual supportive structures seem to be of less functional significance.

Expression of periglandular tenascin-C and basement membrane laminin in normal prostate, benign prostatic hyperplasia and prostate carcinoma

NARCIS (Netherlands)

Xue, Y.; Li, J.; Latijnhouwers, M. A.; Smedts, F.; Umbas, R.; Aalders, T. W.; Debruyne, F. M.; de la Rosette, J. J.; Schalken, J. A.

1998-01-01

To evaluate the structural relationship of the distribution between tenascin (tenascin-C, an extra-cellular matrix glycoprotein involved in stromal-epithelial interactions in both normal and pathological conditions) and laminin, an important component of the basement membrane, in normal and

Evaluation and Comparison of the Biopathology of Collagen and Inflammation in the Extracellular Matrix of Oral Epithelial Dysplasias and Inflammatory Fibrous Hyperplasia Using Picrosirius Red Stain and Polarising Microscopy: A Preliminary Study.

Science.gov (United States)

Varghese, Soma Susan; Sarojini, Sreenivasan Bargavan; George, Giju Baby; Vinod, Sankar; Mathew, Philips; Babu, Anulekh; Sebastian, Joseph

2015-12-01

The role of tumour inflammation and the dysplastic epithelial-stromal interactions on the nature of collagen fibres in the extracellular matrix of dysplastic epithelium is not fully understood. The present study was aimed to evaluate and compare the inflammation and pathological stromal collagen (loosely packed thin disorganized collagen) present in mild, moderate and severe epithelial dysplasias with that of inflammatory fibrous hyperplasias. The basement membrane intactness of epithelial dysplasias was also evaluated to determine if dysplastic epithelial mesenchymal interaction has any role in the integrity of stromal collagen in epithelial dysplasia. Oral epithelial dysplasias, inflammatory fibrous hyperplasia and normal oral mucosal samples were used for the study. Packing, thickness and orientation of collagen fibres in mild, moderate and severe grades of oral epithelial dysplasias (n = 24), inflammatory fibrous hyperplasia (n = 8) and normal oral mucosal samples (n = 8) were analysed based on the polarisation of collagen fibres in picrosirius red polarising stain under polarising microscope. All the grades of epithelial dysplasias showed greenish yellow birefringence confirming the presence of loosely arranged pathological collagen in the presence of moderate inflammation. All the cases of inflammatory fibrous hyperplasia showed red polarisation hue and moderate inflammation. A statistically significant difference was found in the packing and orientation of collagen when epithelial dysplasias and inflammatory fibrous hyperplasia were compared (P collagen even in mild epithelial dysplasia suggests that tumourigenic factors are released to connective tissue stroma much earlier than expected. Hence we suggest considering the integrity of extracellular matrix collagen, intactness of basement membrane and inflammation associated with dysplasia along with the anaplasia of epithelial cells in the microscopic assessment of dysplastic epithelium.

Collagen VII deficient mice show morphologic and histologic corneal changes that phenotypically mimic human dystrophic epidermolysis bullosa of the eye.

Science.gov (United States)

Chen, Vicki M; Shelke, Rajani; Nyström, Alexander; Laver, Nora; Sampson, James F; Zhiyi, Cao; Bhat, Najma; Panjwani, Noorjahan

2018-06-16

Absence of collagen VII causes blistering of the skin, eyes and many other tissues. This disease is termed dystrophic epidermolysis bullosa (DEB). Corneal fibrosis occurs in up to 41% and vision loss in up to 64% of patients. Standard treatments are supportive and there is no cure. The immune-histologic and morphologic changes in the corneas of the mouse model for this disease have not been described in the literature. Our purpose is to characterize the eyes of these mice to determine if this is an appropriate model for study of human therapeutics. Western blot analysis (WB) and immunohistochemistry (IHC) were performed to assess the relative collagen VII protein levels and its location within the cornea. Additional IHC for inflammatory and fibrotic biomarkers alpha-smooth muscle actin (α-SMA), transforming growth factor-beta (TGF-β), connective tissue growth factor (CTGF), proteinase 3, tenascin C and collagen III were performed. Clinical photographs documenting opacification of the corneas of animals of differing ages were assessed and scored independently by 2 examiners. Histology was then used to investigate morphologic changes. IHC and WB confirmed that these mice are deficient in collagen VII production at the level of the basement membrane when compared with wild-types. IHC showed anomalous deposition of collagen III throughout the stroma. Of the 5 biomarkers tested, TGF-β showed the strongest and most consistently staining. Photographs documented corneal opacities only in mice older than 10 weeks, opacities were not seen in younger animals. Histology showed multiple abnormalities, including epithelial hyperplasia, ulceration, fibrosis, edema, dysplasia, neovascularization and bullae formation. The collagen VII hypomorphic mouse shows reduced collagen VII production at the level of the corneal basement membrane. Corneal changes are similar to pathology seen in humans with this disease. The presence of anomalous stromal collagen III and TGF-β appear to be

Permeation of macromolecules into the renal glomerular basement membrane and capture by the tubules

Science.gov (United States)

Lawrence, Marlon G.; Altenburg, Michael K.; Sanford, Ryan; Willett, Julian D.; Bleasdale, Benjamin; Ballou, Byron; Wilder, Jennifer; Li, Feng; Miner, Jeffrey H.; Berg, Ulla B.; Smithies, Oliver

2017-01-01

How the kidney prevents urinary excretion of plasma proteins continues to be debated. Here, using unfixed whole-mount mouse kidneys, we show that fluorescent-tagged proteins and neutral dextrans permeate into the glomerular basement membrane (GBM), in general agreement with Ogston's 1958 equation describing how permeation into gels is related to molecular size. Electron-microscopic analyses of kidneys fixed seconds to hours after injecting gold-tagged albumin, negatively charged gold nanoparticles, and stable oligoclusters of gold nanoparticles show that permeation into the lamina densa of the GBM is size-sensitive. Nanoparticles comparable in size with IgG dimers do not permeate into it. IgG monomer-sized particles permeate to some extent. Albumin-sized particles permeate extensively into the lamina densa. Particles traversing the lamina densa tend to accumulate upstream of the podocyte glycocalyx that spans the slit, but none are observed upstream of the slit diaphragm. At low concentrations, ovalbumin-sized nanoparticles reach the primary filtrate, are captured by proximal tubule cells, and are endocytosed. At higher concentrations, tubular capture is saturated, and they reach the urine. In mouse models of Pierson’s or Alport’s proteinuric syndromes resulting from defects in GBM structural proteins (laminin β2 or collagen α3 IV), the GBM is irregularly swollen, the lamina densa is absent, and permeation is increased. Our observations indicate that size-dependent permeation into the lamina densa of the GBM and the podocyte glycocalyx, together with saturable tubular capture, determines which macromolecules reach the urine without the need to invoke direct size selection by the slit diaphragm. PMID:28246329

[Morphology of basement membrane and associated matrix proteins in normal and pathological tissues].

Science.gov (United States)

Nerlich, A

1995-01-01

Basement membranes (BM) are specialized structures of the extracellular matrix. Their composition is of particular importance for the maintenance of normal morphological and functional properties of a multitude of organs and tissue systems and it is thus required for regular homeostasis of body function. Generally, they possess three main functions, i.e. participation in the maintenance of tissue structure, control of fluid and substrate exchange, and regulation of cell growth and differentiation. BMs are made up by various components which are in part specifically localized within the BM zone, or which represent ubiquitous matrix constituents with specific quantitative and/or qualitative differences in their localization. On the basis of a thorough immunohistochemical analysis of normal and diseased tissues, we provide here a concept of "functional morphology/pathomorphology" of the different BM components analyzed: 1.) The ubiquitous BM-constituent collagen IV primarily stabilizes the BM-zone and thus represents the "backbone" of the BM providing mechanical strength. Its loss leads to cystic tissue transformation as it is evidenced from the analysis of polycystic nephropathies. Thus, in other cystic tissue transformations a similar formal pathogenesis may be present. 2.) The specific localization of collagen VII as the main structural component of anchoring fibrils underlines the mechanical anchoring function of this collagenous protein. Defects in this protein lead to hereditary epidermolysis. The rapid re-occurrence of epidermal collagen VII during normal human wound healing indicates a quick reconstitution of the mechanical tensile strength of healing wounds. 3.) The BM-specific heparan sulfate proteoglycan (HSPG, Perlecan) with its highly negative anionic charge can be assumed to exert filter control. This assumption is corroborated by the localizatory findings of a preferential deposition of HSPG in endothelial and particularly in glomerular BM. Similarly

Spumiform basement membrane aberrations in the microvasculature of the midbrain periaqueductal gray region in hamster : Rostro-caudal pathogenesis?

NARCIS (Netherlands)

Gerrits, P.O.; Kortekaas, R.; de Weerd, Heleen; Luiten, P.G.M.; van der Want, J.J.L.; Veening, Jan

2013-01-01

Spumiform basement membrane degeneration (sbmd) is a specific kind of aberration present in the capillaries of the midbrain periaqueductal gray (PAG) region of the senescent hamster. These capillaries, separated by the ependymal cell layer, are bordering the Sylvian cerebral aqueduct. The aqueduct,

Spumiform basement membrane aberrations in the microvasculature of the midbrain periaqueductal gray region in hamster: rostro-caudal pathogenesis?

NARCIS (Netherlands)

Gerrits, P.O.; Kortekaas, R.; Weerd, H. de; Luiten, P.G.M.; Want, J.J. van der; Veening, J.G.

2013-01-01

Spumiform basement membrane degeneration (sbmd) is a specific kind of aberration present in the capillaries of the midbrain periaqueductal gray (PAG) region of the senescent hamster. These capillaries, separated by the ependymal cell layer, are bordering the Sylvian cerebral aqueduct. The aqueduct,

Expression and deposition of basement membrane proteins by brain capillary endothelial cells in a primary murine model of the blood-brain barrier

DEFF Research Database (Denmark)

Thomsen, Maj Schneider; Birkelund, Svend; Larsen, Annette Burkhart

2016-01-01

The blood-brain barrier (BBB) represents the interface between the blood and the brain parenchyma and consists of endothelial cells which are tightly sealed together by tight junction proteins. The endothelial cells are in addition supported by pericytes, which are embedded in the vascular basement...... of the present study was to create four different in vitro constructs of the murine BBB to characterise if the expression and secretion of basement membrane proteins by the murine brain capillary endothelial cells (mBCECs) was affected by co-culturing with pericytes, mixed glial cells, or both. Primary m......BCECs and pericytes were isolated from brains of adult mice. Mixed glial cells were prepared from cerebral cortices of newborn mice. The mBCECs were grown as mono-culture, or co-cultured with pericytes, mixed glial cells, or both. To study the expression of basement membrane proteins RT-qPCR, mass spectrometry...

Basement membrane heparan sulfate proteoglycan from the L2 rat yolk sac carcinoma

DEFF Research Database (Denmark)

Fenger, M; Wewer, U; Albrechtsen, R

1984-01-01

Heparan sulfate proteoglycan from the L2 rat yolk sac carcinoma has been purified and partially characterized. The proteoglycan has an apparent Mr of 750 000, 35% of which represents the core protein. The core protein seems to be homogeneous, whereas the heparan sulfate chains are heterogeneous...... with an Mr of about 50 000-70 000, with 30% of the glucosamine being N-sulfated. Antibodies raised against the core protein of the heparan sulfate proteoglycan reacted with basement membranes of various rat and human tissue....

Evaluation of 3D printed PCL/PLGA/β-TCP versus collagen membranes for guided bone regeneration in a beagle implant model.

Science.gov (United States)

Won, J-Y; Park, C-Y; Bae, J-H; Ahn, G; Kim, C; Lim, D-H; Cho, D-W; Yun, W-S; Shim, J-H; Huh, J-B

2016-10-07

Here, we compared 3D-printed polycaprolactone/poly(lactic-co-glycolic acid)/β-tricalcium phosphate (PCL/PLGA/β-TCP) membranes with the widely used collagen membranes for guided bone regeneration (GBR) in beagle implant models. For mechanical property comparison in dry and wet conditions and cytocompatibility determination, we analyzed the rate and pattern of cell proliferation of seeded fibroblasts and preosteoblasts using the cell counting kit-8 assay and scanning electron microscopy. Osteogenic differentiation was verified using alizarin red S staining. At 8 weeks following implantation in vivo using beagle dogs, computed tomography and histological analyses were performed after sacrifice. Cell proliferation rates in vitro indicated that early cell attachment was higher in collagen than in PCL/PLGA/β-TCP membranes; however, the difference subsided by day 7. Similar outcomes were found for osteogenic differentiation, with approximately 2.5 times greater staining in collagen than PCL/PLGA/β-TCP, but without significant difference by day 14. In vivo, bone regeneration in the defect area, represented by new bone formation and bone-to-implant contact, paralleled those associated with collagen membranes. However, tensile testing revealed that whereas the PCL/PLGA/β-TCP membrane mechanical properties were conserved in both wet and dry states, the tensile property of collagen was reduced by 99% under wet conditions. Our results demonstrate in vitro and in vivo that PCL/PLGA/β-TCP membranes have similar levels of biocompatibility and bone regeneration as collagen membranes. In particular, considering that GBR is always applied to a wet environment (e.g. blood, saliva), we demonstrated that PCL/PLGA/β-TCP membranes maintained their form more reliably than collagen membranes in a wet setting, confirming their appropriateness as a GBR membrane.

Agrin is a major heparan sulfate proteoglycan in the human glomerular basement membrane.

Science.gov (United States)

Groffen, A J; Ruegg, M A; Dijkman, H; van de Velden, T J; Buskens, C A; van den Born, J; Assmann, K J; Monnens, L A; Veerkamp, J H; van den Heuvel, L P

1998-01-01

Agrin is a heparan sulfate proteoglycan (HSPG) that is highly concentrated in the synaptic basal lamina at the neuromuscular junction (NMJ). Agrin-like immunoreactivity is also detected outside the NMJ. Here we show that agrin is a major HSPG component of the human glomerular basement membrane (GBM). This is in addition to perlecan, a previously characterized HSPG of basement membranes. Antibodies against agrin and against an unidentified GBM HSPG produced a strong staining of the GBM and the NMJ, different from that observed with anti-perlecan antibodies. In addition, anti-agrin antisera recognized purified GBM HSPG and competed with an anti-GBM HSPG monoclonal antibody in ELISA. Furthermore, both antibodies recognized a molecule that migrated in SDS-PAGE as a smear and had a molecular mass of approximately 200-210 kD after deglycosylation. In immunoelectron microscopy, agrin showed a linear distribution along the GBM and was present throughout the width of the GBM. This was again different from perlecan, which was exclusively present on the endothelial side of the GBM and was distributed in a nonlinear manner. Quantitative ELISA showed that, compared with perlecan, the agrin-like GBM HSPG showed a sixfold higher molarity in crude glomerular extract. These results show that agrin is a major component of the GBM, indicating that it may play a role in renal ultrafiltration and cell matrix interaction. (J Histochem Cytochem 46:19-27, 1998)

Extraction Socket Preservation Using Porcine-Derived Collagen Membrane Alone or Associated with Porcine-Derived Bone. Clinical Results of Randomized Controlled Study

Directory of Open Access Journals (Sweden)

Renzo Guarnieri

2017-03-01

Full Text Available Objectives: The aim of present randomized controlled clinical trial was to clinically evaluate hard tissue changes after extraction socket preservation procedures compared to natural spontaneous healing. Material and Methods: Thirty patients were enrolled in the present study and underwent single-tooth extraction in the premolar/molar areas. Ten sites were grafted with porcine-derived bone covered by collagen membrane, 10 covered by porcine-derived collagen membrane alone, and 10 underwent natural spontaneous healing. Vertical and horizontal bone changes after 3-month were evaluated at implant placement. Results: The vertical and horizontal bone changes at the extraction sockets treated with collagen membrane alone (vertical: -0.55 [SD 0.11] mm, and horizontal: -1.21 [SD 0.69] mm and collagen membrane plus porcine-derived bone (vertical: -0.37 [SD 0.7] mm, and horizontal: -0.91 [SD 0.53] mm were found significantly lower (P < 0.001, when compared to non-grafted sockets (vertical: -2.09 [SD 0.19] mm, and horizontal: -3.96 [SD 0.87] mm. In type 1 extraction sockets, in premolar sites, and in presence of vestibular bone thicknesses ≥ 1.5 mm, the use of collagen membrane alone revealed similar outcomes to those with additional graft material. Conclusions: At the re-entry surgery, extraction sockets grafted with porcine-derived bone and covered by collagen membrane, and extraction sockets covered by porcine-derived collagen membrane alone, showed significantly lower vertical and horizontal bone changes, compared to extraction sockets sites underwent natural spontaneous healing. However, a complete prevention of remodelling is not achievable, irrespective of the technique used.

Intercellular deposits of basement membrane material in active human pituitary adenomas detected by immunostaining for laminin and electron microscopy

DEFF Research Database (Denmark)

Holck, S; Wewer, U M; Albrechtsen, R

1986-01-01

and one patient with Cushing's syndrome). Concurrently, at the ultrastructural level, bunches of basement membrane-like material intermingled between the adenoma cells were demonstrated in seven of these ten active adenomas. Furthermore, secretory granules were entrapped occasionally in this intercellular...

Quaternary epitopes of α345(IV) collagen initiate Alport post-transplant anti-GBM nephritis

DEFF Research Database (Denmark)

Olaru, Florina; Luo, Wentian; Wang, Xu-Ping

2013-01-01

Alport post-transplant nephritis (APTN) is an aggressive form of anti-glomerular basement membrane disease that targets the allograft in transplanted patients with X-linked Alport syndrome. Alloantibodies develop against the NC1 domain of α5(IV) collagen, which occurs in normal kidneys, including...... of alloantibodies against allogeneic collagen IV. Some alloantibodies targeted alloepitopes within α5NC1 monomers, shared by α345NC1 and α1256NC1 hexamers. Other alloantibodies specifically targeted alloepitopes that depended on the quaternary structure of α345NC1 hexamers. In Col4a5-null mice, immunization...... with native forms of allogeneic collagen IV exclusively elicited antibodies to quaternary α345NC1 alloepitopes, whereas alloimmunogens lacking native quaternary structure elicited antibodies to shared α5NC1 alloepitopes. These results imply that quaternary epitopes within α345NC1 hexamers may initiate...

Overexpression of β1-chain-containing laminins in capillary basement membranes of human breast cancer and its metastases

Science.gov (United States)

Fujita, Manabu; Khazenzon, Natalya M; Bose, Shikha; Sekiguchi, Kiyotoshi; Sasaki, Takako; Carter, William G; Ljubimov, Alexander V; Black, Keith L; Ljubimova, Julia Y

2005-01-01

Introduction Laminins are the major components of vascular and parenchymal basement membranes. We previously documented a switch in the expression of vascular laminins containing the α4 chain from predominantly laminin-9 (α4β2γ1) to predominantly laminin-8 (α4β1γ1) during progression of human brain gliomas to high-grade glioblastoma multiforme. Here, differential expression of laminins was studied in blood vessels and ductal epithelium of the breast. Method In the present study the expressions of laminin isoforms α1–α5, β1–β3, γ1, and γ2 were examined during progression of breast cancer. Forty-five clinical samples of breast tissues including normal breast, ductal carcinomas in situ, invasive ductal carcinomas, and their metastases to the brain were compared using Western blot analysis and immunohistochemistry for various chains of laminin, in particular laminin-8 and laminin-9. Results Laminin α4 chain was observed in vascular basement membranes of most studied tissues, with the highest expression in metastases. At the same time, the expression of laminin β2 chain (a constituent of laminin-9) was mostly seen in normal breast and carcinomas in situ but not in invasive carcinomas or metastases. In contrast, laminin β1 chain (a constituent of laminin-8) was typically found in vessel walls of carcinomas and their metastases but not in those of normal breast. The expression of laminin-8 increased in a progression-dependent manner. A similar change was observed from laminin-11 (α5β2γ1) to laminin-10 (α5β1γ1) during breast tumor progression. Additionally, laminin-2 (α2β1γ1) appeared in vascular basement membranes of invasive carcinomas and metastases. Chains of laminin-5 (α3β3γ2) were expressed in the ductal epithelium basement membranes of the breast and diminished with tumor progression. Conclusion These results suggest that laminin-2, laminin-8, and laminin-10 are important components of tumor microvessels and may associate with breast

Overexpression of β1-chain-containing laminins in capillary basement membranes of human breast cancer and its metastases

International Nuclear Information System (INIS)

Fujita, Manabu; Khazenzon, Natalya M; Bose, Shikha; Sekiguchi, Kiyotoshi; Sasaki, Takako; Carter, William G; Ljubimov, Alexander V; Black, Keith L; Ljubimova, Julia Y

2005-01-01

Laminins are the major components of vascular and parenchymal basement membranes. We previously documented a switch in the expression of vascular laminins containing the α4 chain from predominantly laminin-9 (α4β2γ1) to predominantly laminin-8 (α4β1γ1) during progression of human brain gliomas to high-grade glioblastoma multiforme. Here, differential expression of laminins was studied in blood vessels and ductal epithelium of the breast. In the present study the expressions of laminin isoforms α1–α5, β1–β3, γ1, and γ2 were examined during progression of breast cancer. Forty-five clinical samples of breast tissues including normal breast, ductal carcinomas in situ, invasive ductal carcinomas, and their metastases to the brain were compared using Western blot analysis and immunohistochemistry for various chains of laminin, in particular laminin-8 and laminin-9. Laminin α4 chain was observed in vascular basement membranes of most studied tissues, with the highest expression in metastases. At the same time, the expression of laminin β2 chain (a constituent of laminin-9) was mostly seen in normal breast and carcinomas in situ but not in invasive carcinomas or metastases. In contrast, laminin β1 chain (a constituent of laminin-8) was typically found in vessel walls of carcinomas and their metastases but not in those of normal breast. The expression of laminin-8 increased in a progression-dependent manner. A similar change was observed from laminin-11 (α5β2γ1) to laminin-10 (α5β1γ1) during breast tumor progression. Additionally, laminin-2 (α2β1γ1) appeared in vascular basement membranes of invasive carcinomas and metastases. Chains of laminin-5 (α3β3γ2) were expressed in the ductal epithelium basement membranes of the breast and diminished with tumor progression. These results suggest that laminin-2, laminin-8, and laminin-10 are important components of tumor microvessels and may associate with breast tumor progression. Angiogenic switch

Rat mesangial cells in vitro synthesize a spectrum of proteoglycan species including those of the basement membrane and interstitium

DEFF Research Database (Denmark)

Thomas, G J; Shewring, L; McCarthy, K J

1995-01-01

is localized in the mesangium but is not found in the pericapillary glomerular basement membrane (GBM). Further characterization of the proteoglycans synthesized by RMC in vitro revealed: (i) a second large CSPG, identified as versican; (ii) two small dermatan sulphate proteoglycans identified as biglycan...

Chondrocyte-seeded type I/III collagen membrane for autologous chondrocyte transplantation

DEFF Research Database (Denmark)

Niemeyer, Philipp; Lenz, Philipp; Kreuz, Peter C

2010-01-01

PURPOSE: We report the 2-year clinical results and identify prognostic factors in patients treated with autologous chondrocyte transplantation by use of a collagen membrane to seed the chondrocytes (ACT-CS). METHODS: This is a prospective study of 59 patients who were treated with ACT......-CS represents a technical modification of membrane-associated autologous chondrocyte transplantation that combines easy handling and attractive application properties with reliable clinical results 24 months after surgery, especially in patients with isolated cartilage defects. Even though the failure rate...

Treatment of gingival recession with collagen membrane and DFDBA: a histometric study in dogs

Directory of Open Access Journals (Sweden)

Elizabeth Pimentel Rosetti

2009-09-01

Full Text Available In a previous study, we evaluated the findings related to the use of resorbable collagen membranes in humans along with DFDBA (demineralized freeze-dried bone allograft. The aim of this subsequent study was to histometrically evaluate in dogs, the healing response of gingival recessions treated with collagen membrane + DFDBA (Guided Tissue Regeneration, GTR compared to a coronally positioned flap (CPF. Two types of treatment were randomly carried out in a split-mouth study. Group 1 was considered as test (GTR: collagen membrane + DFDBA, whereas Group 2 stood for the control (only CPF. The dogs were given chemical bacterial plaque control with 0.2% chlorhexidine digluconate during a 90-day repair period. Afterwards, the animals were killed to obtain biopsies and histometric evaluation of the process of cementum and bone formation, epithelial migration and gingival level. A statistically significant difference was found between groups with a larger extension of neoformed cementum (GTR = 32.72%; CPF = 18.82%; p = 0.0004, new bone (GTR = 23.20%; CPF = 09.90%; p = 0.0401 and with a smaller area of residual gingival recession in the test group (GTR = 50.69%; CPF = 59.73%; p = 0.0055 compared to the control group. The only item assessed that showed no statistical difference was epithelial proliferation on the root surface, with means of 15.14% for the GTR group and 20.34% for the CPF group (p = 0.0890. Within the limits of this study we concluded that the treatment of gingival recession defects with GTR, associating collagen membrane with DFDBA, showed better outcomes in terms of a larger extension of neoformed cementum and bone, as well as in terms of a smaller proportion of residual recessions.

The Peri-islet Basement Membrane, a Barrier to Infiltrating Leukocytes in Type 1 Diabetes in Mouse and Human

DEFF Research Database (Denmark)

Korpos, Eva; Kadri, Nadir; Kappelhoff, Reinhild

2013-01-01

We provide the first comprehensive analysis of the extracellular matrix (ECM) composition of peri-islet capsules, composed of the peri-islet basement membrane (BM) and subjacent interstitial matrix (IM), in development of type 1 diabetes in NOD mice and in human type 1 diabetes. Our data demonstr...... IM are reconstituted once inflammation subsides, indicating that the peri-islet BM-producing cells are not lost due to the inflammation, which has important ramifications to islet transplantation studies.......We provide the first comprehensive analysis of the extracellular matrix (ECM) composition of peri-islet capsules, composed of the peri-islet basement membrane (BM) and subjacent interstitial matrix (IM), in development of type 1 diabetes in NOD mice and in human type 1 diabetes. Our data...... demonstrate global loss of peri-islet BM and IM components only at sites of leukocyte infiltration into the islet. Stereological analyses reveal a correlation between incidence of insulitis and the number of islets showing loss of peri-islet BM versus islets with intact BMs, suggesting that leukocyte...

The Role of Type IV Collagen in Developing Lens in Mouse Fetuses

Directory of Open Access Journals (Sweden)

Mehdi Jalali

2009-09-01

Full Text Available Objective(sExtracellular matrix (ECM and basement membrane (BM play important roles in many developmental processes during development and after birth. Among the components of the BM, collagen fibers specially type IV are the most important parts. The aim of this study was to determine the time when collagen type IV appears in the BM of lens structure during mouse embryonic development.Materials and MethodsIn this experimental study, 22 female Balb/C mice were randomly selected and were kept under normal condition, finding vaginal plug was assumed as day zero of pregnancy. From embryonic day 10 to 20, all specimens were sacrificed by cervical dislocation and their heads were fixed, serially sectioned and immunohistochemistry study for tracing collagen type IV in lens were carried out.ResultsOur data revealed that collagen type IV appeared at the early stage of gestation day 12 in BM of anterior epithelial lens cells and the amount of this protein gradually increased until days 15-17 in ECM and posterior capsule epithelium. After this period, severe reaction was not observed in any part of the lens.ConclusionThese findings establish the important role of collagen IV in developing optic cup and any changes during critical period of pregnancy may be result in severe visual system defect

Guided bone regeneration with asymmetric collagen-chitosan membranes containing aspirin-loaded chitosan nanoparticles

Directory of Open Access Journals (Sweden)

Zhang J

2017-12-01

Full Text Available Jiayu Zhang,1 Shiqing Ma,1 Zihao Liu,1 Hongjuan Geng,1 Xin Lu,1 Xi Zhang,1 Hongjie Li,1 Chenyuan Gao,2 Xu Zhang,1 Ping Gao1 1School of Dentistry, Hospital of Stomatology, Tianjin Medical University, Tianjin, 2Beijing Laboratory of Biomedical Materials, Beijing University of Chemical Technology, Beijing, People’s Republic of China Introduction: Membranes allowing the sustained release of drugs that can achieve cell adhesion are very promising for guided bone regeneration. Previous studies have suggested that aspirin has the potential to promote bone regeneration. The purpose of this study was to prepare a local drug delivery system with aspirin-loaded chitosan nanoparticles (ACS contained in an asymmetric collagen-chitosan membrane (CCM. Methods: In this study, the ACS were fabricated using different concentrations of aspirin (5 mg, 25 mg, 50 mg, and 75 mg. The drug release behavior of ACS was studied. Transmission electron microscopy (TEM and scanning electron microscopy (SEM were used to examine the micromorphology of ACS and aspirin-loaded chitosan nanoparticles contained in chitosan-collagen membranes (ACS-CCM. In vitro bone mesenchymal stem cells (BMSCs were cultured and critical-sized cranial defects on Sprague-Dawley rats were made to evaluate the effect of the ACS-CCM on bone regeneration.Results: Drug release behavior results of ACS showed that the nanoparticles fabricated in this study could successfully sustain the release of the drug. TEM showed the morphology of the nanoparticles. SEM images indicated that the asymmetric membrane comprised a loose collagen layer and a dense chitosan layer. In vitro studies showed that ACS-CCM could promote the proliferation of BMSCs, and that the degree of differentiated BMSCs seeded on CCMs containing 50 mg of ACS was higher than that of other membranes. Micro-computed tomography showed that 50 mg of ACS-CCM resulted in enhanced bone regeneration compared with the control group.Conclusion: This

Goodpasture's autoimmune disease - A collagen IV disorder.

Science.gov (United States)

Pedchenko, Vadim; Richard Kitching, A; Hudson, Billy G

2018-05-12

Goodpasture's (GP) disease is an autoimmune disorder characterized by the deposition of pathogenic autoantibodies in basement membranes of kidney and lung eliciting rapidly progressive glomerulonephritis and pulmonary hemorrhage. The principal autoantigen is the α345 network of collagen IV, which expression is restricted to target tissues. Recent discoveries include a key role of chloride and bromide for network assembly, a novel posttranslational modification of the antigen, a sulfilimine bond that crosslinks the antigen, and the mechanistic role of HLA in genetic susceptibility and resistance to GP disease. These advances provide further insights into molecular mechanisms of initiation and progression of GP disease and serve as a basis for developing of novel diagnostic tools and therapies for treatment of Goodpasture's disease. Copyright © 2017. Published by Elsevier B.V.

Fluid Mechanics of the Vascular Basement Membrane in the Brain

Science.gov (United States)

Coloma, Mikhail; Hui, Jonathan; Chiarot, Paul; Huang, Peter; Carare, Roxana; McLeod, Kenneth; Schaffer, David

2013-11-01

Beta-amyloid is a normal product of brain metabolic function and is found within the interstitial fluid of the brain. Failure of the clearance of beta-amyloid from the aging brain leads to its accumulation within the walls of arteries and to Alzheimer's disease. The vascular basement membrane (VBM) within the walls of cerebral arteries surrounds the spirally arranged smooth muscle cells and represents an essential pathway for removal of beta-amyloid from the brain. This process fails with the stiffening of arterial walls associated with aging. In this study we hypothesize that the deformation of the VBM associated with arterial pulsations drives the interstitial fluid to drain in the direction opposite of the arterial blood flow. This hypothesis is theoretically investigated by modeling the VBM as a thin, coaxial, fluid-filled porous medium surrounding a periodically deforming cylindrical tube. Flow and boundary conditions required to achieve such a backward clearance are derived through a control volume analysis of mass, momentum, and energy.

In vivo turnover of the basement membrane and other heparan sulfate proteoglycans of rat glomerulus

DEFF Research Database (Denmark)

Beavan, L A; Davies, M; Couchman, J R

1989-01-01

The metabolic turnover of rat glomerular proteoglycans in vivo was investigated. Newly synthesized proteoglycans were labeled during a 7-h period after injecting sodium [35S]sulfate intraperitoneally. At the end of the labeling period a chase dose of sodium sulfate was given. Subsequently......-propanesulfonate-4 M guanidine hydrochloride, a procedure which solubilized greater than 95% of the 35S-labeled macromolecules. Of these 11-13% was immunoprecipitated by an antiserum against heparan sulfate proteoglycan which, in immunolocalization experiments, showed specificity for staining the basement membrane...

Collagen macromolecular drug delivery systems

International Nuclear Information System (INIS)

Gilbert, D.L.

1988-01-01

The objective of this study was to examine collagen for use as a macromolecular drug delivery system by determining the mechanism of release through a matrix. Collagen membranes varying in porosity, crosslinking density, structure and crosslinker were fabricated. Collagen characterized by infrared spectroscopy and solution viscosity was determined to be pure and native. The collagen membranes were determined to possess native vs. non-native quaternary structure and porous vs. dense aggregate membranes by electron microscopy. Collagen monolithic devices containing a model macromolecule (inulin) were fabricated. In vitro release rates were found to be linear with respect to t 1/2 and were affected by crosslinking density, crosslinker and structure. The biodegradation of the collagen matrix was also examined. In vivo biocompatibility, degradation and 14 C-inulin release rates were evaluated subcutaneously in rats

Preparation and characterisation of Punica granatum pericarp aqueous extract loaded chitosan-collagen-starch membrane: role in wound healing process.

Science.gov (United States)

Amal, B; Veena, B; Jayachandran, V P; Shilpa, Joy

2015-05-01

Engineered scaffolds made from natural biomaterials are crucial elements in tissue engineering strategies. In this study, biological scaffold like chitosan-collagen-starch membrane (CCSM) loaded with the antibacterial agent, Punica granatum pericarp aqueous extract was explored for enhanced regeneration of epithelial tissue during wound healing. Collagen was extracted from Rachycentron canadum fish skin. Membranous scaffold was prepared by mixing collagen, starch and chitosan in a fixed proportion, loaded with aqueous extract of P. granatum and its anti-pseudomonal activity was studied. Morphological characterization by SEM and mechanical property like tensile strength of the membrane were studied. Excision wound of 2 cm(2) size was induced in Guinea pig and the effect of P. granatum extract loaded CCSM in wound healing was studied. The SEM image showed deep pores in the membrane and also possessed good tensile strength. Wound surface area was reduced prominently in the experimental group with P. granatum extract loaded CCSM when compared to the group with unloaded membrane and the one with no membrane. Punica granatum extract loaded CCSM has antipseudomonal property and supported enhanced epithelial cell proliferation without leaving a scar after wound healing. This has significant therapeutic application in membranous scaffold mediated skin repair and regeneration.

Marine-derived collagen biomaterials from echinoderm connective tissues

KAUST Repository

Ferrario, Cinzia; Leggio, Livio; Leone, Roberta; Di Benedetto, Cristiano; Guidetti, Luca; Coccè , Valentina; Ascagni, Miriam; Bonasoro, Francesco; La Porta, Caterina A.M.; Candia Carnevali, M. Daniela; Sugni, Michela

2016-01-01

The use of marine collagens is a hot topic in the field of tissue engineering. Echinoderms possess unique connective tissues (Mutable Collagenous Tissues, MCTs) which can represent an innovative source of collagen to develop collagen barrier-membranes for Guided Tissue Regeneration (GTR). In the present work we used MCTs from different echinoderm models (sea urchin, starfish and sea cucumber) to produce echinoderm-derived collagen membranes (EDCMs). Commercial membranes for GTR or soluble/reassembled (fibrillar) bovine collagen substrates were used as controls. The three EDCMs were similar among each other in terms of structure and mechanical performances and were much thinner and mechanically more resistant than the commercial membranes. Number of fibroblasts seeded on sea-urchin membranes were comparable to the bovine collagen substrates. Cell morphology on all EDCMs was similar to that of structurally comparable (reassembled) bovine collagen substrates. Overall, echinoderms, and sea urchins particularly, are alternative collagen sources to produce efficient GTR membranes. Sea urchins display a further advantage in terms of eco-sustainability by recycling tissues from food wastes.

Marine-derived collagen biomaterials from echinoderm connective tissues

KAUST Repository

Ferrario, Cinzia

2016-03-31

The use of marine collagens is a hot topic in the field of tissue engineering. Echinoderms possess unique connective tissues (Mutable Collagenous Tissues, MCTs) which can represent an innovative source of collagen to develop collagen barrier-membranes for Guided Tissue Regeneration (GTR). In the present work we used MCTs from different echinoderm models (sea urchin, starfish and sea cucumber) to produce echinoderm-derived collagen membranes (EDCMs). Commercial membranes for GTR or soluble/reassembled (fibrillar) bovine collagen substrates were used as controls. The three EDCMs were similar among each other in terms of structure and mechanical performances and were much thinner and mechanically more resistant than the commercial membranes. Number of fibroblasts seeded on sea-urchin membranes were comparable to the bovine collagen substrates. Cell morphology on all EDCMs was similar to that of structurally comparable (reassembled) bovine collagen substrates. Overall, echinoderms, and sea urchins particularly, are alternative collagen sources to produce efficient GTR membranes. Sea urchins display a further advantage in terms of eco-sustainability by recycling tissues from food wastes.

Connective matrix organization in human pulmonary fibrosis. Collagen polymorphism analysis in fibrotic deposits by immunohistological methods.

Science.gov (United States)

Takiya, C; Peyrol, S; Cordier, J F; Grimaud, J A

1983-01-01

In the interstitium of the alveolar septa in the peripheral parts of the lung, four molecular types of collagen (I, III, IV and V) each with different morphological appearances, can be identified. The structural integrity of collagens accounts for the physiological efficiency of the lung. Fibrous thickening of alveolar septa is an invariable result of various diseases affecting the interstitium of the lung. The light and electron microscopic findings, and the immunological typing of collagens in six cases of fibrotic alveolar disease, are described. In the alveolar septa, two different compartments (the alveolo-capillary junction and the supportive axis) were affected by fibrosis: the alveolo-capillary junction was widened by the addition of interstitial collagens to basement membranes. In the axis, the increase of interstitial (types I and III) collagen gave rise to different patterns of connective matrix organization, graded as Loose or Dense depending on quantitative alterations of the type I/III ratio. The mode of organization of the fibrotic lung connective matrix, which depends on the quality of deposits in the matrix, may be correlated with the evolution of interstitial pulmonary fibrosis, in terms of its stability, remodelling ability and reversibility.

Heparan sulfate proteoglycans made by different basement-membrane-producing tumors have immunological and structural similarities

DEFF Research Database (Denmark)

Wewer, U M; Albrechtsen, R; Hassell, J R

1985-01-01

in the native basement membrane of surrounding normal murine tissues. Blocking and ELISA assays demonstrated that the antibodies recognized both antigens. Using techniques involving the chemical and enzymatic degradation of 35S-sulfate-labeled glycosaminoglycans, the mouse EHS tumor cells were found to produce...... proteoglycans obtained from these two sources immunoprecipitated the same precursor protein with a molecular mass of 400,000 daltons from 35S-methionine pulse-labeled cells of both tumors. Immunohistochemistry showed the heparan sulfate proteoglycan to be distributed in the extracellular matrix and also...

Mutations in collagen 18A1 (COL18A1 and their relevance to the human phenotype

Directory of Open Access Journals (Sweden)

Passos-Bueno Maria Rita

2006-01-01

Full Text Available Collagen XVIII, a proteoglycan, is a component of basement membranes (BMs. There are three distinct isoforms that differ only by their N-terminal, but with a specific pattern of tissue and developmental expression. Cleavage of its C-terminal produces endostatin, an inhibitor of angiogenesis. In its N-terminal, there is a frizzled motif which seems to be involved in Wnt signaling. Mutations in this gene cause Knobloch syndrome KS, an autosomal recessive disorder characterized by vitreoretinal and macular degeneration and occipital encephalocele. This review discusses the effect of both rare and polymorphic alleles in the human phenotype, showing that deficiency of one of the collagen XVIII isoforms is sufficient to cause KS and that null alleles causing deficiency of all collagen XVIII isoforms are associated with a more severe ocular defect. This review besides illustrating the functional importance of collagen XVIII in eye development and its structure maintenance throughout life, it also shows its role in other tissues and organs, such as nervous system and kidney.

Frequently relapsing anti-glomerular basement membrane antibody disease with changing clinical phenotype and antibody characteristics over time

OpenAIRE

Gu, Bobby; Magil, Alex B.; Barbour, Sean J.

2016-01-01

Anti-glomerular basement membrane (GBM) antibody disease is a typically monophasic autoimmune disease with severe pulmonary and renal involvement. We report an atypical case of frequently relapsing anti-GBM antibody disease with both anti-GBM antibody?positive flares with pulmonary and renal involvement, and anti-GBM antibody?negative flares that were pulmonary limited with no histologic renal disease. This is the first report of alternating disease phenotype and anti-GBM antibody status over...

Mechanisms of Vascular Damage by Hemorrhagic Snake Venom Metalloproteinases: Tissue Distribution and In Situ Hydrolysis

Science.gov (United States)

Baldo, Cristiani; Jamora, Colin; Yamanouye, Norma; Zorn, Telma M.; Moura-da-Silva, Ana M.

2010-01-01

Background Envenoming by viper snakes constitutes an important public health problem in Brazil and other developing countries. Local hemorrhage is an important symptom of these accidents and is correlated with the action of snake venom metalloproteinases (SVMPs). The degradation of vascular basement membrane has been proposed as a key event for the capillary vessel disruption. However, SVMPs that present similar catalytic activity towards extracellular matrix proteins differ in their hemorrhagic activity, suggesting that other mechanisms might be contributing to the accumulation of SVMPs at the snakebite area allowing capillary disruption. Methodology/Principal Findings In this work, we compared the tissue distribution and degradation of extracellular matrix proteins induced by jararhagin (highly hemorrhagic SVMP) and BnP1 (weakly hemorrhagic SVMP) using the mouse skin as experimental model. Jararhagin induced strong hemorrhage accompanied by hydrolysis of collagen fibers in the hypodermis and a marked degradation of type IV collagen at the vascular basement membrane. In contrast, BnP1 induced only a mild hemorrhage and did not disrupt collagen fibers or type IV collagen. Injection of Alexa488-labeled jararhagin revealed fluorescent staining around capillary vessels and co-localization with basement membrane type IV collagen. The same distribution pattern was detected with jararhagin-C (disintegrin-like/cysteine-rich domains of jararhagin). In opposition, BnP1 did not accumulate in the tissues. Conclusions/Significance These results show a particular tissue distribution of hemorrhagic toxins accumulating at the basement membrane. This probably occurs through binding to collagens, which are drastically hydrolyzed at the sites of hemorrhagic lesions. Toxin accumulation near blood vessels explains enhanced catalysis of basement membrane components, resulting in the strong hemorrhagic activity of SVMPs. This is a novel mechanism that underlies the difference between

Laminin, a noncollagenous component of epithelial basement membranes synthesized by a rat yolk sac tumor

DEFF Research Database (Denmark)

Wewer, U; Albrechtsen, R; Ruoslahti, E

1981-01-01

Laminin, a glycoprotein antigenically similar or identical to a component of epithelial basement membranes, was identified as a major component of the abundant extracellular matrix synthesized by an experimentally induced rat yolk sac tumor. Immunocytochemical staining revealed laminin in cultured...... polypeptides with molecular weights of approximately 200,000 and 400,000. These comigrated with the polypeptides of mouse laminin isolated previously. The yolk sac tumor tissue grown in vivo contained laminin in the tumor cells and in the extracellular material as evidenced by immunofluorescence...... membranes in rat tissues in a manner indistinguishable from antilaminin. The presence of laminin in rat yolk sac cells, the presumed origin of our yolk sac tumor, was studied in some detail. Laminin was found to be present in normal cells of the visceral as well as the parietal yolk sac layer...

Graphene oxide improves the biocompatibility of collagen membranes in an in vitro model of human primary gingival fibroblasts.

Science.gov (United States)

De Marco, Patrizia; Zara, Susi; De Colli, Marianna; Radunovic, Milena; Lazović, Vladimir; Ettorre, Valeria; Di Crescenzo, Antonello; Piattelli, Adriano; Cataldi, Amelia; Fontana, Antonella

2017-09-13

Commercial collagen membranes are used in oral surgical procedures as scaffolds for bone deposition in guided bone regeneration. Here, we have enriched them with graphene oxide (GO) via a simple non-covalent functionalization, exploiting the capacity of oxygenated carbon functional moieties of GO to interact through hydrogen bonding with collagen. In the present paper, the GO-coated membranes have been characterized in terms of stability, nano-roughness, biocompatibility and induction of inflammatory response in human primary gingival fibroblast cells. The obtained coated membranes are demonstrated not to leak GO in the bulk solution, and to change some features of the membrane, such as stiffness and adhesion between the membrane and the atomic force microscopy (AFM) tip. Moreover, the presence of GO increases the roughness and the total surface exposed to the cells, as demonstrated by AFM analyses. The obtained material is biocompatible, and does not induce inflammation in the tested cells.

Disease: H01721 [KEGG MEDICUS

Lifescience Database Archive (English)

Full Text Available H01721 Anti-glomerular basement membrane (GBM) disease; Goodpasture syndrome Goodp...asture syndrome (GS), or anti-glomerular basement membrane (anti-GBM) disease, is a rare and organ-specific ...n of the alpha 3 chain of type IV collagen [alpha3(IV)NC1], found in the glomerular and alveolar basement me

In vivo turnover of the basement membrane and other heparan sulfate proteoglycans of rat glomerulus

International Nuclear Information System (INIS)

Beavan, L.A.; Davies, M.; Couchman, J.R.; Williams, M.A.; Mason, R.M.

1989-01-01

The metabolic turnover of rat glomerular proteoglycans in vivo was investigated. Newly synthesized proteoglycans were labeled during a 7-h period after injecting sodium [35S]sulfate intraperitoneally. At the end of the labeling period a chase dose of sodium sulfate was given. Subsequently at defined times (0-163 h) the kidneys were perfused in situ with 0.01% cetylpyridinium chloride in phosphate-buffered saline to maximize the recovery of 35S-proteoglycans. Glomeruli were isolated from the renal cortex and analyzed for 35S-proteoglycans by autoradiographic, biochemical, and immunochemical methods. Grain counting of autoradiographs revealed a complex turnover pattern of 35S-labeled macromolecules, commencing with a rapid phase followed by a slower phase. Biochemical analysis confirmed the biphasic pattern and showed that the total population of [35S]heparan sulfate proteoglycans had a metabolic half-life (t1/2) of 20 and 60 h in the early and late phases, respectively. Heparan sulfate proteoglycans accounted for 80% of total 35S-proteoglycans, the remainder being chondroitin/dermatan sulfate proteoglycans. Whole glomeruli were extracted with 4% 3-[(cholamidopropyl)dimethy-lammonio]-1-propanesulfonate-4 M guanidine hydrochloride, a procedure which solubilized greater than 95% of the 35S-labeled macromolecules. Of these 11-13% was immunoprecipitated by an antiserum against heparan sulfate proteoglycan which, in immunolocalization experiments, showed specificity for staining the basement membrane of rat glomeruli. Autoradiographic analysis showed that 18% of total radioactivity present at the end of the labeling period was associated with the glomerular basement membrane

Multiple recurrences of anti-glomerular basement membrane disease with variable antibody detection: can the laboratory be trusted?

OpenAIRE

Liu, Patricia; Waheed, Sana; Boujelbane, Lamya; Maursetter, Laura J.

2016-01-01

Anti-glomerular basement membrane (GBM) disease is commonly a monophasic illness. We present the case of multiple recurrences of anti-GBM disease with varying serum anti-GBM antibody findings. A 33-year-old female tobacco user presenting with hematuria was diagnosed with anti-GBM disease by renal biopsy. Five years later, she presented with alveolar hemorrhage and positive anti-GBM antibody. She presented a third time with alveolar hemorrhage but undetectable anti-GBM antibody. With each occu...

Collagen XII and XIV, New Partners of Cartilage Oligomeric Matrix Protein in the Skin Extracellular Matrix Suprastructure*

Science.gov (United States)

Agarwal, Pallavi; Zwolanek, Daniela; Keene, Douglas R.; Schulz, Jan-Niklas; Blumbach, Katrin; Heinegård, Dick; Zaucke, Frank; Paulsson, Mats; Krieg, Thomas; Koch, Manuel; Eckes, Beate

2012-01-01

The tensile and scaffolding properties of skin rely on the complex extracellular matrix (ECM) that surrounds cells, vasculature, nerves, and adnexus structures and supports the epidermis. In the skin, collagen I fibrils are the major structural component of the dermal ECM, decorated by proteoglycans and by fibril-associated collagens with interrupted triple helices such as collagens XII and XIV. Here we show that the cartilage oligomeric matrix protein (COMP), an abundant component of cartilage ECM, is expressed in healthy human skin. COMP expression is detected in the dermal compartment of skin and in cultured fibroblasts, whereas epidermis and HaCaT cells are negative. In addition to binding collagen I, COMP binds to collagens XII and XIV via their C-terminal collagenous domains. All three proteins codistribute in a characteristic narrow zone in the superficial papillary dermis of healthy human skin. Ultrastructural analysis by immunogold labeling confirmed colocalization and further revealed the presence of COMP along with collagens XII and XIV in anchoring plaques. On the basis of these observations, we postulate that COMP functions as an adapter protein in human skin, similar to its function in cartilage ECM, by organizing collagen I fibrils into a suprastructure, mainly in the vicinity of anchoring plaques that stabilize the cohesion between the upper dermis and the basement membrane zone. PMID:22573329

Collagen XII and XIV, new partners of cartilage oligomeric matrix protein in the skin extracellular matrix suprastructure.

Science.gov (United States)

Agarwal, Pallavi; Zwolanek, Daniela; Keene, Douglas R; Schulz, Jan-Niklas; Blumbach, Katrin; Heinegård, Dick; Zaucke, Frank; Paulsson, Mats; Krieg, Thomas; Koch, Manuel; Eckes, Beate

2012-06-29

The tensile and scaffolding properties of skin rely on the complex extracellular matrix (ECM) that surrounds cells, vasculature, nerves, and adnexus structures and supports the epidermis. In the skin, collagen I fibrils are the major structural component of the dermal ECM, decorated by proteoglycans and by fibril-associated collagens with interrupted triple helices such as collagens XII and XIV. Here we show that the cartilage oligomeric matrix protein (COMP), an abundant component of cartilage ECM, is expressed in healthy human skin. COMP expression is detected in the dermal compartment of skin and in cultured fibroblasts, whereas epidermis and HaCaT cells are negative. In addition to binding collagen I, COMP binds to collagens XII and XIV via their C-terminal collagenous domains. All three proteins codistribute in a characteristic narrow zone in the superficial papillary dermis of healthy human skin. Ultrastructural analysis by immunogold labeling confirmed colocalization and further revealed the presence of COMP along with collagens XII and XIV in anchoring plaques. On the basis of these observations, we postulate that COMP functions as an adapter protein in human skin, similar to its function in cartilage ECM, by organizing collagen I fibrils into a suprastructure, mainly in the vicinity of anchoring plaques that stabilize the cohesion between the upper dermis and the basement membrane zone.

Collagen-chitosan-glycerol bio-composite as artificial tympanic membrane for ruptured inner ear organ

Science.gov (United States)

Widiyanti, Prihartini; Setya Angtika, Rara; Githanadi, Brillyana; Hanif Kharisma, Ditya; Asyraf, Tarikh Omar; Wardani, Adita

2017-05-01

WHO data in 2012 shows that 5.3% of world population highly suffers from hearing loss and deafness. One of the deafness causes is rupture of tympanic membrane. Tympanic membrane damage which occurs often is perforated tympanic membrane, and it is also commonly known in medical term as tympanic membrane perforation. The causes, for instance, are high frequency of using earphones, traumatic accidents, noise, bacteria, viruses, and infectious microorganism. Tympanoplasty becomes the only treatment that can be widely accepted despite of deficiencies in postoperative complications. Therefore, this research aims to create artificial tympanic membrane made of natural materials such as type I collagen composited with chitosan and made of addition of glycerol to improve its mechanical strength and biodegradability. The method included the process of dissolving acetic acid in distilled water and mixation with chitosan. The solution is next added with glycerol and stirred to be homogeneous. After that, it was minted in petri dish and aerated before characterized. The sample characterization included tensile strength of which tensile test results showed that the value of the elasticity modulus tended to decrease with an increase in collagen concentration. The elasticity modulus values in a row for the variations of 7: 3, 8: 2, and 9: 1 were 35.10 MPa, 54,52MPa, and 47,45MPa respectively. The morphological test with 1000x, 2500x, and 5000x magnification showed their interaction in the formation of pores. Cytotoxicity results, moreover, showed that those samples were non-toxic and safe for the body due to the percentage of living cells. The sound absorption coefficient was between 1000 Hz - 2000 Hz which means that it could use as sound absorbing material. The antibacterial test results showed that all the sample variations were anti-bacterial due to the diameter of the clear zone. In conclusion, collagen and chitosan composite with addition of glycerol could be used for

PDGF-metronidazole-encapsulated nanofibrous functional layers on collagen membrane promote alveolar ridge regeneration

Directory of Open Access Journals (Sweden)

Ho MH

2017-08-01

Full Text Available Ming-Hua Ho,1 Hao-Chieh Chang,2,3 Yu-Chia Chang,3 Jeiannete Claudia,1 Tzu-Chiao Lin,2 Po-Chun Chang2,3 1Department of Chemical Engineering, College of Engineering, National Taiwan University of Science and Technology, Taipei, Taiwan; 2Graduate Institute of Clinical Dentistry, School of Dentistry, National Taiwan University, Taipei, Taiwan; 3Department of Dentistry, National Taiwan University Hospital, Taipei, Taiwan Abstract: This study aimed to develop a functionally graded membrane (FGM to prevent infection and promote tissue regeneration. Poly(L-lactide-co-D,L-lactide encapsulating platelet-derived growth factor (PDLLA-PDGF or metronidazole (PDLLA-MTZ was electrospun to form a nanofibrous layer on the inner or outer surface of a clinically available collagen membrane, respectively. The membrane was characterized for the morphology, molecule release profile, in vitro and in vivo biocompatibility, and preclinical efficiency for alveolar ridge regeneration. The PDLLA-MTZ and PDLLA-PDGF nanofibers were 800–900 nm in diameter, and the thicknesses of the functional layers were 20–30 µm, with sustained molecule release over 28 days. All of the membranes tested were compatible with cell survival in vitro and showed good tissue integration with minimal fibrous capsule formation or inflammation. Cell proliferation was especially prominent on the PDLLA-PDGF layer in vivo. On the alveolar ridge, all FGMs reduced wound dehiscence compared with the control collagen membrane, and the FGM with PDLLA-PDGF promoted osteogenesis significantly. In conclusion, the FGMs with PDLLA-PDGF and PDLLA-MTZ showed high biocompatibility and facilitated wound healing compared with conventional membrane, and the FGM with PDLLA-PDGF enhanced alveolar ridge regeneration in vivo. The design represents a beneficial modification, which may be easily adapted for future clinical use. Keywords: tissue engineering, platelet-derived growth factor, metronidazole, alveolar process

Modeling pulmonary fibrosis by abnormal expression of telomerase/apoptosis/collagen V in experimental usual interstitial pneumonia

International Nuclear Information System (INIS)

Parra, E.R.; Pincelli, M.S.; Teodoro, W.R.; Velosa, A.P.P.; Martins, V.; Rangel, M.P.; Barbas-Filho, J.V.; Capelozzi, V.L.

2014-01-01

Limitations on tissue proliferation capacity determined by telomerase/apoptosis balance have been implicated in pathogenesis of idiopathic pulmonary fibrosis. In addition, collagen V shows promise as an inductor of apoptosis. We evaluated the quantitative relationship between the telomerase/apoptosis index, collagen V synthesis, and epithelial/fibroblast replication in mice exposed to butylated hydroxytoluene (BHT) at high oxygen concentration. Two groups of mice were analyzed: 20 mice received BHT, and 10 control mice received corn oil. Telomerase expression, apoptosis, collagen I, III, and V fibers, and hydroxyproline were evaluated by immunohistochemistry, in situ detection of apoptosis, electron microscopy, immunofluorescence, and histomorphometry. Electron microscopy confirmed the presence of increased alveolar epithelial cells type 1 (AEC1) in apoptosis. Immunostaining showed increased nuclear expression of telomerase in AEC type 2 (AEC2) between normal and chronic scarring areas of usual interstitial pneumonia (UIP). Control lungs and normal areas from UIP lungs showed weak green birefringence of type I and III collagens in the alveolar wall and type V collagen in the basement membrane of alveolar capillaries. The increase in collagen V was greater than collagens I and III in scarring areas of UIP. A significant direct association was found between collagen V and AEC2 apoptosis. We concluded that telomerase, collagen V fiber density, and apoptosis evaluation in experimental UIP offers the potential to control reepithelization of alveolar septa and fibroblast proliferation. Strategies aimed at preventing high rates of collagen V synthesis, or local responses to high rates of cell apoptosis, may have a significant impact in pulmonary fibrosis

Modeling pulmonary fibrosis by abnormal expression of telomerase/apoptosis/collagen V in experimental usual interstitial pneumonia

Energy Technology Data Exchange (ETDEWEB)

Parra, E.R.; Pincelli, M.S. [Departamento de Patologia, Faculdade de Medicina, Universidade de São Paulo, São Paulo, SP (Brazil); Teodoro, W.R.; Velosa, A.P.P. [Disciplina de Reumatologia, Faculdade de Medicina, Universidade de São Paulo, São Paulo, SP (Brazil); Martins, V.; Rangel, M.P.; Barbas-Filho, J.V.; Capelozzi, V.L. [Departamento de Patologia, Faculdade de Medicina, Universidade de São Paulo, São Paulo, SP (Brazil)

2014-06-04

Limitations on tissue proliferation capacity determined by telomerase/apoptosis balance have been implicated in pathogenesis of idiopathic pulmonary fibrosis. In addition, collagen V shows promise as an inductor of apoptosis. We evaluated the quantitative relationship between the telomerase/apoptosis index, collagen V synthesis, and epithelial/fibroblast replication in mice exposed to butylated hydroxytoluene (BHT) at high oxygen concentration. Two groups of mice were analyzed: 20 mice received BHT, and 10 control mice received corn oil. Telomerase expression, apoptosis, collagen I, III, and V fibers, and hydroxyproline were evaluated by immunohistochemistry, in situ detection of apoptosis, electron microscopy, immunofluorescence, and histomorphometry. Electron microscopy confirmed the presence of increased alveolar epithelial cells type 1 (AEC1) in apoptosis. Immunostaining showed increased nuclear expression of telomerase in AEC type 2 (AEC2) between normal and chronic scarring areas of usual interstitial pneumonia (UIP). Control lungs and normal areas from UIP lungs showed weak green birefringence of type I and III collagens in the alveolar wall and type V collagen in the basement membrane of alveolar capillaries. The increase in collagen V was greater than collagens I and III in scarring areas of UIP. A significant direct association was found between collagen V and AEC2 apoptosis. We concluded that telomerase, collagen V fiber density, and apoptosis evaluation in experimental UIP offers the potential to control reepithelization of alveolar septa and fibroblast proliferation. Strategies aimed at preventing high rates of collagen V synthesis, or local responses to high rates of cell apoptosis, may have a significant impact in pulmonary fibrosis.

Laminin peptide YIGSR induces collagen synthesis in Hs27 human dermal fibroblasts

International Nuclear Information System (INIS)

Yoon, Jong Hyuk; Kim, Jaeyoon; Lee, Hyeongjoo; Kim, So Young; Jang, Hwan-Hee; Ryu, Sung Ho; Kim, Beom Joon; Lee, Taehoon G.

2012-01-01

Highlights: ► We identify a function of the YIGSR peptide to enhance collagen synthesis in Hs27. ► YIGSR peptide enhanced collagen type 1 synthesis both of gene and protein levels. ► There were no changes in cell proliferation and MMP-1 level in YIGSR treatment. ► The YIGSR effect on collagen synthesis mediated activation of FAK, pyk2 and ERK. ► The YIGSR-induced FAK and ERK activation was modulated by FAK and MEK inhibitors. -- Abstract: The dermal ECM is synthesized from fibroblasts and is primarily compromised of fibrillar collagen and elastic fibers, which support the mechanical strength and resiliency of skin, respectively. Laminin, a major glycoprotein located in the basement membrane, promotes cell adhesion, cell growth, differentiation, and migration. The laminin tyrosine-isoleucine-glycine-serine-arginine (YIGSR) peptide, corresponding to the 929–933 sequence of the β1 chain, is known to be a functional motif with effects on the inhibition of tumor metastasis, the regulation of sensory axonal response and the inhibition of angiogenesis through high affinity to the 67 kDa laminin receptor. In this study, we identified a novel function of the YIGSR peptide to enhance collagen synthesis in human dermal fibroblasts. To elucidate this novel function regarding collagen synthesis, we treated human dermal fibroblasts with YIGSR peptide in both a time- and dose-dependent manner. According to subsequent experiments, we found that the YIGSR peptide strongly enhanced collagen type 1 synthesis without changing cell proliferation or cellular MMP-1 level. This YIGSR peptide-mediated collagen type 1 synthesis was modulated by FAK inhibitor and MEK inhibitor. This study clearly reveals that YIGSR peptide plays a novel function on the collagen type 1 synthesis of dermal fibroblasts and also suggests that YIGSR is a strong candidate peptide for the treatment of skin aging and wrinkles.

Quantitative analysis of the synthesis and secretion of type VII collagen in cultured human dermal fibroblasts with a sensitive sandwich enzyme-linked immunoassay.

Science.gov (United States)

Amano, Satoshi; Ogura, Yuki; Akutsu, Nobuko; Nishiyama, Toshio

2007-02-01

Type VII collagen is the major component of anchoring fibrils in the epidermal basement membrane. Its expression has been analyzed by immunostaining or Northern blotting, but rarely at the protein level. In this study, we have quantitatively examined the effects of ascorbic acid and various cytokines/growth factors on the protein synthesis and secretion of type VII collagen by human dermal fibroblasts in culture, using a developed, highly sensitive sandwich enzyme-linked immunoassay with two kinds of specific monoclonal antibodies against the non-collagenous domain-1. Ascorbic acid and its derivative induced a twofold increase in type VII collagen synthesis, and markedly increased the secretion of type VII collagen into the medium when compared with the control culture. This effect was not influenced by the presence of transforming growth factor-beta1 (TGF-beta1). The synthesis of type VII collagen was elevated by TGF-beta1, platelet-derived growth factor, tumor necrosis factor-alpha, and interleukin-1beta, but not by TGF-alpha. Thus, our data indicate that the synthesis and secretion of type VII collagen in human dermal fibroblasts are regulated by ascorbate and the enhancement of type VII collagen gene expression by cytokines/growth factors is accompanied with elevated production of type VII collagen at the protein level.

Immunohistochemical localization of chondroitin sulfate, chondroitin sulfate proteoglycan, heparan sulfate proteoglycan, entactin, and laminin in basement membranes of postnatal developing and adult rat lungs

DEFF Research Database (Denmark)

Sannes, P L; Burch, K K; Khosla, J

1993-01-01

Histologic preparations of lungs from 1-, 5-, 10-, 18-, and 25-day-old postnatal and adult rats were examined immunohistochemically with antibodies specific against chondroitin sulfate (CS), basement membrane chondroitin sulfate proteoglycan (BM-CSPG), heparan sulfate proteoglycan (HSPG), entactin...

Guided Bone Regeneration in Long-Bone Defects with a Structural Hydroxyapatite Graft and Collagen Membrane

Science.gov (United States)

2013-01-01

Original Articles Guided Bone Regeneration in Long-Bone Defects with a Structural Hydroxyapatite Graft and Collagen Membrane Teja Guda, PhD,1,2 John...Joint Surg Br 90-B, 1617, 2008. 6. Carlo Reis, E.C., Borges AaPB, Araujo, M.V.F., Mendes, V.C., Guan, L., and Davies, J.E. Periodontal regeneration...Regeneration of periodontal tissues: combinations of barrier membranes and grafting materials–biological foundation and preclinical evi- dence: a

Podocytes regulate the glomerular basement membrane protein nephronectin by means of miR-378a-3p in glomerular diseases.

Science.gov (United States)

Müller-Deile, Janina; Dannenberg, Jan; Schroder, Patricia; Lin, Meei-Hua; Miner, Jeffrey H; Chen, Rongjun; Bräsen, Jan-Hinrich; Thum, Thomas; Nyström, Jenny; Staggs, Lynne Beverly; Haller, Hermann; Fiedler, Jan; Lorenzen, Johan M; Schiffer, Mario

2017-10-01

The pathophysiology of many proteinuric kidney diseases is poorly understood, and microRNAs (miRs) regulation of these diseases has been largely unexplored. Here, we tested whether miR-378a-3p is a novel regulator of glomerular diseases. MiR-378a-3p has two predicted targets relevant to glomerular function, the glomerular basement membrane matrix component, nephronectin (NPNT), and vascular endothelial growth factor VEGF-A. In zebrafish (Danio rerio), miR-378a-3p mimic injection or npnt knockdown by a morpholino oligomer caused an identical phenotype consisting of edema, proteinuria, podocyte effacement, and widening of the glomerular basement membrane in the lamina rara interna. Zebrafish vegf-A protein could not rescue this phenotype. However, mouse Npnt constructs containing a mutated 3'UTR region prevented the phenotype caused by miR-378a-3p mimic injection. Overexpression of miR-378a-3p in mice confirmed glomerular dysfunction in a mammalian model. Biopsies from patients with focal segmental glomerulosclerosis and membranous nephropathy had increased miR-378a-3p expression and reduced glomerular levels of NPNT. Thus, miR-378a-3p-mediated suppression of the glomerular matrix protein NPNT is a novel mechanism for proteinuria development in active glomerular diseases. Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

Multilayer Membranes of Glycosaminoglycans and Collagen I Biomaterials Modulate the Function and Microvesicle Release of Endothelial Progenitor Cells.

Science.gov (United States)

Dai, Bingyan; Pan, Qunwen; Li, Zhanghua; Zhao, Mingyan; Liao, Xiaorong; Wu, Keng; Ma, Xiaotang

2016-01-01

Multilayer composite membrane of biomaterials can increase the function of adipose stem cells or osteoprogenitor cells. Recent evidence indicates endothelial progenitor cells (EPCs) and EPCs released microvesicles (MVs) play important roles in angiogenesis and vascular repair. Here, we investigated the effects of biomaterial multilayer membranes of hyaluronic acid (HA) or chondroitin sulfate (CS) and Collagen I (Col I) on the functions and MVs release of EPCs. Layer-by-layer (LBL) technology was applied to construct the multilayer composite membranes. Four types of the membranes constructed by adsorbing either HA or CS and Col I alternatively with different top layers were studied. The results showed that all four types of multilayer composite membranes could promote EPCs proliferation and migration and inhibit cell senility, apoptosis, and the expression of activated caspase-3. Interestingly, these biomaterials increased the release and the miR-126 level of EPCs-MVs. Moreover, the CS-Col I membrane with CS on the top layer showed the most effects on promoting EPCs proliferation, EPCs-MV release, and miR-126 level in EPCs-MVs. In conclusion, HA/CS and Collagen I composed multilayer composite membranes can promote EPCs functions and release of miR-126 riched EPCs-MVs, which provides a novel strategy for tissue repair treatment.

Bone Healing in Extraction Sockets Covered With Collagen Membrane Alone or Associated With Porcine-Derived Bone Graft: a Comparative Histological and Histomorphometric Analysis.

Science.gov (United States)

Guarnieri, Renzo; Testarelli, Luca; Stefanelli, Luigi; De Angelis, Francesca; Mencio, Francesca; Pompa, Giorgio; Di Carlo, Stefano

2017-01-01

The present paper reports data of a randomized study aimed to analyse and compare the histologic and histomorphometric aspects of bone healing in extraction sites covered with collagen membrane alone or associated with porcine-derived bone graft. Thirty patients, with single extraction sockets without severe bone wall defects in the premolar/molar region, were included. Ten extraction sockets were grafted with porcine-derived bone and covered with collagen membrane (group 1), 10 sites were covered with collagen membrane alone (group 2), and 10 sites healed spontaneously (group 3). After 4 months of healing, 26 (8 in group 1, 9 in group 2, and 9 in group 3) bone core specimens were harvested for histologic evaluation, then dental implants were placed. Sites in the group 1 and in the group 2 showed similar histologic and histomorphometric results without significantly differences in the percentage of vital bone (57.43% [SD 4.8] vs. 60.01% [SD 3.2]), and non-mineralized connective tissue 22.99% (SD 5.3) vs. 18.53% (SD 6.2). In group 1 a 16.57% (SD 3.8) of residual material was found. Results showed that the use of collagen membrane alone or associated to porcine-derived bone improves the healing bone process compared to that of extraction sites spontaneously healed. Moreover, histomorphometric data related to bone quality, indicated that extraction sites without severe walls defects and with a vestibular bone thickness > 1.5 mm, treated with a low resorbtion rate collagen membrane alone, do not need more than 4 months for dental implant insertion.

Mac-2 binding protein is a cell-adhesive protein of the extracellular matrix which self-assembles into ring-like structures and binds beta1 integrins, collagens and fibronectin

DEFF Research Database (Denmark)

Sasaki, T; Brakebusch, C; Engel, J

1998-01-01

Human Mac-2 binding protein (M2BP) was prepared in recombinant form from the culture medium of 293 kidney cells and consisted of a 92 kDa subunit. The protein was obtained in a native state as indicated by CD spectroscopy, demonstrating alpha-helical and beta-type structure, and by protease resis...... in the extracellular matrix of several mouse tissues....... in solid-phase assays to collagens IV, V and VI, fibronectin and nidogen, but not to fibrillar collagens I and III or other basement membrane proteins. The protein also mediated adhesion of cell lines at comparable strength with laminin. Adhesion to M2BP was inhibited by antibodies to integrin beta1...

Toward guided tissue and bone regeneration: morphology, attachment, proliferation, and migration of cells cultured on collagen barrier membranes. A systematic review.

NARCIS (Netherlands)

Behring, J.; Junker, R.; Walboomers, X.F.; Chessnut, B.; Jansen, J.A.

2008-01-01

Collagen barrier membranes are frequently used in both guided tissue regeneration (GTR) and guided bone regeneration (GBR). Collagen used for these devices is available from different species and is often processed to alter the properties of the final product. This is necessary because unprocessed

Comparing The Efficacy of Hematoxylin and Eosin, Periodic Acid Schiff and Fluorescent Periodic Acid Schiff-Acriflavine Techniques for Demonstration of Basement Membrane in Oral Lichen Planus: A Histochemical Study.

Science.gov (United States)

Pujar, Ashwini; Pereira, Treville; Tamgadge, Avinash; Bhalerao, Sudhir; Tamgadge, Sandhya

2015-01-01

Basement membrane (BM) is a thick sheet of extracellular matrix molecules, upon which epithelial cells attach. Various immunohistochemical studies in the past have been carried out but these advanced staining techniques are expensive and not feasible in routine laboratories. Although hematoxylin and eosin (H-E) is very popular among pathologists for looking at biopsies, the method has some limitations. This is where special stains come handy. The aim of the present study was to demonstrate and compare the efficacy of H-E, periodic acid Schiff (PAS) and fluorescent periodic acid-acriflavine staining techniques for the basement membrane and to establish a histochemical stain which could be cost effective, less time consuming, and unambiguous for observation of the basement membrane zone. A total number of 40 paraffin-embedded tissue sections of known basement membrane containing tissues including 10 - Normal oral mucosa (NOM) and 30 - oral lichen planus (OLP) were considered in the study. Four-micron-thick sections of each block were cut and stained with H-E stain, PAS and fluorescent periodic acid-acriflavine stain. Sections were evaluated by three oral pathologists independently for continuity, contrast and pattern. Though all the three stains showed favorable features at different levels, acriflavine stain was better than the other stains in demonstrating BM continuity, contrast and also the pattern followed by PAS stain. Acriflavine stain was the better in demonstrating a fibrillar pattern of a BM. Acriflavine stains a BM distinctly and is less time consuming and easy to carry out using readily available dyes as compared to other stains. The continuity and contrast along with the homogenous pattern and the afibrillar pattern of the BM was better demonstrated by acriflavine followed by the PAS stain.

Comparing the efficacy of hematoxylin and eosin, periodic acid schiff and fluorescent periodic acid schiff-acriflavine techniques for demonstration of basement membrane in oral lichen planus: A histochemical study

Directory of Open Access Journals (Sweden)

Ashwini Pujar

2015-01-01

Full Text Available Background: Basement membrane (BM is a thick sheet of extracellular matrix molecules, upon which epithelial cells attach. Various immunohistochemical studies in the past have been carried out but these advanced staining techniques are expensive and not feasible in routine laboratories. Although hematoxylin and eosin (H-E is very popular among pathologists for looking at biopsies, the method has some limitations. This is where special stains come handy. Aims and Objectives: The aim of the present study was to demonstrate and compare the efficacy of H-E, periodic acid Schiff (PAS and fluorescent periodic acid-acriflavine staining techniques for the basement membrane and to establish a histochemical stain which could be cost effective, less time consuming, and unambiguous for observation of the basement membrane zone. Materials and Methods: A total number of 40 paraffin-embedded tissue sections of known basement membrane containing tissues including 10 - Normal oral mucosa (NOM and 30 - oral lichen planus (OLP were considered in the study. Four-micron-thick sections of each block were cut and stained with H-E stain, PAS and fluorescent periodic acid-acriflavine stain. Sections were evaluated by three oral pathologists independently for continuity, contrast and pattern. Results: Though all the three stains showed favorable features at different levels, acriflavine stain was better than the other stains in demonstrating BM continuity, contrast and also the pattern followed by PAS stain. Acriflavine stain was the better in demonstrating a fibrillar pattern of a BM. Acriflavine stains a BM distinctly and is less time consuming and easy to carry out using readily available dyes as compared to other stains. Conclusion: The continuity and contrast along with the homogenous pattern and the afibrillar pattern of the BM was better demonstrated by acriflavine followed by the PAS stain.

Deletion of PPAR-γ in immune cells enhances susceptibility to antiglomerular basement membrane disease

Directory of Open Access Journals (Sweden)

Cristen Chafin

2010-10-01

Full Text Available Cristen Chafin2, Sarah Muse2, Raquel Hontecillas5, Josep Bassaganya-Riera5, David L Caudell2, Samuel K Shimp III4, M Nichole Rylander4, John Zhang6, Liwu Li3, Christopher M Reilly1,21Virginia College of Osteopathic Medicine, 2Department of Biomedical Sciences and Pathobiology, Virginia-Maryland Regional College of Veterinary Medicine, Virginia Polytechnic Institute and State University, Blacksburg, VA, USA; 3Department of Biological Sciences, 4Department of Mechanical Engineering, Virginia Polytechnic Institute and State University, Blacksburg, VA, USA; 5Nutritional Immunology and Molecular Medicine Laboratory, Virginia Bioinformatics Institute, Virginia Polytechnic Institute and State University, Blacksburg, VA, USA; 6Medical University of SC, Charleston, SC, USAAbstract: Activation of the nuclear hormone receptor peroxisome proliferator-activated receptor gamma (PPAR-γ has been shown to be immunoregulatory in autoimmune diseases by inhibiting production of a number of inflammatory mediators. We investigated whether PPAR-γ gene deletion in hematopoietic cells would alter disease pathogenesis in the antiglomerular basement membrane (anti-GBM mouse model. PPAR-γ+/+ and PPAR-γ-/- mice were immunized with rabbit antimouse GBM antibodies and lipopolysaccharide and evaluated for two weeks. Although both the PPAR-γ+/+ and PPAR-γ-/- mice had IgG deposition in the glomerulus and showed proteinuria two weeks after injection, glomerular and tubulointerstitial disease in PPAR-γ-/- mice were significantly more severe compared with the PPAR-γ+/+ animals. We observed that the PPAR-γ-/- mice had decreased CD4+CD25+ regulatory T cells and an increased CD8+:CD4+ ratio as compared with the PPAR-γ+/+ mice, suggesting that PPAR-γ has a role in the regulation of T cells. Furthermore, plasma interleukin-6 levels were significantly increased in the PPAR-γ-/- mice at two weeks as compared with the PPAR-γ+/+ animals. Taken together, these studies show that

Quantitative Assessment of Ultrastructure and Light Scatter in Mouse Corneal Debridement Wounds

Science.gov (United States)

Boote, Craig; Du, Yiqin; Morgan, Sian; Harris, Jonathan; Kamma-Lorger, Christina S.; Hayes, Sally; Lathrop, Kira L.; Roh, Danny S.; Burrow, Michael K.; Hiller, Jennifer; Terrill, Nicholas J.; Funderburgh, James L.; Meek, Keith M.

2012-01-01

Purpose. The mouse has become an important wound healing model with which to study corneal fibrosis, a frequent complication of refractive surgery. The aim of the current study was to quantify changes in stromal ultrastructure and light scatter that characterize fibrosis in mouse corneal debridement wounds. Methods. Epithelial debridement wounds, with and without removal of basement membrane, were produced in C57BL/6 mice. Corneal opacity was measured using optical coherence tomography, and collagen diameter and matrix order were quantified by x-ray scattering. Electron microscopy was used to visualize proteoglycans. Quantitative PCR (Q-PCR) measured mRNA transcript levels for several quiescent and fibrotic markers. Results. Epithelial debridement without basement membrane disruption produced a significant increase in matrix disorder at 8 weeks, but minimal corneal opacity. In contrast, basement membrane penetration led to increases in light scatter, matrix disorder, and collagen diameter, accompanied by the appearance of abnormally large proteoglycans in the subepithelial stroma. This group also demonstrated upregulation of several quiescent and fibrotic markers 2 to 4 weeks after wounding. Conclusions. Fibrotic corneal wound healing in mice involves extensive changes to collagen and proteoglycan ultrastructure, consistent with deposition of opaque scar tissue. Epithelial basement membrane penetration is a deciding factor determining the degree of ultrastructural changes and resulting opacity. PMID:22467580

Functional differentiation and alveolar morphogenesis of primary mammary cultures on reconstituted basement membrane

Energy Technology Data Exchange (ETDEWEB)

BARCELLOS-HOFF, M. H; AGGELER, J.; RAM, T. G; BISSELL, M. J

1989-02-01

An essential feature of mammary gland differentiation during pregnancy is the formation of alveoli composed of polarized epithelial cells, which, under the influence of lactogenic hormones, secrete vectorially and sequester milk proteins. Previous culture studies have described either organization of cells polarized towards lumina containing little or no demonstrable tissue-specific protein, or establishment of functional secretory cells exhibiting little or no glandular architecture. In this paper, we report that tissue-specific vectorial secretion coincides with the formation of functional alveoli-like structures by primary mammary epithelial cells cultured on a reconstituted basement membrane matrix (derived from Engelbreth-Holm-Swarm murine tumour). Morphogenesis of these unique three-dimensional structures was initiated by cell-directed remodelling of the exogenous matrix leading to reorganization of cells into matrixensheathed aggregates by 24 h after plating. The aggregates subsequently cavitated, so that by day 6 the cells were organized into hollow spheres in which apical cell surfaces faced lumina sealed by tight junctions and basal surfaces were surrounded by a distinct basal lamina. The profiles of proteins secreted into the apical (luminal) and basal (medium) compartments indicated that these alveoli-like structures were capable of an appreciable amount of vectorial secretion. Immunoprecipitation with a broad spectrum milk antiserum showed that more than 80% of caseins were secreted into the lumina, whereas iron-binding proteins (both lactoferrin and transferrin) were present in comparable amounts in each compartment. Thus, these mammary cells established protein targeting pathways directing milk-specific proteins to the luminal compartment. A time course monitoring secretory activity demonstrated that establishment of tissue-specific vectorial secretion and increased total and milk protein secretion coincided with functional alveolar

Type IV collagen is a novel DEJ biomarker that is reduced by radiotherapy.

Science.gov (United States)

McGuire, J D; Gorski, J P; Dusevich, V; Wang, Y; Walker, M P

2014-10-01

The dental basement membrane (BM) is composed of collagen types IV, VI, VII, and XVII, fibronectin, and laminin and plays an inductive role in epithelial-mesenchymal interactions during tooth development. The BM is degraded and removed during later-stage tooth morphogenesis; however, its original position defines the location of the dentin-enamel junction (DEJ) in mature teeth. We recently demonstrated that type VII collagen is a novel component of the inner enamel organic matrix layer contiguous with the DEJ. Since it is frequently co-expressed with and forms functional complexes with type VII collagen, we hypothesized that type IV collagen should also be localized to the DEJ in mature human teeth. To identify collagen IV, we first evaluated defect-free erupted teeth from various donors. To investigate a possible stabilizing role, we also evaluated extracted teeth exposed to high-dose radiotherapy--teeth that manifest post-radiotherapy DEJ instability. We now show that type IV collagen is a component within the morphological DEJ of posterior and anterior teeth from individuals aged 18 to 80 yr. Confocal microscopy revealed that immunostained type IV collagen was restricted to the 5- to 10-µm-wide optical DEJ, while collagenase treatment or previous in vivo tooth-level exposure to > 60 Gray irradiation severely reduced immunoreactivity. This assignment was confirmed by Western blotting with whole-tooth crown and enamel extracts. Without reduction, type IV collagen contained macromolecular α-chains of 225 and 250 kDa. Compositionally, our results identify type IV collagen as the first macromolecular biomarker of the morphological DEJ of mature teeth. Given its network structure and propensity to stabilize the dermal-epidermal junction, we propose that a collagen-IV-enriched DEJ may, in part, explain its well-known fracture toughness, crack propagation resistance, and stability. In contrast, loss of type IV collagen may represent a biochemical rationale for the DEJ

Two-layer membranes of calcium phosphate/collagen/PLGA nanofibres: in vitro biomineralisation and osteogenic differentiation of human mesenchymal stem cells

Science.gov (United States)

Hild, Nora; Schneider, Oliver D.; Mohn, Dirk; Luechinger, Norman A.; Koehler, Fabian M.; Hofmann, Sandra; Vetsch, Jolanda R.; Thimm, Benjamin W.; Müller, Ralph; Stark, Wendelin J.

2011-02-01

The present study evaluates the in vitro biomedical performance of an electrospun, flexible, anisotropic bilayer with one layer containing a collagen to mineral ratio similar to that in bone. The double membrane consists of a poly(lactide-co-glycolide) (PLGA) layer and an amorphous calcium phosphate (a-CaP)/collagen (Col)/PLGA layer. In vitro biomineralisation and a cell culture study with human mesenchymal stem cells (hMSC) were conducted to characterise such membranes for possible application as biomaterials. Nanofibres with different a-CaP/Col/PLGA compositions were synthesised by electrospinning to mimic the actual composition of bone tissue. Immersion in simulated body fluid and in cell culture medium resulted in the deposition of a hydroxyapatite layer. Incubation of hMSC for 4 weeks allowed for assessment of the proliferation and osteogenic differentiation of the cells on both sides of the double membrane. Confocal laser scanning microscopy was used to observe the proper adhesion of the cells. Calcium and collagen content was proven by Alizarin red S and Sirius red assays. Acute cytotoxic effects of the nanoparticles or the chemicals used in the scaffold preparation could be excluded based on viability assays (alamarBlue and alkaline phosphatase activity). The findings suggest possible application of such double membranes is in treatment of bone defects with complex geometries as wound dressing material.The present study evaluates the in vitro biomedical performance of an electrospun, flexible, anisotropic bilayer with one layer containing a collagen to mineral ratio similar to that in bone. The double membrane consists of a poly(lactide-co-glycolide) (PLGA) layer and an amorphous calcium phosphate (a-CaP)/collagen (Col)/PLGA layer. In vitro biomineralisation and a cell culture study with human mesenchymal stem cells (hMSC) were conducted to characterise such membranes for possible application as biomaterials. Nanofibres with different a

The vascular basement membrane as "soil" in brain metastasis.

Directory of Open Access Journals (Sweden)

W Shawn Carbonell

2009-06-01

Full Text Available Brain-specific homing and direct interactions with the neural substance are prominent hypotheses for brain metastasis formation and a modern manifestation of Paget's "seed and soil" concept. However, there is little direct evidence for this "neurotropic" growth in vivo. In contrast, many experimental studies have anecdotally noted the propensity of metastatic cells to grow along the exterior of pre-existing vessels of the CNS, a process termed vascular cooption. These observations suggest the "soil" for malignant cells in the CNS may well be vascular, rather than neuronal. We used in vivo experimental models of brain metastasis and analysis of human clinical specimens to test this hypothesis. Indeed, over 95% of early micrometastases examined demonstrated vascular cooption with little evidence for isolated neurotropic growth. This vessel interaction was adhesive in nature implicating the vascular basement membrane (VBM as the active substrate for tumor cell growth in the brain. Accordingly, VBM promoted adhesion and invasion of malignant cells and was sufficient for tumor growth prior to any evidence of angiogenesis. Blockade or loss of the beta1 integrin subunit in tumor cells prevented adhesion to VBM and attenuated metastasis establishment and growth in vivo. Our data establishes a new understanding of CNS metastasis formation and identifies the neurovasculature as the critical partner for such growth. Further, we have elucidated the mechanism of vascular cooption for the first time. These findings may help inform the design of effective molecular therapies for patients with fatal CNS malignancies.

Membrane-associated 41-kDa GTP-binding protein in collagen-induced platelet activation

International Nuclear Information System (INIS)

Walker, G.; Bourguignon, L.Y.

1990-01-01

Initially we established that the binding of collagen to human blood platelets stimulates both the rapid loss of PIP2 and the generation of inositol-4,5-bisphosphate (IP2) and inositol-1,4,5-triphosphate (IP3). These results indicate that the binding of collagen stimulates inositol phospholipid-specific phospholipase C during platelet activation. The fact that GTP or GTP-gamma-S augments, and pertussis toxin inhibits, collagen-induced IP3 formation suggests that a GTP-binding protein or (or proteins) may be directly involved in the regulation of phospholipase C-mediated phosphoinositide turnover in human platelets. We have used several complementary techniques to isolate and characterize a platelet 41-kDa polypeptide (or polypeptides) that has a number of structural and functional similarities to the regulatory alpha i subunit of the GTP-binding proteins isolated from bovine brain. This 41-kDa polypeptide (or polypeptides) is found to be closely associated with at least four membrane glycoproteins (e.g., gp180, gp110, gp95, and gp75) in a 330-kDa complex that can be dissociated by treatment with high salt plus urea. Most important, we have demonstrated that antilymphoma 41-kDa (alpha i subunit of GTP-binding proteins) antibody cross-reacts with the platelet 41-kDa protein (or proteins) and the alpha i subunit of bovine brain Gi alpha proteins, and blocks GTP/collagen-induced IP3 formation. These data provide strong evidence that the 41-kDa platelet GTP-binding protein (or proteins) is directly involved in collagen-induced signal transduction during platelet activation

Membrane-associated 41-kDa GTP-binding protein in collagen-induced platelet activation

Energy Technology Data Exchange (ETDEWEB)

Walker, G.; Bourguignon, L.Y. (Univ. of Miami Medical School, FL (USA))

1990-08-01

Initially we established that the binding of collagen to human blood platelets stimulates both the rapid loss of PIP2 and the generation of inositol-4,5-bisphosphate (IP2) and inositol-1,4,5-triphosphate (IP3). These results indicate that the binding of collagen stimulates inositol phospholipid-specific phospholipase C during platelet activation. The fact that GTP or GTP-gamma-S augments, and pertussis toxin inhibits, collagen-induced IP3 formation suggests that a GTP-binding protein or (or proteins) may be directly involved in the regulation of phospholipase C-mediated phosphoinositide turnover in human platelets. We have used several complementary techniques to isolate and characterize a platelet 41-kDa polypeptide (or polypeptides) that has a number of structural and functional similarities to the regulatory alpha i subunit of the GTP-binding proteins isolated from bovine brain. This 41-kDa polypeptide (or polypeptides) is found to be closely associated with at least four membrane glycoproteins (e.g., gp180, gp110, gp95, and gp75) in a 330-kDa complex that can be dissociated by treatment with high salt plus urea. Most important, we have demonstrated that antilymphoma 41-kDa (alpha i subunit of GTP-binding proteins) antibody cross-reacts with the platelet 41-kDa protein (or proteins) and the alpha i subunit of bovine brain Gi alpha proteins, and blocks GTP/collagen-induced IP3 formation. These data provide strong evidence that the 41-kDa platelet GTP-binding protein (or proteins) is directly involved in collagen-induced signal transduction during platelet activation.

Laminin peptide YIGSR induces collagen synthesis in Hs27 human dermal fibroblasts

Energy Technology Data Exchange (ETDEWEB)

Yoon, Jong Hyuk; Kim, Jaeyoon; Lee, Hyeongjoo [NovaCell Technology Inc., Pohang, Kyungbuk 790-784 (Korea, Republic of); Kim, So Young [Department of Dermatology, Chung-Ang University College of Medicine, Seoul 156-756 (Korea, Republic of); Department of Convergence Medicine and Pharmaceutical Biosciences, Graduate School, Chung-Ang University, Seoul 156-756 (Korea, Republic of); Jang, Hwan-Hee [Functional Food and Nutrition Division, Department of Agrofood Resources, Rural Development Administration, Suwon 441-853 (Korea, Republic of); Ryu, Sung Ho [Division of Integrative Biosciences and Biotechnology, Pohang University of Science and Technology (POSTECH), Pohang, Kyungbuk 790-784 (Korea, Republic of); Kim, Beom Joon [Department of Dermatology, Chung-Ang University College of Medicine, Seoul 156-756 (Korea, Republic of); Department of Convergence Medicine and Pharmaceutical Biosciences, Graduate School, Chung-Ang University, Seoul 156-756 (Korea, Republic of); Lee, Taehoon G., E-mail: taehoon@novacelltech.com [NovaCell Technology Inc., Pohang, Kyungbuk 790-784 (Korea, Republic of)

2012-11-23

Highlights: Black-Right-Pointing-Pointer We identify a function of the YIGSR peptide to enhance collagen synthesis in Hs27. Black-Right-Pointing-Pointer YIGSR peptide enhanced collagen type 1 synthesis both of gene and protein levels. Black-Right-Pointing-Pointer There were no changes in cell proliferation and MMP-1 level in YIGSR treatment. Black-Right-Pointing-Pointer The YIGSR effect on collagen synthesis mediated activation of FAK, pyk2 and ERK. Black-Right-Pointing-Pointer The YIGSR-induced FAK and ERK activation was modulated by FAK and MEK inhibitors. -- Abstract: The dermal ECM is synthesized from fibroblasts and is primarily compromised of fibrillar collagen and elastic fibers, which support the mechanical strength and resiliency of skin, respectively. Laminin, a major glycoprotein located in the basement membrane, promotes cell adhesion, cell growth, differentiation, and migration. The laminin tyrosine-isoleucine-glycine-serine-arginine (YIGSR) peptide, corresponding to the 929-933 sequence of the {beta}1 chain, is known to be a functional motif with effects on the inhibition of tumor metastasis, the regulation of sensory axonal response and the inhibition of angiogenesis through high affinity to the 67 kDa laminin receptor. In this study, we identified a novel function of the YIGSR peptide to enhance collagen synthesis in human dermal fibroblasts. To elucidate this novel function regarding collagen synthesis, we treated human dermal fibroblasts with YIGSR peptide in both a time- and dose-dependent manner. According to subsequent experiments, we found that the YIGSR peptide strongly enhanced collagen type 1 synthesis without changing cell proliferation or cellular MMP-1 level. This YIGSR peptide-mediated collagen type 1 synthesis was modulated by FAK inhibitor and MEK inhibitor. This study clearly reveals that YIGSR peptide plays a novel function on the collagen type 1 synthesis of dermal fibroblasts and also suggests that YIGSR is a strong candidate

UPR transducer BBF2H7 allows export of type II collagen in a cargo- and developmental stage–specific manner

Science.gov (United States)

Toyama, Takuya; Nakamura, Yuki; Tamada, Kentaro; Shimizu, Hitomi; Ninagawa, Satoshi; Okada, Tetsuya; Ishikawa-Fujiwara, Tomoko; Aoyama, Eriko; Takigawa, Masaharu

2017-01-01

The unfolded protein response (UPR) handles unfolded/misfolded proteins accumulated in the endoplasmic reticulum (ER). However, it is unclear how vertebrates correctly use the total of ten UPR transducers. We have found that ER stress occurs physiologically during early embryonic development in medaka fish and that the smooth alignment of notochord cells requires ATF6 as a UPR transducer, which induces ER chaperones for folding of type VIII (short-chain) collagen. After secretion of hedgehog for tissue patterning, notochord cells differentiate into sheath cells, which synthesize type II collagen. In this study, we show that this vacuolization step requires both ATF6 and BBF2H7 as UPR transducers and that BBF2H7 regulates a complete set of genes (Sec23/24/13/31, Tango1, Sedlin, and KLHL12) essential for the enlargement of COPII vesicles to accommodate long-chain collagen for export, leading to the formation of the perinotochordal basement membrane. Thus, the most appropriate UPR transducer is activated to cope with the differing physiological ER stresses of different content types depending on developmental stage. PMID:28500182

Clinical and radiographical evaluation of a bioresorbable collagen membrane of fish origin in the treatment of periodontal intrabony defects: A preliminary study.

Science.gov (United States)

Santosh Kumar, B B; Aruna, D R; Gowda, Vinayak S; Galagali, Sushama R; Prashanthy, R; Navaneetha, H

2013-09-01

Recently, there has been interest in non-mammalian collagen sources such as fish collagen in periodontal regeneration. In the present study, collagen barrier membrane of fish origin was assessed in the treatment of periodontal intrabony defects. Ten systemically healthy chronic periodontitis patients having a paired osseous defect in the mandibular posterior teeth were selected and randomly assigned to receive a collagen membrane (test) or open flap debridement (control) in a split mouth design. Clinical parameters such as Plaque index, Gingival bleeding index, Probing pocket depth, Relative attachment level, and Recession were recorded at baseline, 3, 6, and at 9 months, while radiographic evaluation was done to assess alveolar crestal bone level and percentage of defect fill at 6 and 9 months using autoCAD 2007 software. Student's t test (two-tailed, dependent) was used to find the significance of study parameters on continuous scale. Significance was set at 5% level of significance. Wilcoxon signed rank test was used to find the significance of percentage change of defect fill. The comparison between the two groups did not show any statistically significant differences in the parameters assessed (P > 0.05) but, within each group, clinical parameters showed statistically significant differences from baseline to 9 months (P < 0.05). Within the limits of the study, it can be inferred that no significant differences were found either by using collagen membrane of fish origin or open flap debridement in the treatment of periodontal intrabony defects.

Mutation in the alpha 5(IV) collagen chain in juvenile-onset Alport syndrome without hearing loss or ocular lesions

DEFF Research Database (Denmark)

Zhou, J; Hertz, Jens Michael; Tryggvason, K

1992-01-01

A single base mutation was identified in the type IV collagen alpha 5 chain gene (COL4A5) of a Danish kindred with Alport syndrome. The 27-year-old male proband developed hematuria in childhood and terminal renal failure at the age of 25 years. He has no hearing loss or ocular lesions. Electron...... microscopy demonstrated splitting of the lamina densa of the glomerular basement membrane. The proband's mother has had persistent microscopic hematuria since the age of 40 years, but no other manifestations. Southern analysis of MspI-digested genomic DNA from the proband showed the absence of 1.3-kb and 0...

Ridge Preservation Comparing a Nonresorbable PTFE Membrane to a Resorbable Collagen Membrane: A Clinical and Histologic Study in Humans.

Science.gov (United States)

Arbab, Hussain; Greenwell, Henry; Hill, Margaret; Morton, Dean; Vidal, Ricardo; Shumway, Brian; Allan, Nicholas D

2016-02-01

The primary aim of this randomized, controlled, blinded clinical trial was to compare the effect of a resorbable collagen membrane (CM group) versus a nonresorbable high-density polytetrafluoroethylene membrane (PTFE group) on the clinical and histologic outcomes of a ridge preservation procedure. All 24 sites received an intrasocket cancellous allograft and a buccal overlay bovine derived xenograft. The change in horizontal crestal ridge width was -1.4 ± 1.2 mm for the CM group, whereas the PTFE group lost -2.2 ± 1.5 mm, which was not statistically significant between groups (P > 0.05). Vertical ridge height change was -1.2 ± 1.5 for the CM group, whereas the PTFE group lost -0.5 ± 1.6, which was not significantly different between groups (P > 0.05). The percent vital bone was similar and not significantly different between groups. Primary closure was not obtained and the exposed membrane portion over the socket opening healed with keratinized tissue. The choice of a resorbable versus a nonresorbable barrier membrane did not affect the clinical or the histologic outcome of ridge preservation treatment.

Disease: H00582 [KEGG MEDICUS

Lifescience Database Archive (English)

Full Text Available H00582 Benign familial hematuria; Thin basement membrane nephropathy Benign famili...hildhood. The glomerular basement membrane is uniformly thin, but renal function is normal. Heterozygous mut...ations in COL4A3 or COL4A4 lead to reduced collagen network levels in the basemen...MIM: 141200 PMID:18094725 (gene, description) ... AUTHORS ... Gubler MC ... TITLE ... Inherited diseases of the glomerular basemen...Wang YY ... TITLE ... Thin basement membrane nephropathy. ... JOURNAL ... Kidney Int 64:1169-78 (2003) DOI:10.1046/j.1523-1755.2003.00234.x

19-DEJ-1, a hemidesmosome-anchoring filament complex-associated monoclonal antibody. Definition of a new skin basement membrane antigenic defect in junctional and dystrophic epidermolysis bullosa

DEFF Research Database (Denmark)

Fine, J D; Horiguchi, Y; Couchman, J R

1989-01-01

A murine monoclonal antibody (19-DEJ-1) was recently produced that recognizes a unique antigenic epitope of human skin basement membrane localized to the midlamina lucida exclusively in those areas bordered by overlying hemidesmosomes. To determine whether the antigen defined by 19-DEJ-1 is norma...

The microenvironment determines the breast cancer cells' phenotype: organization of MCF7 cells in 3D cultures

International Nuclear Information System (INIS)

Krause, Silva; Maffini, Maricel V; Soto, Ana M; Sonnenschein, Carlos

2010-01-01

Stromal-epithelial interactions mediate breast development, and the initiation and progression of breast cancer. In the present study, we developed 3-dimensional (3D) in vitro models to study breast cancer tissue organization and the role of the microenvironment in phenotypic determination. The human breast cancer MCF7 cells were grown alone or co-cultured with primary human breast fibroblasts. Cells were embedded in matrices containing either type I collagen or a combination of reconstituted basement membrane proteins and type I collagen. The cultures were carried out for up to 6 weeks. For every time point (1-6 weeks), the gels were fixed and processed for histology, and whole-mounted for confocal microscopy evaluation. The epithelial structures were characterized utilizing immunohistochemical techniques; their area and proliferation index were measured using computerized morphometric analysis. Statistical differences between groups were analyzed by ANOVA, Dunnett's T3 post-hoc test and chi-square. Most of the MCF7 cells grown alone within a collagen matrix died during the first two weeks; those that survived organized into large, round and solid clusters. The presence of fibroblasts in collagen gels reduced MCF7 cell death, induced cell polarity, and the formation of round and elongated epithelial structures containing a lumen. The addition of reconstituted basement membrane to collagen gels by itself had also survival and organizational effects on the MCF7 cells. Regardless of the presence of fibroblasts, the MCF7 cells both polarized and formed a lumen. The addition of fibroblasts to the gel containing reconstituted basement membrane and collagen induced the formation of elongated structures. Our results indicate that a matrix containing both type I collagen and reconstituted basement membrane, and the presence of normal breast fibroblasts constitute the minimal permissive microenvironment to induce near-complete tumor phenotype reversion. These human

Degradation of endothelial basement membrane by human breast cancer cell lines

International Nuclear Information System (INIS)

Yee, C.; Shiu, R.P.

1986-01-01

During metastasis, it is believed that tumor cells destroy the basement membrane (BM) of blood vessels in order to disseminate through the circulatory system. By radioactively labeling the extracellular matrix produced by primary endothelial cells in vitro, the ability of human breast cancer cells to degrade BM components was studied. We found that T-47D, a human breast cancer line, was able to degrade significant amounts of [35S]methionine-labeled and [3H]proline-labeled BM, but not 35SO4-labeled BM. Six other tumor cell lines of human breast origin were assayed in the same manner and were found to degrade BM to varying degrees. Several non-tumor cell lines tested showed relatively little degrading activity. The use of serum-free medium greatly enhanced degradation of the BM by tumor cells, suggesting a role for naturally occurring enzyme inhibitors in the serum. Direct cell contact with the BM was required for BM degradation, suggesting that the active enzymes are cell associated. The addition of hormones implicated in the etiology of breast cancer did not significantly alter the ability of T-47D cells to degrade the BM. The use of this assay affords future studies on the mechanism of invasion and metastasis of human breast cancer

Study on the Impact of Coagulation Bath Temperature on the Surface Morphology and Performance of Polyethylene Membrane Prepared by TIPS Method in Purification of Collagen Protein

Directory of Open Access Journals (Sweden)

Ali Akbari

2015-11-01

Full Text Available Fabrication of an efficient microfiltration polymeric membrane with low fouling characteristic and high permeation flux is an essential task for developing membrane-related researches and membrane industries. Surface skin layer which decreases the membrane permeation and accelerates the membrane fouling in purification and separation of protein solution is usually observed for all membranes fabricated via thermally induced phase separation (TIPS method. In this work, the impact of coagulation bath temperature on the skin layer thickness and performance of fabricated membranes was investigated. Collagen protein purification tests were carried out to investigate the impact of skin layer on the performance and determine the fouling mechanisms of the membranes. Obtained results showed that when coagulation bath temperature increases, the thickness of skin layer decreases. In membranes with lower surface porosity, decline in protein permeation is mainly due to the standard blocking fouling mechanism which is a kind of the irreversible fouling phenomenon. In membranes with higher surface porosity, however, decline in protein permeation is mainly due to the intermediate blocking fouling mechanism which is a kind of reversible fouling phenomenon. Obtained results from permeation flux and spectrophotometric analyses of inlet feed and retentate streams within 800 min showed that the collagen recovery ratio for modified and unmodified membranes were 5.6 and less than 1%, respectively. It is worth to mention that for membrane with lower surface porosity the collagen filtration process was stopped within 400 min due to the membrane fouling. For membrane with higher surface porosity, however there was no halting in filtration process within 800 min.

The use of allodermis prepared from Euro skin bank to prepare autologous tissue engineered skin for clinical use.

Science.gov (United States)

Deshpande, P; Ralston, D R; MacNeil, S

2013-09-01

Over the past two decades a range of 3D models for human skin have been described. Some include native collagen and intrinsic basement membrane proteins and fibroblasts, others are based on xenogeneic collagen or synthetic supports often without fibroblasts. The aim of this study was to look at the influence of media calcium, basement membrane and fibroblasts on the quality of 3D tissue engineered skin produced using human de-epidermized acellular dermis. In this study we deliberately used Euro skin as the source of the donor dermis to examine to what extent this could provide an effective dermal substrate for producing 3D skin for clinical use. Keratinocytes were cultured in the presence and absence of fibroblasts and both with and without basement membrane on decellularized dermis at calcium concentrations ranging from 250μM to 1.6mM over a period of 14 days. Results showed the formation of a well attached epithelium with many of the features of normal skin in the presence of a basement membrane. This was largely independent of the presence of fibroblasts and not greatly influenced by the concentration of calcium in the media. However there was a clear requirement for physiological levels of calcium in the formation of a stratified epithelium in the absence of a basement membrane. Copyright © 2013 Elsevier Ltd and ISBI. All rights reserved.

Ultrastructural study of electron dense deposits in renal tubular basement membrane: prevalence and relationship to epithelial atrophy.

Science.gov (United States)

Yong, Jim L C; Killingsworth, Murray C

2014-08-01

This study reports the prevalence of immune deposits associated with the proximal and distal tubules in a series of routine renal biopsies received in our department during a single calendar year. From 87 cases, 65 (74%) were found to have glomerular immune deposits by immunofluorescence. Tubular immune deposits were found in 12 cases (18%), 3 of which had no glomerular deposits. By transmission electron microscopy (EM), 58 cases (66%) were found to have deposits of granular or vesicular material associated with the tubular basement membranes (TBM). Finely granular electron dense deposits appeared to correspond to the immune deposits seen by immunofluorescence microscopy (IF) and may be a sensitive marker of immune deposition.

In vitro aging of mineralized collagen-based composite as guided tissue regeneration membrane

Energy Technology Data Exchange (ETDEWEB)

Pan, S.X. [Department of Prothodontics, School of Stomatology, Peking University, Beijing 100875 (China)]. E-mail: sx_pan@sina.com; Li, Y. [Department of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China); Feng, H.L. [Department of Prothodontics, School of Stomatology, Peking University, Beijing 100875 (China); Bai, W. [Department of Prothodontics, School of Stomatology, Peking University, Beijing 100875 (China); Gu, Y.Y. [Department of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China)

2006-05-15

The technique of guided tissue regeneration (GTR) has been developed for the regeneration of periodontal tissues, bone around natural teeth and dental implants. The aim of this study is to investigate the biodegradability and mechanic behavior of a novel mineralized nano-hydroxyapatite/collagen/poly (lactic acid) (nHAC/PLA) composite as GTR membrane in vitro. The elastic modulus and maximum tensile strength of GTR film samples with different nHAC/PLA ratio were measured to get an optimal nHAC/PLA ratio. Thermogravimetric analysis was conducted to evaluate the change of the inorganic component in the samples during the process of in vitro aging. Morphology of samples was checked by using scanning electron microscopy. On the basis of the above results, it can be concluded that the GTR membranes maintained integrity and the original appearance throughout the 1-month in vitro aging. There is an active dissolution and deposition process of crystals which is propitious to the bone formation on the surface of the composite membrane. The optimal nHAC/PLA ratio of the novel membrane is 0.4:1. For a longer period of bone repair, PLA with higher molecular weight should be chosen as the scaffold for the GTR membrane.

Type IV collagen as marker of fibrosis in nonalcoholic liver disease

Directory of Open Access Journals (Sweden)

Alvina Alvina

2016-02-01

Full Text Available Currently nonalcoholic fatty liver disease (NAFLD and nonalcoholic steatohepatitis (NASH are medical problems associated with the increasing prevalence of diabetes mellitus, obesity, hypertension and hypertriglyceridemia, usually designated as the metabolic syndrome associated with insulin resistance. One study demonstrated an increase in NAFLD prevalence of around 17-33% and in NASH prevalence of 5.7-16.5%. NAFLD comprises a range of mild to severe conditions, from simple steatosis to steatohepatitis, hepatic fibrosis and cirrhosis. The diagnosis of hepatic fibrosis is important for prognosis, stratification for treatment, and monitoring of treatment efficacy. Ultrasonography (USG is a simple method for detecting fatty infiltrates in the liver. USG has a sensitivity of 82-89% and a specificity of 93%, but cannot differentiate between hepatic steatosis and fibrosis. The gold standard for evaluation of hepatic fibrosis is liver biopsy, which however is a painful and invasive procedure. Currently determination of serum type IV collagen has been suggested as an alternative to liver biopsy among the non-invasive methods for evaluation of hepatic fibrosis, as its serum concentration is closely correlated with advanced hepatic fibrosis in NASH. Type IV collagen is one of the components of basement membrane and its serum concentration is indicative of degradation of the extracellular matrix.

Type IV collagen as marker of fibrosis in nonalcoholic liver disease

Directory of Open Access Journals (Sweden)

Alvina

2010-08-01

Full Text Available Currently nonalcoholic fatty liver disease (NAFLD and nonalcoholic steatohepatitis (NASH are medical problems associated with the increasing prevalence of diabetes mellitus, obesity, hypertension and hypertriglyceridemia, usually designated as the metabolic syndrome associated with insulin resistance. One study demonstrated an increase in NAFLD prevalence of around 17-33% and in NASH prevalence of 5.7-16.5%. NAFLD comprises a range of mild to severe conditions, from simple steatosis to steatohepatitis, hepatic fibrosis and cirrhosis. The diagnosis of hepatic fibrosis is important for prognosis, stratification for treatment, and monitoring of treatment efficacy. Ultrasonography (USG is a simple method for detecting fatty infiltrates in the liver. USG has a sensitivity of 82-89% and a specificity of 93%, but cannot differentiate between hepatic steatosis and fibrosis. The gold standard for evaluation of hepatic fibrosis is liver biopsy, which however is a painful and invasive procedure. Currently determination of serum type IV collagen has been suggested as an alternative to liver biopsy among the non-invasive methods for evaluation of hepatic fibrosis, as its serum concentration is closely correlated with advanced hepatic fibrosis in NASH. Type IV collagen is one of the components of basement membrane and its serum concentration is indicative of degradation of the extracellular matrix.

Large proteoglycan complexes and disturbed collagen architecture in the corneal extracellular matrix of mucopolysaccharidosis type VII (Sly syndrome).

Science.gov (United States)

Young, Robert D; Liskova, Petra; Pinali, Christian; Palka, Barbara P; Palos, Michalis; Jirsova, Katerina; Hrdlickova, Enkela; Tesarova, Marketa; Elleder, Milan; Zeman, Jiri; Meek, Keith M; Knupp, Carlo; Quantock, Andrew J

2011-08-24

Deficiencies in enzymes involved in proteoglycan (PG) turnover underlie a number of rare mucopolysaccharidoses (MPS), investigations of which can considerably aid understanding of the roles of PGs in corneal matrix biology. Here, the authors analyze novel pathologic changes in MPS VII (Sly syndrome) to determine the nature of PG-collagen associations in stromal ultrastructure. Transmission electron microscopy and electron tomography were used to investigate PG-collagen architectures and interactions in a cornea obtained at keratoplasty from a 22-year-old man with MPS VII, which was caused by a compound heterozygous mutation in the GUSB gene. Transmission electron microscopy showed atypical morphology of the epithelial basement membrane and Bowman's layer in MPS VII. Keratocytes were packed with cytoplasmic vacuoles containing abnormal glycosaminoglycan (GAG) material, and collagen fibrils were thinner than in normal cornea and varied considerably throughout anterior (14-32 nm), mid (13-42 nm), and posterior (17-39 nm) regions of the MPS VII stroma. PGs viewed in three dimensions were striking in appearance in that they were significantly larger than PGs in normal cornea and formed highly extended linkages with multiple collagen fibrils. Cellular changes in the MPS VII cornea resemble those in other MPS. However, the wide range of collagen fibril diameters throughout the stroma and the extensive matrix presence of supranormal-sized PG structures appear to be unique features of this disorder. The findings suggest that the accumulation of stromal chondroitin-, dermatan-, and heparan-sulfate glycosaminoglycans in the absence of β-glucuronidase-mediated degradation can modulate collagen fibrillogenesis.

Membrane-type-3 matrix metalloproteinase (MT3-MMP functions as a matrix composition-dependent effector of melanoma cell invasion.

Directory of Open Access Journals (Sweden)

Olga Tatti

Full Text Available In primary human melanoma, the membrane-type matrix metalloproteinase, MT3-MMP, is overexpressed in the most aggressive nodular-type tumors. Unlike MT1-MMP and MT2-MMP, which promote cell invasion through basement membranes and collagen type I-rich tissues, the function of MT3-MMP in tumor progression remains unclear. Here, we demonstrate that MT3-MMP inhibits MT1-MMP-driven melanoma cell invasion in three-dimensional collagen, while yielding an altered, yet MT1-MMP-dependent, form of expansive growth behavior that phenocopies the formation of nodular cell colonies. In melanoma cell lines originating from advanced primary or metastatic lesions, endogenous MT3-MMP expression was associated with limited collagen-invasive potential. In the cell lines with highest MT3-MMP expression relative to MT1-MMP, collagen-invasive activity was increased following stable MT3-MMP gene silencing. Consistently, MT3-MMP overexpression in cells derived from less advanced superficially spreading melanoma lesions, or in the MT3-MMP knockdown cells, reduced MT1-MMP-dependent collagen invasion. Rather than altering MT1-MMP transcription, MT3-MMP interacted with MT1-MMP in membrane complexes and reduced its cell surface expression. By contrast, as a potent fibrinolytic enzyme, MT3-MMP induced efficient invasion of the cells in fibrin, a provisional matrix component frequently found at tumor-host tissue interfaces and perivascular spaces of melanoma. Since MT3-MMP was significantly upregulated in biopsies of human melanoma metastases, these results identify MT3-MMP as a matrix-dependent modifier of the invasive tumor cell functions during melanoma progression.

Defective muscle basement membrane and lack of M-laminin in the dystrophic dy/dy mouse

DEFF Research Database (Denmark)

Xu, H; Christmas, P; Wu, X R

1994-01-01

-linked Duchenne and Becker muscular dystrophies. We have examined M-laminin expression in mice with autosomal recessive muscular dystrophy caused by the mutation dy. The heavy chain of M-laminin was undetectable in skeletal muscle, heart muscle, and peripheral nerve by immunofluorescence and immunoblotting......M-laminin is a major member of the laminin family of basement membrane proteins. It is prominently expressed in striated muscle and peripheral nerve. M-laminin is deficient in patients with the autosomal recessive Fukuyama congenital muscular dystrophy but is normal in patients with the sex...... tissue from dy/dy mice, suggesting that M-laminin heavy-chain mRNA may be produced at very low levels or is unstable. Information about the chromosomal localization of the M heavy-chain in human and mouse suggests that a mutation in the M-chain gene causes the muscular dystrophy in dy/dy mice. The dy...

[Development of a novel absorbable nanofiber chitosan-collagen membrane by electrospinning and the preliminary study on guided bone regeneration].

Science.gov (United States)

Gao, B; Li, X J; Lin, M; Li, Y Y; Dong, Y

2018-02-09

Objective: To evaluate the application effect of nanofiber chitosan-collagen membrane (NCM) on guided bone regeneration (GBR). Methods: The mixture of collagen, chitosan, polyethylene oxide was used to make up the NCM by electrospinning, then the NCM was crosslinked by glutaraldehyde vapor. The physical property of the NCM was measured by scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR). MC3T3-E1 osteoblasts were cultured on NCM to characterize the biocompatibility. The effectiveness of four groups [contrast group, Bio-gide membrane (BGM), compressed chitosan-collagen menbrane (CCM), NCM/CCM] on bone regeneration were evaluated in critical-sized defects (diameter = 5 mm) in SD rats. Results: When the mixed solution consists of 4.0% collagen, 1.0% chitosan and 3.5% polyethylene oxide, the NCM could be validly fabricated by electrospinning. After cross-linking by glutaraldehyde vapor, the tensile strength and the stability of NCM in damp was enhanced. No cytotoxicity of the NCM was detected on MC3T3-E1 osteoblasts. In vivo study showed that the new bone regeneration ratio of NCM/CCM group was [(43.10±1.49)%], and this was similar to that of the group of BGM [(41.36±2.60)%] ( P> 0.05), but higher than that of the CCM group [(33.10±1.41)%] and the contrast group [(7.22±2.46)%] ( P< 0.05). Conclusions: The NCM can promote new bone regeneration effectively in GBR procedure.

Fluorescent and radiolabelling of pepsin-digested human glomerular basement membrane with a newly developed hydroxy-coumarin derivative (CASE)

International Nuclear Information System (INIS)

Rand-Weaver, M.; Abuknesha, R.A.; Price, R.G.

1985-01-01

The labelling of pepsin-digested human glomerular basement membrane (pHGBM) with a newly developed fluorescent iodine acceptor 7-hydroxy-coumarin-3-acetic acid N-hydroxysucciniimydyl ester (CASE) is described. The binding of a monoclonal antibody to pHGBM was assessed by radiobinding assays, and when directly iodinated pHGBM was used there was no apparent binding. When CASE was conjugated to pHGBM prior to iodination 11% binding was achieved. CASE acting as an iodine acceptor may be useful for proteins containing few or inaccessible tyrosine residues or which are destroyed by introduction of 125 I. Since CASE is fluorescent, small amounts of material can be detected during isolation prior to iodination. (orig.)

Enhancement of tendon–bone healing via the combination of biodegradable collagen-loaded nanofibrous membranes and a three-dimensional printed bone-anchoring bolt

Directory of Open Access Journals (Sweden)

Chou YC

2016-08-01

Full Text Available Ying-Chao Chou,1,2 Wen-Lin Yeh,2 Chien-Lin Chao,1 Yung-Heng Hsu,1,2 Yi-Hsun Yu,1,2 Jan-Kan Chen,3 Shih-Jung Liu1,2 1Department of Mechanical Engineering, Chang Gung University, 2Department of Orthopedic Surgery, Chang Gung Memorial Hospital, 3Department of Physiology and Pharmacology, Chang Gung University, Taoyuan, Taiwan Abstract: A composite biodegradable polymeric model was developed to enhance tendon graft healing. This model included a biodegradable polylactide (PLA bolt as the bone anchor and a poly(D,L-lactide-co-glycolide (PLGA nanofibrous membrane embedded with collagen as a biomimic patch to promote tendon–bone interface integration. Degradation rate and compressive strength of the PLA bolt were measured after immersion in a buffer solution for 3 months. In vitro biochemical characteristics and the nanofibrous matrix were assessed using a water contact angle analyzer, pH meter, and tetrazolium reduction assay. In vivo efficacies of PLGA/collagen nanofibers and PLA bolts for tendon–bone healing were investigated on a rabbit bone tunnel model with histological and tendon pullout tests. The PLGA/collagen-blended nanofibrous membrane was a hydrophilic, stable, and biocompatible scaffold. The PLA bolt was durable for tendon–bone anchoring. Histology showed adequate biocompatibility of the PLA bolt on a medial cortex with progressive bone ingrowth and without tissue overreaction. PLGA nanofibers within the bone tunnel also decreased the tunnel enlargement phenomenon and enhanced tendon–bone integration. Composite polymers of the PLA bolt and PLGA/collagen nanofibrous membrane can effectively promote outcomes of tendon reconstruction in a rabbit model. The composite biodegradable polymeric system may be useful in humans for tendon reconstruction. Keywords: polylactide–polyglycolide nanofibers, PLGA, collagen, 3D printing, polylactide, PLA, bone-anchoring bolts, tendon healing

Anti-glomerular basement membrane autoantibodies in the Brown Norway rat: detection by a solid-phase radioimmunoassay

International Nuclear Information System (INIS)

Bowman, C.; Peters, D.K.; Lockwood, C.M.

1983-01-01

A solid-phase radioimmunoassay (RIA) is described for the detection of IgG autoantibodies to glomerular basement membrane (GBM) induced in the Brown Norway rat by mercuric chloride. The assay involves the adsorption of a collagenase digest of GBM to plastic microtitre plates and detection of bound antibody with affinity purified radiolabelled rabbit anti-rat IgG. Comparison with existing immunofluorescence methods for detection of anti-GBM antibody showed that the solid-phase RIA is highly sensitive, allowing detection of antibody in solutions with as low as 0.5 ng protein/ml. The assay is suitable for detection of anti-GBM antibody both in serum and in eluates from nephritic kidneys. The assay proved to be specific in competitive studies of inhibition brought about by GBM, keyhole limpet antigen and ovalbumin. This solid-phase RIA is reproducible, robust and easy to perform. (Auth.)

Identification of the NC1 domain of {alpha}3 chain as critical for {alpha}3{alpha}4{alpha}5 type IV collagen network assembly.

Science.gov (United States)

LeBleu, Valerie; Sund, Malin; Sugimoto, Hikaru; Birrane, Gabriel; Kanasaki, Keizo; Finan, Elizabeth; Miller, Caroline A; Gattone, Vincent H; McLaughlin, Heather; Shield, Charles F; Kalluri, Raghu

2010-12-31

The network organization of type IV collagen consisting of α3, α4, and α5 chains in the glomerular basement membrane (GBM) is speculated to involve interactions of the triple helical and NC1 domain of individual α-chains, but in vivo evidence is lacking. To specifically address the contribution of the NC1 domain in the GBM collagen network organization, we generated a mouse with specific loss of α3NC1 domain while keeping the triple helical α3 chain intact by connecting it to the human α5NC1 domain. The absence of α3NC1 domain leads to the complete loss of the α4 chain. The α3 collagenous domain is incapable of incorporating the α5 chain, resulting in the impaired organization of the α3α4α5 chain-containing network. Although the α5 chain can assemble with the α1, α2, and α6 chains, such assembly is incapable of functionally replacing the α3α4α5 protomer. This novel approach to explore the assembly type IV collagen in vivo offers novel insights in the specific role of the NC1 domain in the assembly and function of GBM during health and disease.

Development of a human corneal epithelium model utilizing a collagen vitrigel membrane and the changes of its barrier function induced by exposing eye irritant chemicals.

Science.gov (United States)

Takezawa, Toshiaki; Nishikawa, Kazunori; Wang, Pi-Chao

2011-09-01

The brief TEER (trans-epithelial electrical resistance) assay after exposing chemicals to corneal epithelium in vivo is known as a suitable method for evaluating corneal irritancy and permeability quantitatively and continuously. A collagen vitrigel membrane we previously developed is a thin (about 20 μm thick) and transparent membrane composed of high density collagen fibrils equivalent to connective tissues in vivo, e.g. corneal Bowman's membrane. To develop such a TEER assay system in vitro utilizing a human corneal epithelial model, HCE-T cells (a human corneal epithelial cell line) were cultured on the collagen vitrigel membrane substratum prepared in a Millicell chamber suitable for TEER measurement. Human corneal epithelium model possessing 5-6 cell layers sufficient for TEER assay was successfully reconstructed on the substratum in the Millicell chamber by culturing the cells in monolayer for 2 days and subsequently in air-liquid interface for 7 days. The exposure of chemicals to the model induced the time-dependent relative changes of TEER in response to the characteristic of each chemical within a few minutes. These results suggest that the TEER assay using the human corneal epithelial model is very useful for an ocular irritancy evaluation as an alternative to the Draize eye irritation test. Copyright © 2011 Elsevier Ltd. All rights reserved.

A direct contact between astrocyte and vitreous body is possible in the rabbit eye due to discontinuities in the basement membrane of the retinal inner limiting membrane

Directory of Open Access Journals (Sweden)

A. Haddad

2003-02-01

Full Text Available Different from most mammalian species, the optic nerve of the rabbit eye is initially formed inside the retina where myelination of the axons of the ganglion cells starts and vascularization occurs. Astrocytes are confined to these regions. The aforementioned nerve fibers known as medullated nerve fibers form two bundles that may be identified with the naked eye. The blood vessels run on the inner surface of these nerve fiber bundles (epivascularization and, accordingly, the accompanying astrocytes lie mostly facing the vitreous body from which they are separated only by the inner limiting membrane of the retina. The arrangement of the astrocytes around blood vessels leads to the formation of structures known as glial tufts. Fragments (N = 3 or whole pieces (N = 3 of the medullated nerve fiber region of three-month-old male rabbits (Orictolagus cuniculus were fixed in glutaraldehyde followed by osmium tetroxide, and their thin sections were examined with a transmission electron microscope. Randomly located discontinuities (up to a few micrometers long of the basement membrane of the inner limiting membrane of the retina were observed in the glial tufts. As a consequence, a direct contact between the astrocyte plasma membrane and vitreous elements was demonstrated, making possible functional interactions such as macromolecular exchanges between this glial cell type and the components of the vitreous body.

Still more complexity in mammalian basement membranes

DEFF Research Database (Denmark)

Erickson, A C; Couchman, J R

2000-01-01

laminins, entactin-1/nidogen-1, Type IV collagen, and proteoglycans. However, within the past few years this complexity has increased as new components are described. The entactin/nidogen (E/N) family has expanded with the recent description of a new isoform, E/N-2/osteonidogen. Agrin and Type XVIII...... to be regulated through multiple, mostly domain-specific mechanisms. Understanding the functions of individual BM components and their assembly into macromolecular complexes is a considerable challenge that may increase as further BM and cell surface ligands are discovered for these proteins....

Anti-glomerular basement membrane glomerulonephritis in an HIV positive patient: case report

Directory of Open Access Journals (Sweden)

Eduardo José Bellotto Monteiro

Full Text Available We report on a case of a patient with HIV infection, diagnosed 18 months prior to the development of an anti-glomerular basement membrane (anti-GBM rapidly progressive glomerulonephritis; this is probably the first report of such an association. A 30-year-old white man presented with elevation of serum creatinine (1.3 - 13.5 mg/dL within one month. At admission, the urinalysis showed proteinuria of 7.2 g/L and 8,000,000 erythrocytes/mL. Renal biopsy corresponded to a crescentic diffuse proliferative glomerulonephritis mediated by anti-GBM, and serum testing for anti-GBM antibodies was positive; antinuclear antibodies (ANA and anti-neutrophilic cytoplasmic antibodies (ANCA were also positive. The patient underwent hemodyalisis and was treated with plasmapheresis, cyclophosphamide and prednisone. The association described here is not casual, as crescentic glomerulonephritis is not common in HIV-positive patients, anti-GBM glomerulonephritis is rare and anti-GBM antibodies are frequently observed in HIV-positive subjects when compared to the overall population. Based on the current case and on the elevated frequency of the positivity for such antibodies in this group of patients, it is advisable to be aware of the eventual association between these two conditions and to promote an active search for anti-GBM antibodies and early diagnosis of eventual urinary abnormalities in HIV-positive subjects, considering the severity of anti-GBM glomerulonephritis.

Functional Imaging of Proteolysis: Stromal and Inflammatory Cells Increase Tumor Proteolysis

Directory of Open Access Journals (Sweden)

Mansoureh Sameni

2003-07-01

Full Text Available The underlying basement membrane is degraded during progression of breast and colon carcinoma. Thus, we imaged degradation of a quenched fluorescent derivative of basement membrane type IV collagen (DQ-collagen IV by living human breast and colon tumor spheroids. Proteolysis of DQ-collagen IV by HCT 116 and HKh-2 human colon tumor spheroids was both intracellular and pericellular. In contrast, proteolysis of DQ-collagen IV by BT20 human breast tumor spheroids was pericellular. As stromal elements can contribute to proteolytic activities associated with tumors, we also examined degradation of DQ-collagen IV by human monocytes/macrophages and colon and breast fibroblasts. Fibroblasts themselves exhibited a modest amount of pericellular degradation. Degradation was increased 4–17-fold in cocultures of fibroblasts and tumor cells as compared to either cell type alone. Inhibitors of matrix metalloproteinases, plasmin, and the cysteine protease, cathepsin B, all reduced degradation in the cocultures. Monocytes did not degrade DQ-collagen IV; however, macrophages degraded DQ-collagen IV intracellularly. In coculture of tumor cells, fibroblasts, and macrophages, degradation of DQ-collagen IV was further increased. Imaging of living tumor and stromal cells has, thus, allowed us to establish that tumor proteolysis occurs pericellularly and intracellularly and that tumor, stromal, and inflammatory cells all contribute to degradative processes.

Smad4-dependent pathways control basement membrane deposition and endodermal cell migration at early stages of mouse development

Directory of Open Access Journals (Sweden)

Taylor Jennifer M

2009-10-01

Full Text Available Abstract Background Smad4 mutant embryos arrest shortly after implantation and display a characteristic shortened proximodistal axis, a significantly reduced epiblast, as well as a thickened visceral endoderm layer. Conditional rescue experiments demonstrate that bypassing the primary requirement for Smad4 in the extra-embryonic endoderm allows the epiblast to gastrulate. Smad4-independent TGF-β signals are thus sufficient to promote mesoderm formation and patterning. To further analyse essential Smad4 activities contributed by the extra-embryonic tissues, and characterise Smad4 dependent pathways in the early embryo, here we performed transcriptional profiling of Smad4 null embryonic stem (ES cells and day 4 embryoid bodies (EBs. Results Transcripts from wild-type versus Smad4 null ES cells and day 4 EBs were analysed using Illumina arrays. In addition to several known TGF-β/BMP target genes, we identified numerous Smad4-dependent transcripts that are mis-expressed in the mutants. As expected, mesodermal cell markers were dramatically down-regulated. We also observed an increase in non-canonical potency markers (Pramel7, Tbx3, Zscan4, germ cell markers (Aire, Tuba3a, Dnmt3l as well as early endoderm markers (Dpp4, H19, Dcn. Additionally, expression of the extracellular matrix (ECM remodelling enzymes Mmp14 and Mmp9 was decreased in Smad4 mutant ES and EB populations. These changes, in combination with increased levels of laminin alpha1, cause excessive basement membrane deposition. Similarly, in the context of the Smad4 null E6.5 embryos we observed an expanded basement membrane (BM associated with the thickened endoderm layer. Conclusion Smad4 functional loss results in a dramatic shift in gene expression patterns and in the endodermal cell lineage causes an excess deposition of, or an inability to breakdown and remodel, the underlying BM layer. These structural abnormalities probably disrupt reciprocal signalling between the epiblast and

Optically Highlighting Basement Membrane Components in C. elegans

OpenAIRE

sprotocols

2015-01-01

Authors: Elliott Hagedorn & David Sherwood ### Abstract Green fluorescent protein (GFP) and other genetically encoded fluorescent proteins provide a means to study gene expression pattern and protein localization in living tissues. Recently discovered GFP-like fluorophores and engineered variants have further expanded the fluorescent protein toolkit for in vivo imaging. Here we describe a technique using transgenic C. elegans that contain laminin or type IV collagen fused to the green...

Comparative evaluation of a bioabsorbable collagen membrane and connective tissue graft in the treatment of localized gingival recession: A clinical study

Directory of Open Access Journals (Sweden)

Harsha Mysore Babu

2011-01-01

Full Text Available Background: Gingival recession (GR can result in root sensitivity, esthetic concern to the patient, and predilection to root caries. The purpose of this randomized clinical study was to evaluate (1 the effect of guided tissue regeneration (GTR procedure using a bioabsorbable collagen membrane, in comparison to autogenous subepithelial connective tissue graft (SCTG for root coverage in localized gingival recession defects; and (2 the change in width of keratinized gingiva following these two procedures. Materials and Methods: A total of 10 cases, showing at least two localized Miller′s Class I or Class II gingival recession, participated in this study. In a split mouth design, the pairs of defects were randomly assigned for treatment with either SCTG (SCTG Group or GTR-based collagen membrane (GTRC Group. Both the grafts were covered with coronally advanced flap. Recession depth (RD, recession width (RW, width of keratinized gingiva (KG, probing depth (PD, relative attachment level (RAL, plaque index (PI, and gingival index (GI were recorded at baseline, 3 and 6 months postoperatively. Results: Six months following root coverage procedures, the mean root coverage was found to be 84.84% ± 16.81% and 84.0% ± 15.19% in SCTG Group and GTRC Group, respectively. The mean keratinized gingival width increase was 1.50 ± 0.70 mm and 2.30 ± 0.67 mm in the SCTG and GTRC group, respectively, which was not statistically significant. Conclusion: It may be concluded that resorbable collagen membrane can be a reliable alternative to autogenous connective tissue graft in the treatment of gingival recession.

Differences in collagen distribution of healthy and regenerated periodontium. Histomorphometric study in dogs.

Science.gov (United States)

Souza, Sérgio L S; Macedo, Guilherme O; Silveira E Souza, Adriana M M; Taba, Mário; Novaes, Arthur B; Oliver, Constance; Jamur, Maria C; Correa, Vani M A

2013-10-01

Previous studies have shown that there is a relationship between periodontal disease and the distribution of collagen fibers. This study evaluated the distribution of collagen types I and III in regenerated bone and periodontal ligament, comparing them to the tissues near the regenerated area and to the healthy periodontium. In the third (P3) and fourth (P4) mandibular premolars of 5 healthy mongrel dogs, bilaterally, buccal class 2 furcation lesions were surgically created and chronified for 3 weeks. After that, full flaps were elevated and expanded polytetrafluoroethylene (e-PTFE) membranes were adapted, sutured and recovered by the flaps. Two weeks after surgery, two membranes on the same side were removed and the other membranes were removed four weeks after surgery. The dogs were euthanized at 12 weeks following placement of the e-PTFE membranes. P3 and P4 teeth as well as the second premolars (healthy control teeth) and their periodontal tissues were removed and histologically processed for Collagen Quantification (COLQ). The amount of type III collagen was higher in native bone compared to the regenerated area. For periodontal ligament, COLQ for type I collagen showed statistically significant differences (Tukeys's Multiple Comparison, p⟨0.05) between the regenerated groups and the control group. These differences were not found for type III COLQ. There are significant differences in collagen distribution among the regenerated, native and control tissues. Membrane removal 2 or 4 weeks postoperatively did not influence the collagen composition.

Irradiation Alters MMP-2/TIMP-2 System and Collagen Type IV Degradation in Brain

Energy Technology Data Exchange (ETDEWEB)

Lee, Won Hee [School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia (United States); Warrington, Junie P.; Sonntag, William E. [Reynolds Oklahoma Center on Aging, Department of Geriatric Medicine, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma (United States); Lee, Yong Woo, E-mail: ywlee@vt.edu [School of Biomedical Engineering and Sciences, Virginia Polytechnic Institute and State University, Blacksburg, Virginia (United States); Department of Biomedical Sciences and Pathobiology, Virginia Polytechnic Institute and State University, Blacksburg, Virginia (United States)

2012-04-01

Purpose: Blood-brain barrier (BBB) disruption is one of the major consequences of radiation-induced normal tissue injury in the central nervous system. We examined the effects of whole-brain irradiation on matrix metalloproteinases (MMPs)/tissue inhibitors of metalloproteinases (TIMPs) and extracellular matrix (ECM) degradation in the brain. Methods and Materials: Animals received either whole-brain irradiation (a single dose of 10 Gy {gamma}-rays or a fractionated dose of 40 Gy {gamma}-rays, total) or sham-irradiation and were maintained for 4, 8, and 24 h following irradiation. mRNA expression levels of MMPs and TIMPs in the brain were analyzed by real-time reverse transcriptase-polymerase chain reaction (PCR). The functional activity of MMPs was measured by in situ zymography, and degradation of ECM was visualized by collagen type IV immunofluorescent staining. Results: A significant increase in mRNA expression levels of MMP-2, MMP-9, and TIMP-1 was observed in irradiated brains compared to that in sham-irradiated controls. In situ zymography revealed a strong gelatinolytic activity in the brain 24 h postirradiation, and the enhanced gelatinolytic activity mediated by irradiation was significantly attenuated in the presence of anti-MMP-2 antibody. A significant reduction in collagen type IV immunoreactivity was also detected in the brain at 24 h after irradiation. In contrast, the levels of collagen type IV were not significantly changed at 4 and 8 h after irradiation compared with the sham-irradiated controls. Conclusions: The present study demonstrates for the first time that radiation induces an imbalance between MMP-2 and TIMP-2 levels and suggests that degradation of collagen type IV, a major ECM component of BBB basement membrane, may have a role in the pathogenesis of brain injury.

Irradiation Alters MMP-2/TIMP-2 System and Collagen Type IV Degradation in Brain

International Nuclear Information System (INIS)

Lee, Won Hee; Warrington, Junie P.; Sonntag, William E.; Lee, Yong Woo

2012-01-01

Purpose: Blood-brain barrier (BBB) disruption is one of the major consequences of radiation-induced normal tissue injury in the central nervous system. We examined the effects of whole-brain irradiation on matrix metalloproteinases (MMPs)/tissue inhibitors of metalloproteinases (TIMPs) and extracellular matrix (ECM) degradation in the brain. Methods and Materials: Animals received either whole-brain irradiation (a single dose of 10 Gy γ-rays or a fractionated dose of 40 Gy γ-rays, total) or sham-irradiation and were maintained for 4, 8, and 24 h following irradiation. mRNA expression levels of MMPs and TIMPs in the brain were analyzed by real-time reverse transcriptase-polymerase chain reaction (PCR). The functional activity of MMPs was measured by in situ zymography, and degradation of ECM was visualized by collagen type IV immunofluorescent staining. Results: A significant increase in mRNA expression levels of MMP-2, MMP-9, and TIMP-1 was observed in irradiated brains compared to that in sham-irradiated controls. In situ zymography revealed a strong gelatinolytic activity in the brain 24 h postirradiation, and the enhanced gelatinolytic activity mediated by irradiation was significantly attenuated in the presence of anti-MMP-2 antibody. A significant reduction in collagen type IV immunoreactivity was also detected in the brain at 24 h after irradiation. In contrast, the levels of collagen type IV were not significantly changed at 4 and 8 h after irradiation compared with the sham-irradiated controls. Conclusions: The present study demonstrates for the first time that radiation induces an imbalance between MMP-2 and TIMP-2 levels and suggests that degradation of collagen type IV, a major ECM component of BBB basement membrane, may have a role in the pathogenesis of brain injury.

The collagen receptor uPARAP/Endo180

DEFF Research Database (Denmark)

Engelholm, Lars H; Ingvarsen, Signe; Jürgensen, Henrik J

2009-01-01

The uPAR-associated protein (uPARAP/Endo180), a type-1 membrane protein belonging to the mannose receptor family, is an endocytic receptor for collagen. Through this endocytic function, the protein takes part in a previously unrecognized mechanism of collagen turnover. uPARAP/Endo180 can bind...... and internalize both intact and partially degraded collagens. In some turnover pathways, the function of the receptor probably involves an interplay with certain matrix-degrading proteases whereas, in other physiological processes, redundant mechanisms involving both endocytic and pericellular collagenolysis seem...... in collagen breakdown seems to be involved in invasive tumor growth Udgivelsesdato: 2009...

Airway basement membrane perimeter in human airways is not a constant; potential implications for airway remodeling in asthma.

Science.gov (United States)

McParland, Brent E; Paré, Peter D; Johnson, Peter R A; Armour, Carol L; Black, Judith L

2004-08-01

Many studies that demonstrate an increase in airway smooth muscle in asthmatic patients rely on the assumption that bronchial internal perimeter (P(i)) or basement membrane perimeter (P(bm)) is a constant, i.e., not affected by fixation pressure or the degree of smooth muscle shortening. Because it is the basement membrane that has been purported to be the indistensible structure, this study examines the assumption that P(bm) is not affected by fixation pressure. P(bm) was determined for the same human airway segment (n = 12) fixed at distending pressures of 0 cmH(2)O and 21 cmH(2)O in the absence of smooth muscle tone. P(bm) for the segment fixed at 0 cmH(2)O was determined morphometrically, and the P(bm) for the same segment, had the segment been fixed at 21 cmH(2)O, was predicted from knowing the luminal volume and length of the airway when distended to 21 cmH(2)O (organ bath-derived P(i)). To ensure an accurate transformation of the organ bath-derived P(i) value to a morphometry-derived P(bm) value, had the segment been fixed at 21 cmH(2)O, the relationship between organ bath-derived P(i) and morphometry-derived P(bm) was determined for five different bronchial segments distended to 21 cmH(2)O and fixed at 21 cmH(2)O (r(2) = 0.99, P < 0.0001). Mean P(bm) for bronchial segments fixed at 0 cmH(2)O was 9.4 +/- 0.4 mm, whereas mean predicted P(bm), had the segments been fixed at 21 cmH(2)O, was 14.1 +/- 0.5 mm (P < 0.0001). This indicates that P(bm) is not a constant when isolated airway segments without smooth muscle tone are fixed distended to 21 cmH(2)O. The implication of these results is that the increase in smooth muscle mass in asthma may have been overestimated in some previous studies. Therefore, further studies are required to examine the potential artifact using whole lungs with and without abolition of airway smooth muscle tone and/or inflation.

Novel Vanadium-Loaded Ordered Collagen Scaffold Promotes Osteochondral Differentiation of Bone Marrow Progenitor Cells

Directory of Open Access Journals (Sweden)

Ana M. Cortizo

2016-01-01

Full Text Available Bone and cartilage regeneration can be improved by designing a functionalized biomaterial that includes bioactive drugs in a biocompatible and biodegradable scaffold. Based on our previous studies, we designed a vanadium-loaded collagen scaffold for osteochondral tissue engineering. Collagen-vanadium loaded scaffolds were characterized by SEM, FTIR, and permeability studies. Rat bone marrow progenitor cells were plated on collagen or vanadium-loaded membranes to evaluate differences in cell attachment, growth and osteogenic or chondrocytic differentiation. The potential cytotoxicity of the scaffolds was assessed by the MTT assay and by evaluation of morphological changes in cultured RAW 264.7 macrophages. Our results show that loading of VOAsc did not alter the grooved ordered structure of the collagen membrane although it increased membrane permeability, suggesting a more open structure. The VOAsc was released to the media, suggesting diffusion-controlled drug release. Vanadium-loaded membranes proved to be a better substratum than C0 for all evaluated aspects of BMPC biocompatibility (adhesion, growth, and osteoblastic and chondrocytic differentiation. In addition, there was no detectable effect of collagen or vanadium-loaded scaffolds on macrophage viability or cytotoxicity. Based on these findings, we have developed a new ordered collagen scaffold loaded with VOAsc that shows potential for osteochondral tissue engineering.

Clinical comparison of guided tissue regeneration, with collagen membrane and bone graft, versus connective tissue graft in the treatment of gingival recessions

Directory of Open Access Journals (Sweden)

Haghighati F

2006-06-01

Full Text Available Background and Aim: Increasing patient demands for esthetic, put the root coverage procedures in particular attention. Periodontal regeneration with GTR based root coverage methods is the most common treatment used. The purpose of this study was to compare guided tissue regeneration (GTR with collagen membrane and a bone graft, with sub-epithelial connective tissue graft (SCTG, in treatment of gingival recession. Materials and Methods: In this randomized clinical trial study, eleven healthy patients with no systemic diseases who had miller’s class I or II recession defects (gingival recession  2mm were treated with SCTG or GTR using a collagen membrane and a bone graft. Clinical measurements were obtained at baseline and 6 months after surgery. These clinical measurements included recession depth (RD, recession width (RW, probing depth (PD, and clinical attachment level (CAL. Data were analyzed using independent t test with p<0.05 as the limit of significance. Results: Both treatment methods resulted in a statistically significant reduction of recession depth (SCTG=2.3mm, GTR=2.1mm; P<0.0001. CAL gain after 6 months was also improved in both groups (SCG= 2.5mm, GTR=2.1mm, compared to baseline (P<0.0001. No statistical differences were observed in RD, RW, CAL between test and control groups. Root coverage was similar in both methods (SCTG= 74.2%, GTR= 62.6%, P=0.87. Conclusion: Based on the results of this study, the two techniques are clinically comparable. Therefore the use of collagen membrane and a bovine derived xenograft may alleviate the need for connective tissue graft.

A urokinase receptor-associated protein with specific collagen binding properties

DEFF Research Database (Denmark)

Behrendt, N; Jensen, O N; Engelholm, L H

2000-01-01

membrane-bound lectin with hitherto unknown function. The human cDNA was cloned and sequenced. The protein, designated uPARAP, is a member of the macrophage mannose receptor protein family and contains a putative collagen-binding (fibronectin type II) domain in addition to 8 C-type carbohydrate recognition...... domains. It proved capable of binding strongly to a single type of collagen, collagen V. This collagen binding reaction at the exact site of plasminogen activation on the cell may lead to adhesive functions as well as a contribution to cellular degradation of collagen matrices....

Stabilization of collagen nanofibers with l-lysine improves the ability of carbodiimide cross-linked amniotic membranes to preserve limbal epithelial progenitor cells

Directory of Open Access Journals (Sweden)

Lai JY

2014-11-01

Full Text Available Jui-Yang Lai,1–3 Pei-Ran Wang,1 Li-Jyuan Luo,1 Si-Tan Chen1 1Institute of Biochemical and Biomedical Engineering, 2Biomedical Engineering Research Center, 3Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan, Republic of ChinaAbstract: To overcome the drawbacks associated with limited cross-linking efficiency of carbodiimide modified amniotic membrane, this study investigated the use of L-lysine as an additional amino acid bridge to enhance the stability of a nanofibrous tissue matrix for a limbal epithelial cell culture platform. Results of ninhydrin assays and zeta potential measurements showed that the amount of positively charged amino acid residues incorporated into the tissue collagen chains is highly correlated with the L-lysine -pretreated concentration. The cross-linked structure and hydrophilicity of amniotic membrane scaffolding materials affected by the lysine molecular bridging effects were determined. With an increase in the L-lysine-pretreated concentration from 1 to 30 mM, the cross-linking density was significantly increased and water content was markedly decreased. The variations in resistance to thermal denaturation and enzymatic degradation were in accordance with the number of cross-links per unit mass of amniotic membrane, indicating L-lysine-modulated stabilization of collagen molecules. It was also noteworthy that the carbodiimide cross-linked tissue samples prepared using a relatively high L-lysine-pretreated concentration (ie, 30 mM appeared to have decreased light transmittance and biocompatibility, probably due to the influence of a large nanofiber size and a high charge density. The rise in stemness gene and protein expression levels was dependent on improved cross-link formation, suggesting the crucial role of amino acid bridges in constructing suitable scaffolds to preserve limbal progenitor cells. It is concluded that mild to moderate pretreatment conditions (ie, 3–10 mM L-lysine can

Cell cytoskeletal changes effected by static compressive stress lead to changes in the contractile properties of tissue regenerative collagen membranes

Directory of Open Access Journals (Sweden)

K Gellynck

2013-06-01

Full Text Available Static compressive stress can influence the matrix, which subsequently affects cell behaviour and the cell’s ability to further transform the matrix. This study aimed to assess response to static compressive stress at different stages of osteoblast differentiation and assess the cell cytoskeleton’s role as a conduit of matrix-derived stimuli. Mouse bone marrow mesenchymal stem cells (MSCs (D1 ORL UVA, osteoblastic cells (MC3T3-E1 and post-osteoblast/pre-osteocyte-like cells (MLO-A5 were seeded in hydrated and compressed collagen gels. Contraction was quantified macroscopically, and cell morphology, survival, differentiation and mineralisation assessed using confocal microscopy, alamarBlue® assay, real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR and histological stains, respectively. Confocal microscopy demonstrated cell shape changes and favourable microfilament organisation with static compressive stress of the collagen matrix; furthermore, cell survival was greater compared to the hydrated gels. The stage of osteoblast differentiation determined the degree of matrix contraction, with MSCs demonstrating the greatest amount. Introduction of microfilament disrupting inhibitors confirmed that pre-stress and tensegrity forces were under the influence of gel density, and there was increased survival and differentiation of the cells within the compressed collagen compared to the hydrated collagen. There was also relative stiffening and differentiation with time of the compressed cell-seeded collagen, allowing for greater manipulation. In conclusion, the combined collagen chemistry and increased density of the microenvironment can promote upregulation of osteogenic genes and mineralisation; MSCs can facilitate matrix contraction to form an engineered membrane with the potential to serve as a ‘pseudo-periosteum’ in the regeneration of bone defects.

A bioartificial environment for kidney epithelial cells based on a supramolecular polymer basement membrane mimic and an organotypical culture system.

Science.gov (United States)

Mollet, Björne B; Bogaerts, Iven L J; van Almen, Geert C; Dankers, Patricia Y W

2017-06-01

Renal applications in healthcare, such as renal replacement therapies and nephrotoxicity tests, could potentially benefit from bioartificial kidney membranes with fully differentiated and functional human tubular epithelial cells. A replacement of the natural environment of these cells is required to maintain and study cell functionality cell differentiation in vitro. Our approach was based on synthetic supramolecular biomaterials to mimic the natural basement membrane (BM) on which these cells grow and a bioreactor to provide the desired organotypical culture parameters. The BM mimics were constructed from ureidopyrimidinone (UPy)-functionalized polymer and bioactive peptides by electrospinning. The resultant membranes were shown to have a hierarchical fibrous BM-like structure consisting of self-assembled nanofibres within the electrospun microfibres. Human kidney-2 (HK-2) epithelial cells were cultured on the BM mimics under organotypical conditions in a custom-built bioreactor. The bioreactor facilitated in situ monitoring and functionality testing of the cultures. Cell viability and the integrity of the epithelial cell barrier were demonstrated inside the bioreactor by microscopy and transmembrane leakage of fluorescently labelled inulin, respectively. Furthermore, HK-2 cells maintained a polarized cell layer and showed modulation of both gene expression of membrane transporter proteins and metabolic activity of brush border enzymes when subjected to a continuous flow of culture medium inside the new bioreactor for 21 days. These results demonstrated that both the culture and study of renal epithelial cells was facilitated by the bioartificial in vitro environment that is formed by synthetic supramolecular BM mimics in our custom-built bioreactor. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

A role for PDGF-C/PDGFRα signaling in the formation of the meningeal basement membranes surrounding the cerebral cortex

Science.gov (United States)

Andrae, Johanna; Gouveia, Leonor; Gallini, Radiosa; He, Liqun; Fredriksson, Linda; Nilsson, Ingrid; Johansson, Bengt R.; Eriksson, Ulf; Betsholtz, Christer

2016-01-01

ABSTRACT Platelet-derived growth factor-C (PDGF-C) is one of three known ligands for the tyrosine kinase receptor PDGFRα. Analysis of Pdgfc null mice has demonstrated roles for PDGF-C in palate closure and the formation of cerebral ventricles, but redundancy with other PDGFRα ligands might obscure additional functions. In search of further developmental roles for PDGF-C, we generated mice that were double mutants for Pdgfc−/− and PdgfraGFP/+. These mice display a range of severe phenotypes including spina bifida, lung emphysema, abnormal meninges and neuronal over-migration in the cerebral cortex. We focused our analysis on the central nervous system (CNS), where PDGF-C was identified as a critical factor for the formation of meninges and assembly of the glia limitans basement membrane. We also present expression data on Pdgfa, Pdgfc and Pdgfra in the cerebral cortex and microarray data on cerebral meninges. PMID:26988758

Making more matrix: enhancing the deposition of dermal-epidermal junction components in vitro and accelerating organotypic skin culture development, using macromolecular crowding.

Science.gov (United States)

Benny, Paula; Badowski, Cedric; Lane, E Birgitte; Raghunath, Michael

2015-01-01

Skin is one of the most accessible tissues for experimental biomedical sciences, and cultured skin cells represent one of the longest-running clinical applications of stem cell therapy. However, culture-generated skin mimetic multicellular structures are still limited in their application by the time taken to develop these constructs in vitro and by their incomplete differentiation. The development of a functional dermal-epidermal junction (DEJ) is one of the most sought after aspects of cultured skin, and one of the hardest to recreate in vitro. At the DEJ, dermal fibroblasts and epidermal keratinocytes interact to form an interlinked basement membrane of extracellular matrix (ECM), which forms as a concerted action of both keratinocytes and fibroblasts. Successful formation of this basement membrane is essential for take and stability of cultured skin autografts. We studied interactive matrix production by monocultures and cocultures of primary human keratinocytes and fibroblasts in an attempt to improve the efficiency of basement membrane production in culture using mixed macromolecular crowding (mMMC); resulting ECM were enriched with the deposition of collagens I, IV, fibronectin, and laminin 332 (laminin 5) and also in collagen VII, the anchoring fibril component. Our in vitro data point to fibroblasts, rather than keratinocytes, as the major cellular contributors of the DEJ. Not only did we find more collagen VII production and deposition by fibroblasts in comparison to keratinocytes, but also observed that decellularized fibroblast ECM stimulated the production and deposition of collagen VII by keratinocytes, over and above that of keratinocyte monocultures. In confrontation cultures, keratinocytes and fibroblasts showed spontaneous segregation and demarcation of cell boundaries by DEJ protein deposition. Finally, mMMC was used in a classical organotypic coculture protocol with keratinocytes seeded over fibroblast-containing collagen gels. Applied during

In vitro cultivation of canine multipotent mesenchymal stromal cells on collagen membranes treated with hyaluronic acid for cell therapy and tissue regeneration

International Nuclear Information System (INIS)

Wodewotzky, T.I.; Lima-Neto, J.F.; Pereira-Júnior, O.C.M.; Sudano, M.J.; Lima, S.A.F.; Bersano, P.R.O.; Yoshioka, S.A.; Landim-Alvarenga, F.C.

2012-01-01

Support structures for dermal regeneration are composed of biodegradable and bioresorbable polymers, animal skin or tendons, or are bacteria products. The use of such materials is controversial due to their low efficiency. An important area within tissue engineering is the application of multipotent mesenchymal stromal cells (MSCs) to reparative surgery. The combined use of biodegradable membranes with stem cell therapy may lead to promising results for patients undergoing unsuccessful conventional treatments. Thus, the aim of this study was to test the efficacy of using membranes composed of anionic collagen with or without the addition of hyaluronic acid (HA) as a substrate for adhesion and in vitro differentiation of bone marrow-derived canine MSCs. The benefit of basic fibroblast growth factor (bFGF) on the differentiation of cells in culture was also tested. MSCs were collected from dog bone marrow, isolated and grown on collagen scaffolds with or without HA. Cell viability, proliferation rate, and cellular toxicity were analyzed after 7 days. The cultured cells showed uniform growth and morphological characteristics of undifferentiated MSCs, which demonstrated that MSCs successfully adapted to the culture conditions established by collagen scaffolds with or without HA. This demonstrates that such scaffolds are promising for applications to tissue regeneration. bFGF significantly increased the proliferative rate of MSCs by 63% when compared to groups without the addition of the growth factor. However, the addition of bFGF becomes limiting, since it has an inhibitory effect at high concentrations in culture medium

In vitro cultivation of canine multipotent mesenchymal stromal cells on collagen membranes treated with hyaluronic acid for cell therapy and tissue regeneration

Energy Technology Data Exchange (ETDEWEB)

Wodewotzky, T.I.; Lima-Neto, J.F. [Departamento de Reprodução Animal e Radiologia Veterinária, Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual de São Paulo, Botucatu, SP (Brazil); Pereira-Júnior, O.C.M. [Departamento de Reprodução Animal e Radiologia Veterinária, Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual de São Paulo, Botucatu, SP (Brazil); Departamento de Cirurgia e Anestesiologia Veterinária, Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual de São Paulo, Botucatu, SP (Brazil); Sudano, M.J.; Lima, S.A.F. [Departamento de Reprodução Animal e Radiologia Veterinária, Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual de São Paulo, Botucatu, SP (Brazil); Bersano, P.R.O. [Departamento de Patologia Veterinária, Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual de São Paulo, Botucatu, SP (Brazil); Yoshioka, S.A. [Instituto de Química de São Carlos, Universidade de São Paulo, São Carlos, SP (Brazil); Landim-Alvarenga, F.C. [Departamento de Reprodução Animal e Radiologia Veterinária, Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual de São Paulo, Botucatu, SP (Brazil)

2012-09-21

Support structures for dermal regeneration are composed of biodegradable and bioresorbable polymers, animal skin or tendons, or are bacteria products. The use of such materials is controversial due to their low efficiency. An important area within tissue engineering is the application of multipotent mesenchymal stromal cells (MSCs) to reparative surgery. The combined use of biodegradable membranes with stem cell therapy may lead to promising results for patients undergoing unsuccessful conventional treatments. Thus, the aim of this study was to test the efficacy of using membranes composed of anionic collagen with or without the addition of hyaluronic acid (HA) as a substrate for adhesion and in vitro differentiation of bone marrow-derived canine MSCs. The benefit of basic fibroblast growth factor (bFGF) on the differentiation of cells in culture was also tested. MSCs were collected from dog bone marrow, isolated and grown on collagen scaffolds with or without HA. Cell viability, proliferation rate, and cellular toxicity were analyzed after 7 days. The cultured cells showed uniform growth and morphological characteristics of undifferentiated MSCs, which demonstrated that MSCs successfully adapted to the culture conditions established by collagen scaffolds with or without HA. This demonstrates that such scaffolds are promising for applications to tissue regeneration. bFGF significantly increased the proliferative rate of MSCs by 63% when compared to groups without the addition of the growth factor. However, the addition of bFGF becomes limiting, since it has an inhibitory effect at high concentrations in culture medium.

A biomimetic strategy to form calcium phosphate crystals on type I collagen substrate

Energy Technology Data Exchange (ETDEWEB)

Xu Zhang [Department of Restorative Dentistry, Faculty of Dentistry, National University of Singapore, 5 Lower Kent Ridge Road 119074, Singapore (Singapore); Neoh, Koon Gee [Department of Chemical and Biomolecular Engineering, National University of Singapore, Kent Ridge 119260, Singapore (Singapore); Kishen, Anil, E-mail: anil.kishen@utoronto.ca [Discipline of Endodontics, Faculty of Dentistry, University of Toronto, 124 Edward Street, Toronto, ON (Canada)

2010-07-20

Objective: The aim of this study is to induce mineralization of collagen by introducing phosphate groups onto type I collagen from eggshell membrane (ESM) by treating with sodium trimetaphosphate (STMP). This strategy is based on the hypothesis that phosphate groups introduced on collagen can mimic the nucleating role of phosphorylated non-collagenous proteins bound to collagen for inducing mineralization in natural hard tissue. Method: The collagen membrane was phosphorylated by treating it with a solution of STMP and saturated calcium hydroxide. The phosphorylated collagen was subsequently exposed to a mineralization solution and the pattern of mineralization on the surface of phosphorylated collagen substrate was analyzed. Fourier-transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), field emission electron microscopy (FESEM), energy-dispersive X-ray spectroscopy (EDX), X-ray diffraction (XRD) and microhardness test were used to characterize the collagen substrate and the pattern of minerals formed on the collagen surface. Results: The FTIR and EDX results indicated that the phosphate groups were incorporated onto the collagen surface by treatment with STMP. During the mineralization process, the plate-like mineral, octacalcium phosphate (OCP), which was initially formed on the surface of ESM, was later transformed into needle-like hydroxyapatite (HAP) as indicated by the SEM, FESEM, EDX and XRD findings. The microhardness test displayed significant increase in the Knoop hardness number of the mineralized collagen. Conclusions: Phosphate groups can be introduced onto type I collagen surface by treating it with STMP and such phosphorylated collagen can induce the mineralization of type I collagen.

Neuronal Differentiation Modulated by Polymeric Membrane Properties.

Science.gov (United States)

Morelli, Sabrina; Piscioneri, Antonella; Drioli, Enrico; De Bartolo, Loredana

2017-01-01

In this study, different collagen-blend membranes were successfully constructed by blending collagen with chitosan (CHT) or poly(lactic-co-glycolic acid) (PLGA) to enhance their properties and thus create new biofunctional materials with great potential use for neuronal tissue engineering and regeneration. Collagen blending strongly affected membrane properties in the following ways: (i) it improved the surface hydrophilicity of both pure CHT and PLGA membranes, (ii) it reduced the stiffness of CHT membranes, but (iii) it did not modify the good mechanical properties of PLGA membranes. Then, we investigated the effect of the different collagen concentrations on the neuronal behavior of the membranes developed. Morphological observations, immunocytochemistry, and morphometric measures demonstrated that the membranes developed, especially CHT/Col30, PLGA, and PLGA/Col1, provided suitable microenvironments for neuronal growth owing to their enhanced properties. The most consistent neuronal differentiation was obtained in neurons cultured on PLGA-based membranes, where a well-developed neuronal network was achieved due to their improved mechanical properties. Our findings suggest that tensile strength and elongation at break are key material parameters that have potential influence on both axonal elongation and neuronal structure and organization, which are of fundamental importance for the maintenance of efficient neuronal growth. Hence, our study has provided new insights regarding the effects of membrane mechanical properties on neuronal behavior, and thus it may help to design and improve novel instructive biomaterials for neuronal tissue engineering. © 2017 S. Karger AG, Basel.

Tumor cell invasion of collagen matrices requires coordinate lipid agonist-induced G-protein and membrane-type matrix metalloproteinase-1-dependent signaling

Directory of Open Access Journals (Sweden)

Anthis Nicholas J

2006-12-01

Full Text Available Abstract Background Lysophosphatidic acid (LPA and sphingosine 1-phosphate (S1P are bioactive lipid signaling molecules implicated in tumor dissemination. Membrane-type matrix metalloproteinase 1 (MT1-MMP is a membrane-tethered collagenase thought to be involved in tumor invasion via extracellular matrix degradation. In this study, we investigated the molecular requirements for LPA- and S1P-regulated tumor cell migration in two dimensions (2D and invasion of three-dimensional (3D collagen matrices and, in particular, evaluated the role of MT1-MMP in this process. Results LPA stimulated while S1P inhibited migration of most tumor lines in Boyden chamber assays. Conversely, HT1080 fibrosarcoma cells migrated in response to both lipids. HT1080 cells also markedly invaded 3D collagen matrices (~700 μm over 48 hours in response to either lipid. siRNA targeting of LPA1 and Rac1, or S1P1, Rac1, and Cdc42 specifically inhibited LPA- or S1P-induced HT1080 invasion, respectively. Analysis of LPA-induced HT1080 motility on 2D substrates vs. 3D matrices revealed that synthetic MMP inhibitors markedly reduced the distance (~125 μm vs. ~45 μm and velocity of invasion (~0.09 μm/min vs. ~0.03 μm/min only when cells navigated 3D matrices signifying a role for MMPs exclusively in invasion. Additionally, tissue inhibitors of metalloproteinases (TIMPs-2, -3, and -4, but not TIMP-1, blocked lipid agonist-induced invasion indicating a role for membrane-type (MT-MMPs. Furthermore, MT1-MMP expression in several tumor lines directly correlated with LPA-induced invasion. HEK293s, which neither express MT1-MMP nor invade in the presence of LPA, were transfected with MT1-MMP cDNA, and subsequently invaded in response to LPA. When HT1080 cells were seeded on top of or within collagen matrices, siRNA targeting of MT1-MMP, but not other MMPs, inhibited lipid agonist-induced invasion establishing a requisite role for MT1-MMP in this process. Conclusion LPA is a

In vitro cultivation of canine multipotent mesenchymal stromal cells on collagen membranes treated with hyaluronic acid for cell therapy and tissue regeneration

Directory of Open Access Journals (Sweden)

T.I. Wodewotzky

2012-12-01

Full Text Available Support structures for dermal regeneration are composed of biodegradable and bioresorbable polymers, animal skin or tendons, or are bacteria products. The use of such materials is controversial due to their low efficiency. An important area within tissue engineering is the application of multipotent mesenchymal stromal cells (MSCs to reparative surgery. The combined use of biodegradable membranes with stem cell therapy may lead to promising results for patients undergoing unsuccessful conventional treatments. Thus, the aim of this study was to test the efficacy of using membranes composed of anionic collagen with or without the addition of hyaluronic acid (HA as a substrate for adhesion and in vitro differentiation of bone marrow-derived canine MSCs. The benefit of basic fibroblast growth factor (bFGF on the differentiation of cells in culture was also tested. MSCs were collected from dog bone marrow, isolated and grown on collagen scaffolds with or without HA. Cell viability, proliferation rate, and cellular toxicity were analyzed after 7 days. The cultured cells showed uniform growth and morphological characteristics of undifferentiated MSCs, which demonstrated that MSCs successfully adapted to the culture conditions established by collagen scaffolds with or without HA. This demonstrates that such scaffolds are promising for applications to tissue regeneration. bFGF significantly increased the proliferative rate of MSCs by 63% when compared to groups without the addition of the growth factor. However, the addition of bFGF becomes limiting, since it has an inhibitory effect at high concentrations in culture medium.

Long-term outcome of anti-glomerular basement membrane antibody disease treated with immunoadsorption.

Science.gov (United States)

Biesenbach, Peter; Kain, Renate; Derfler, Kurt; Perkmann, Thomas; Soleiman, Afschin; Benharkou, Alexandra; Druml, Wilfred; Rees, Andrew; Säemann, Marcus D

2014-01-01

Anti-glomerular basement membrane (GBM) antibody disease may lead to acute crescentic glomerulonephritis with poor renal prognosis. Current therapy favours plasma exchange (PE) for removal of pathogenic antibodies. Immunoadsorption (IAS) is superior to PE regarding efficiency of antibody-removal and safety. Apart from anecdotal data, there is no systemic analysis of the long-term effects of IAS on anti-GBM-disease and antibody kinetics. To examine the long-term effect of high-frequency IAS combined with standard immunosuppression on patient and renal survival in patients with anti-GBM-disease and to quantify antibody removal and kinetics through IAS. Retrospective review of patients treated with IAS for anti-GBM-antibody disease confirmed by biopsy and/or anti-GBM-antibodies. University Hospital of Vienna, Austria. 10 patients with anti-GBM-disease treated with IAS. Patient and renal survival, renal histology, anti-GBM-antibodies. Anti-GBM-antibodies were reduced by the first 9 IAS treatments (mean number of 23) to negative levels in all patients. Renal survival was 40% at diagnosis, 70% after the end of IAS, 63% after one year and 50% at the end of observation (mean 84 months, range 9 to 186). Dialysis dependency was successfully reversed in three of six patients. Patient survival was 90% at the end of observation. IAS efficiently eliminates anti-GBM-antibodies suggesting non-inferiority to PE with regard to renal and patient survival. Hence IAS should be considered as a valuable treatment option for anti-GBM-disease, especially in patients presenting with a high percentage of crescents and dialysis dependency due to an unusual high proportion of responders.

Long-term outcome of anti-glomerular basement membrane antibody disease treated with immunoadsorption.

Directory of Open Access Journals (Sweden)

Peter Biesenbach

Full Text Available Anti-glomerular basement membrane (GBM antibody disease may lead to acute crescentic glomerulonephritis with poor renal prognosis. Current therapy favours plasma exchange (PE for removal of pathogenic antibodies. Immunoadsorption (IAS is superior to PE regarding efficiency of antibody-removal and safety. Apart from anecdotal data, there is no systemic analysis of the long-term effects of IAS on anti-GBM-disease and antibody kinetics.To examine the long-term effect of high-frequency IAS combined with standard immunosuppression on patient and renal survival in patients with anti-GBM-disease and to quantify antibody removal and kinetics through IAS.Retrospective review of patients treated with IAS for anti-GBM-antibody disease confirmed by biopsy and/or anti-GBM-antibodies.University Hospital of Vienna, Austria.10 patients with anti-GBM-disease treated with IAS.Patient and renal survival, renal histology, anti-GBM-antibodies.Anti-GBM-antibodies were reduced by the first 9 IAS treatments (mean number of 23 to negative levels in all patients. Renal survival was 40% at diagnosis, 70% after the end of IAS, 63% after one year and 50% at the end of observation (mean 84 months, range 9 to 186. Dialysis dependency was successfully reversed in three of six patients. Patient survival was 90% at the end of observation.IAS efficiently eliminates anti-GBM-antibodies suggesting non-inferiority to PE with regard to renal and patient survival. Hence IAS should be considered as a valuable treatment option for anti-GBM-disease, especially in patients presenting with a high percentage of crescents and dialysis dependency due to an unusual high proportion of responders.

Laminin-332 alters connexin profile, dye coupling and intercellular Ca2+ waves in ciliated tracheal epithelial cells

Directory of Open Access Journals (Sweden)

Olsen Colin E

2006-08-01

Full Text Available Abstract Background Tracheal epithelial cells are anchored to a dynamic basement membrane that contains a variety of extracellular matrix proteins including collagens and laminins. During development, wound repair and disease of the airway epithelium, significant changes in extracellular matrix proteins may directly affect cell migration, differentiation and events mediated by intercellular communication. We hypothesized that alterations in cell matrix, specifically type I collagen and laminin α3β3γ2 (LM-332 proteins within the matrix, directly affect intercellular communication in ciliated rabbit tracheal epithelial cells (RTEC. Methods Functional coupling of RTEC was monitored by microinjection of the negatively charged fluorescent dyes, Lucifer Yellow and Alexa 350, into ciliated RTEC grown on either a LM-332/collagen or collagen matrix. Coupling of physiologically significant molecules was evaluated by the mechanism and extent of propagated intercellular Ca2+ waves. Expression of connexin (Cx mRNA and proteins were assayed by reverse transcriptase – polymerase chain reaction and immunocytochemistry, respectively. Results When compared to RTEC grown on collagen alone, RTEC grown on LM-332/collagen displayed a significant increase in dye transfer. Although mechanical stimulation of RTEC grown on either LM-332/collagen or collagen alone resulted in intercellular Ca2+ waves, the mechanism of transfer was dependent on matrix: RTEC grown on LM-332/collagen propagated Ca2+waves via extracellular purinergic signaling whereas RTEC grown on collagen used gap junctions. Comparison of RTEC grown on collagen or LM-332/collagen matrices revealed a reorganization of Cx26, Cx43 and Cx46 proteins. Conclusion Alterations in airway basement membrane proteins such as LM-332 can induce connexin reorganizations and result in altered cellular communication mechanisms that could contribute to airway tissue function.

Viruses in the Oceanic Basement.

Science.gov (United States)

Nigro, Olivia D; Jungbluth, Sean P; Lin, Huei-Ting; Hsieh, Chih-Chiang; Miranda, Jaclyn A; Schvarcz, Christopher R; Rappé, Michael S; Steward, Grieg F

2017-03-07

Microbial life has been detected well into the igneous crust of the seafloor (i.e., the oceanic basement), but there have been no reports confirming the presence of viruses in this habitat. To detect and characterize an ocean basement virome, geothermally heated fluid samples (ca. 60 to 65°C) were collected from 117 to 292 m deep into the ocean basement using seafloor observatories installed in two boreholes (Integrated Ocean Drilling Program [IODP] U1362A and U1362B) drilled in the eastern sediment-covered flank of the Juan de Fuca Ridge. Concentrations of virus-like particles in the fluid samples were on the order of 0.2 × 10 5 to 2 × 10 5  ml -1 ( n = 8), higher than prokaryote-like cells in the same samples by a factor of 9 on average (range, 1.5 to 27). Electron microscopy revealed diverse viral morphotypes similar to those of viruses known to infect bacteria and thermophilic archaea. An analysis of virus-like sequences in basement microbial metagenomes suggests that those from archaeon-infecting viruses were the most common (63 to 80%). Complete genomes of a putative archaeon-infecting virus and a prophage within an archaeal scaffold were identified among the assembled sequences, and sequence analysis suggests that they represent lineages divergent from known thermophilic viruses. Of the clustered regularly interspaced short palindromic repeat (CRISPR)-containing scaffolds in the metagenomes for which a taxonomy could be inferred (163 out of 737), 51 to 55% appeared to be archaeal and 45 to 49% appeared to be bacterial. These results imply that the warmed, highly altered fluids in deeply buried ocean basement harbor a distinct assemblage of novel viruses, including many that infect archaea, and that these viruses are active participants in the ecology of the basement microbiome. IMPORTANCE The hydrothermally active ocean basement is voluminous and likely provided conditions critical to the origins of life, but the microbiology of this vast habitat is not

Expression of Haemophilus ducreyi collagen binding outer membrane protein NcaA is required for virulence in swine and human challenge models of chancroid.

Science.gov (United States)

Fulcher, Robert A; Cole, Leah E; Janowicz, Diane M; Toffer, Kristen L; Fortney, Kate R; Katz, Barry P; Orndorff, Paul E; Spinola, Stanley M; Kawula, Thomas H

2006-05-01

Haemophilus ducreyi, the etiologic agent of the sexually transmitted genital ulcer disease chancroid, has been shown to associate with dermal collagen fibers within infected skin lesions. Here we describe NcaA, a previously uncharacterized outer membrane protein that is important for H. ducreyi collagen binding and host colonization. An H. ducreyi strain lacking the ncaA gene was impaired in adherence to type I collagen but not fibronectin (plasma or cellular form) or heparin. The mutation had no effect on serum resistance or binding to HaCaT keratinocytes or human foreskin fibroblasts in vitro. Escherichia coli expressing H. ducreyi NcaA bound to type I collagen, demonstrating that NcaA is sufficient to confer collagen attachment. The importance of NcaA in H. ducreyi pathogenesis was assessed using both swine and human experimental models of chancroid. In the swine model, 20% of lesions from sites inoculated with the ncaA mutant were culture positive for H. ducreyi 7 days after inoculation, compared to 73% of wild-type-inoculated sites. The average number of CFU recovered from mutant-inoculated lesions was also significantly reduced compared to that recovered from wild-type-inoculated sites at both 2 and 7 days after inoculation. In the human challenge model, 8 of 30 sites inoculated with wild-type H. ducreyi progressed to the pustular stage, compared to 0 of 30 sites inoculated with the ncaA mutant. Together these results demonstrate that the collagen binding protein NcaA is required for H. ducreyi infection.

Role of collagens and perlecan in microvascular stability: exploring the mechanism of capillary vessel damage by snake venom metalloproteinases.

Directory of Open Access Journals (Sweden)

Teresa Escalante

Full Text Available Hemorrhage is a clinically important manifestation of viperid snakebite envenomings, and is induced by snake venom metalloproteinases (SVMPs. Hemorrhagic and non-hemorrhagic SVMPs hydrolyze some basement membrane (BM and associated extracellular matrix (ECM proteins. Nevertheless, only hemorrhagic SVMPs are able to disrupt microvessels; the mechanisms behind this functional difference remain largely unknown. We compared the proteolytic activity of the hemorrhagic P-I SVMP BaP1, from the venom of Bothrops asper, and the non-hemorrhagic P-I SVMP leucurolysin-a (leuc-a, from the venom of Bothrops leucurus, on several substrates in vitro and in vivo, focusing on BM proteins. When incubated with Matrigel, a soluble extract of BM, both enzymes hydrolyzed laminin, nidogen and perlecan, albeit BaP1 did it at a faster rate. Type IV collagen was readily digested by BaP1 while leuc-a only induced a slight hydrolysis. Degradation of BM proteins in vivo was studied in mouse gastrocnemius muscle. Western blot analysis of muscle tissue homogenates showed a similar degradation of laminin chains by both enzymes, whereas nidogen was cleaved to a higher extent by BaP1, and perlecan and type IV collagen were readily digested by BaP1 but not by leuc-a. Immunohistochemistry of muscle tissue samples showed a decrease in the immunostaining of type IV collagen after injection of BaP1, but not by leuc-a. Proteomic analysis by LC/MS/MS of exudates collected from injected muscle revealed higher amounts of perlecan, and types VI and XV collagens, in exudates from BaP1-injected tissue. The differences in the hemorrhagic activity of these SVMPs could be explained by their variable ability to degrade key BM and associated ECM substrates in vivo, particularly perlecan and several non-fibrillar collagens, which play a mechanical stabilizing role in microvessel structure. These results underscore the key role played by these ECM components in the mechanical stability of

Protein nanocoatings on synthetic polymeric nanofibrous membranes designed as carriers for skin cells.

Science.gov (United States)

Bacakova, Marketa; Pajorova, Julia; Stranska, Denisa; Hadraba, Daniel; Lopot, Frantisek; Riedel, Tomas; Brynda, Eduard; Zaloudkova, Margit; Bacakova, Lucie

2017-01-01

Protein-coated resorbable synthetic polymeric nanofibrous membranes are promising for the fabrication of advanced skin substitutes. We fabricated electrospun polylactic acid and poly(lactide- co -glycolic acid) nanofibrous membranes and coated them with fibrin or collagen I. Fibronectin was attached to a fibrin or collagen nanocoating, in order further to enhance the cell adhesion and spreading. Fibrin regularly formed a coating around individual nanofibers in the membranes, and also formed a thin noncontinuous nanofibrous mesh on top of the membranes. Collagen also coated most of the fibers of the membrane and randomly created a soft gel on the membrane surface. Fibronectin predominantly adsorbed onto a thin fibrin mesh or a collagen gel, and formed a thin nanofibrous structure. Fibrin nanocoating greatly improved the attachment, spreading, and proliferation of human dermal fibroblasts, whereas collagen nanocoating had a positive influence on the behavior of human HaCaT keratinocytes. In addition, fibrin stimulated the fibroblasts to synthesize fibronectin and to deposit it as an extracellular matrix. Fibrin coating also showed a tendency to improve the ultimate tensile strength of the nanofibrous membranes. Fibronectin attached to fibrin or to a collagen coating further enhanced the adhesion, spreading, and proliferation of both cell types.

Magnetotelluric inversion for depth-to-basement estimation

DEFF Research Database (Denmark)

Cai, Hongzhu; Zhdanov, Michael

2015-01-01

The magnetotelluric (MT) method can be effectively applied for depth-to-basement estimation, because there exists a strong contrast in resistivity between a conductive sedimentary basin and a resistive crystalline basement. Conventional inversions of MT data are usually aimed at determining...... the volumetric distribution of the conductivity within the inversion domain. By the nature of the MT method, the recovered distribution of the subsurface conductivity is typically diffusive, which makes it difficult to select the sediment-basement interface. This paper develops a novel approach to 3D MT...... inversion for the depth-to-basement estimate. The key to this approach is selection of the model parameterization with the depth to basement being the major unknown parameter. In order to estimate the depth to the basement, the inversion algorithm recovers both the thickness and the conductivities...

Crosslinked basement membrane-based coatings enhance glucose sensor function and continuous glucose monitoring in vivo.

Science.gov (United States)

Klueh, Ulrike; Ludzinska, Izabela; Czajkowski, Caroline; Qiao, Yi; Kreutzer, Donald L

2018-01-01

Overcoming sensor-induced tissue reactions is an essential element of achieving successful continuous glucose monitoring (CGM) in the management of diabetes, particularly when used in closed loop technology. Recently, we demonstrated that basement membrane (BM)-based glucose sensor coatings significantly reduced tissue reactions at sites of device implantation. However, the biocompatible BM-based biohydrogel sensor coating rapidly degraded over a less than a 3-week period, which effectively eliminated the protective sensor coating. In an effort to increase the stability and effectiveness of the BM coating, we evaluated the impact of crosslinking BM utilizing glutaraldehyde as a crosslinking agent, designated as X-Cultrex. Sensor performance (nonrecalibrated) was evaluated for the impact of these X-Cultrex coatings in vitro and in vivo. Sensor performance was assessed over a 28-day time period in a murine CGM model and expressed as mean absolute relative difference (MARD) values. Tissue reactivity of Cultrex-coated, X-Cultrex-coated, and uncoated glucose sensors was evaluated over a 28-day time period in vivo using standard histological techniques. These studies demonstrated that X-Cultrex-based sensor coatings had no effect on glucose sensor function in vitro. In vivo, glucose sensor performance was significantly enhanced following X-Cultrex coating throughout the 28-day study. Histological evaluations of X-Cultrex-treated sensors demonstrated significantly less tissue reactivity when compared to uncoated sensors. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 7-16, 2018. © 2017 Wiley Periodicals, Inc.

Random/aligned electrospun PCL/PCL-collagen nanofibrous membranes: comparison of neural differentiation of rat AdMSCs and BMSCs

International Nuclear Information System (INIS)

Çapkın, Merve; Gümüşderelioğlu, Menemşe; Çakmak, Soner; Kurt, Feyzan Özdal; Şen, B Hakan; Türk, B Tuğba; Deliloğlu-Gürhan, S İsmet

2012-01-01

In this study, the aligned (A) and randomly oriented (R) polycaprolactone (PCL-A and PCL-R) and PCL/collagen (PCL/Col-A and PCL/Col-R) nanofibers were electrospun onto smooth PCL membranes (PCLMs) prepared by solvent casting. In order to investigate the effects of chemical composition and nanotopography of fibrous surfaces on proliferation and on neural differentiation of mesenchymal stem cells (MSCs), adipose and bone marrow-derived rat MSCs (AdMSCs and BMSCs) were cultivated in suitable media i.e. inducing medium containing basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF), and cell maintenance medium (CMM). BMSCs adhered and proliferated on all nanofibrous membranes more efficiently than AdMSCs. PCL/Col-A was found as the most convenient surface supporting proliferation in both cell types. Immunofluorescence staining indicated that BMSCs and AdMSCs are prone for differentiation to oligodendrocytes more than they differentiate to other neuronal cell types. PCL-A nanofibrous membranes supported differentiation of MSCs to O4 + (an oligodendrocytes surface antigen) cells in both culture media. The intensity of immunoreactivity of O4 + cells differentiated from BMSCs on PCL-A was highest when compared with the other groups (p + cells. In conclusion, this study can be evaluated to establish the cell therapy strategies in neurodegenerative disorders, which are relevant to oligodendrocyte abstinence using BMSCs or AdMSCs on aligned nanofibrous membranes. (paper)

A large-sized bubbling appearance of the glomerular basement membrane in a patient with pulmonary limited AL amyloidosis and a past history of lupus nephritis.

Science.gov (United States)

Suga, Norihiro; Miura, Naoto; Uemura, Yuko; Nakamura, Toshinobu; Morita, Hiroyuki; Banno, Shogo; Imai, Hirokazu

2011-12-01

We report an unusual pathological finding, a large-sized bubbling appearance of the glomerular basement membrane (GBM), in a patient with pulmonary limited AL amyloidosis and a past history of lupus nephritis. The first renal biopsy specimen from 10 years ago, when systemic lupus erythematosus was diagnosed, demonstrated mild mesangial proliferation and subepithelial deposits (WHO classification: III + V). Light microscopy of the current biopsy using periodic acid methenamine silver (PAMS) stain demonstrated a large-sized bubbling appearance of the GBM; however, very weak immunoglobulin and complement deposition was observed in immunofluorescence studies. Routine electron microscopy demonstrated partial subendothelial expansion with electron-lucent materials, but no electron-dense deposits or amyloid fibrils. Electron microscopy with PAMS stain revealed electron-lucent endothelial scalloping, including some cellular components and microspheres in the GBM; however, it is not clear if these materials are derived from endothelial cells. One possibility is that these unique findings represent a recovery phase of lupus membranous nephritis; another is that these findings correspond to a new disease entity.

Modulation of interferon-induced genes by lipoxin analogue in anti-glomerular basement membrane nephritis.

Science.gov (United States)

Ohse, Takamoto; Ota, Tatsuru; Kieran, Niamh; Godson, Catherine; Yamada, Koei; Tanaka, Tetsuhiro; Fujita, Toshiro; Nangaku, Masaomi

2004-04-01

Immune complex deposition is associated with the accumulation of neutrophils, which play an important role in the various immune-mediated diseases. A novel anti-inflammatory agent, the lipoxin A (LXA) analogue (15-epi-16-(FPhO)-LXA-Me)), a stable synthetic analogue of aspirin-triggered 15-epi-lipoxin A4 (ATLa), was used in experimental anti-glomerular basement membrane (GBM) antibody nephritis in mice. ATLa was administered before the induction of the disease, and 2 h later, the animals were killed. ATLa reduced the infiltrating neutrophils and nitrotyrosine staining in glomeruli. Subsequent changes of gene expression in the early phase were evaluated, and 5674 genes were present under the basal conditions in kidneys from normal mice; 54 upregulated genes and 25 downregulated genes were detected in anti-GBM nephritis. Eighteen of these upregulated genes were those induced by IFN-gamma. Real-time quantitative PCR analysis confirmed the results of the microarrays. To investigate a role of IFN-gamma in neutrophil infiltration, anti-GBM nephritis was induced in IFN-gamma knockout mice. The number of infiltrating neutrophils in these mice did not differ from those in wild-type mice. Also examined were CD11b expression on neutrophils from mice treated with ATLa by flow cytometry, but suppression of this adhesion molecule was not observed. Neutrophil infiltration was successfully inhibited by ATLa in the early phase of murine anti-GBM nephritis. Microarray analysis detected the change of mRNA expression in anti-GBM nephritis and demonstrated amelioration of various genes by ATLa, which may provide a clue to the development of novel therapeutic approaches in immune renal injury.

Effects of solid acellular type-I/III collagen biomaterials on in vitro and in vivo chondrogenesis of mesenchymal stem cells.

Science.gov (United States)

Gao, Liang; Orth, Patrick; Cucchiarini, Magali; Madry, Henning

2017-09-01

Type-I/III collagen membranes are advocated for clinical use in articular cartilage repair as being able of inducing chondrogenesis, a technique termed autologous matrix-induced chondrogenesis (AMIC). Area covered: The current in vitro and translational in vivo evidence for chondrogenic effects of solid acellular type-I/III collagen biomaterials. Expert commentary: In vitro, mesenchymal stem cells (MSCs) adhere to the fibers of the type-I/III collagen membrane. No in vitro study provides evidence that a type-I/III collagen matrix alone may induce chondrogenesis. Few in vitro studies compare the effects of type-I and type-II collagen scaffolds on chondrogenesis. Recent investigations suggest better chondrogenesis with type-II collagen scaffolds. A systematic review of the translational in vivo data identified one long-term study showing that covering of cartilage defects treated by microfracture with a type-I/III collagen membrane significantly enhanced the repair tissue volume compared with microfracture alone. Other in vivo evidence is lacking to suggest either improved histological structure or biomechanical function of the repair tissue. Taken together, there is a paucity of in vitro and preclinical in vivo evidence supporting the concept that solid acellular type-I/III collagen scaffolds may be superior to classical approaches to induce in vitro or in vivo chondrogenesis of MSCs.

Antibacterial Membrane with a Bone-Like Structure for Guided Bone Regeneration

Directory of Open Access Journals (Sweden)

YuYuan Zhang

2015-01-01

Full Text Available An antibacterial membrane with a bone-like structure was developed for guided bone regeneration (GBR by mineralising acellular bovine pericardium (ABP and loading it with the antibiotic minocycline. The bovine pericardium (BP membrane was processed using physical and chemical methods to remove the cellular components and obtain ABP membranes. Then, the ABP membranes were biomimetically mineralised using a calcium phosphate-loaded agarose hydrogel system aided by electrophoresis. Minocycline was adsorbed to the mineralised ABP membrane, and the release profile in vitro was studied. The membranes were characterised through scanning electron microscopy, diffuse reflectance-Fourier transform infrared spectroscopy, and X-ray diffraction. Results showed that the ABP membrane had an asymmetric structure with a layer of densely arranged and irregularly aligned collagen fibrils. Collagen fibrils were calcified with the formation of intrafibrillar and interfibrillar hydroxyapatites similar to the bone structure. Minocycline was incorporated into the mineralised collagen membrane and could be released in vitro. This process endowed the membrane with an antibacterial property. This novel composite membrane offers promising applications in bioactive GBR.

Viruses in the Oceanic Basement

Directory of Open Access Journals (Sweden)

Olivia D. Nigro

2017-03-01

Full Text Available Microbial life has been detected well into the igneous crust of the seafloor (i.e., the oceanic basement, but there have been no reports confirming the presence of viruses in this habitat. To detect and characterize an ocean basement virome, geothermally heated fluid samples (ca. 60 to 65°C were collected from 117 to 292 m deep into the ocean basement using seafloor observatories installed in two boreholes (Integrated Ocean Drilling Program [IODP] U1362A and U1362B drilled in the eastern sediment-covered flank of the Juan de Fuca Ridge. Concentrations of virus-like particles in the fluid samples were on the order of 0.2 × 105 to 2 × 105 ml−1 (n = 8, higher than prokaryote-like cells in the same samples by a factor of 9 on average (range, 1.5 to 27. Electron microscopy revealed diverse viral morphotypes similar to those of viruses known to infect bacteria and thermophilic archaea. An analysis of virus-like sequences in basement microbial metagenomes suggests that those from archaeon-infecting viruses were the most common (63 to 80%. Complete genomes of a putative archaeon-infecting virus and a prophage within an archaeal scaffold were identified among the assembled sequences, and sequence analysis suggests that they represent lineages divergent from known thermophilic viruses. Of the clustered regularly interspaced short palindromic repeat (CRISPR-containing scaffolds in the metagenomes for which a taxonomy could be inferred (163 out of 737, 51 to 55% appeared to be archaeal and 45 to 49% appeared to be bacterial. These results imply that the warmed, highly altered fluids in deeply buried ocean basement harbor a distinct assemblage of novel viruses, including many that infect archaea, and that these viruses are active participants in the ecology of the basement microbiome.

A Case of Fibrillary Glomerulonephritis Associated with Thrombotic Microangiopathy and Anti-Glomerular Basement Membrane Antibody

Directory of Open Access Journals (Sweden)

Akishi Momose

2015-02-01

Full Text Available We present the first report of a case of fibrillary glomerulonephritis (FGN associated with thrombotic microangiopathy (TMA and anti-glomerular basement membrane antibody (anti-GBM antibody. A 54-year-old man was admitted to our hospital for high fever and anuria. On the first hospital day, we initiated hemodialysis for renal dysfunction. Laboratory data revealed normocytic-normochromic anemia with schistocytes in the peripheral smear, thrombocytopenia, increased serum lactate dehydrogenase, decreased serum haptoglobin, and negative results for both direct and indirect Coombs tests. Based on these results, we diagnosed TMA. Assays conducted several days later indicated a disintegrin-like and metalloprotease with a thrombospondin motif 13 (ADAMTS13 activity of 31.6%, and ADAMTS13 inhibitors were negative. We started plasma exchange using fresh frozen plasma and steroid pulse therapy. Anti-GBM antibody was found to be positive. Renal biopsy showed FGN. Blood pressure rose on the 46th hospital day, and mild convulsions developed. Based on magnetic resonance imaging of the head, the patient was diagnosed with reversible posterior leukoencephalopathy syndrome. Hypertension persisted despite administration of multiple antihypertensive agents, and the patient experienced a sudden generalized seizure. Computed tomography of the head showed multiple cerebral hemorrhages. However, his blood pressure subsequently decreased and the platelet count increased. TMA remitted following 36 plasma exchange sessions, but renal function was not restored, and maintenance hemodialysis was continued. The patient was discharged on the 119th day of hospitalization. In conclusion, it was shown that TMA, FGN and anti-GBM antibody were closely related.

Expression of matrix metalloproteinase genes during basement membrane degradation in the metamorphosis of Bombyx mori.

Science.gov (United States)

Kawasaki, Hideki; Manickam, Asaithambi; Shahin, Rima; Ote, Manabu; Iwanaga, Masashi

2018-01-05

The present study was conducted to clarify the involvement of the basement membrane (BM) in insect metamorphosis through analysis of the expression profile of two types of metalloproteinase (MMP and ADAMTS) genes in several organs, their ecdysone involvement, and the histological change of BM. BM was observed around wing sac and in the wing cavity and around fat bodies at the W0 stage but disappeared after the W3 stage, and wing discs evaginated and fat body cells scattered after the W3 stage. The disappearance of the BM of midgut and silk glands was not observed after the W3 stage, but degenerated epithelium cells in the midgut and shrunken cells in the silk gland were observed after the W3 stage. BmMMP1 showed a peak at P0 in the wing discs, fat bodies, midgut, and silk gland. BmMMP2 showed a broad peak around pupation in the wing discs, fat bodies, midgut, and silk gland. BmADAMTS-1 showed enhanced expression at W2 in the wing discs, fat bodies, midgut, and hemocyte, while BmADAMTS-L showed enhanced expression at W3 in the fat bodies, midgut, silk gland, and hemocyte. After pupation, they showed a different expression in different organs. All of four genes were induced by 20-hydroxyecdysone in wing discs in vitro. The present results suggested the involvement of MMPs and ADAMTS in the BM digestion and the morphogenesis of organs during Bombyx metamorphosis. Copyright © 2017 Elsevier B.V. All rights reserved.

Bone density of defects treated with lyophilized amniotic membrane versus colagen membrane: a tomographic and histomorfogenic study in a rabbi´s femur.

Directory of Open Access Journals (Sweden)

Liz Ríos

2014-09-01

Full Text Available The aim of this study was to compare the bone density of bone defects treated with lyophilizated amniotic membrane (LAM and collagen Membrane (CM, at 3 and 5 weeks. Two bone defects of 4mm in diameter and 6mm deep were created in left distal femoral diaphysis of New Zealand rabbits (n=12. The animals were randomly divided into 2 groups. One of the defects was covered with lyophilized amniotic membrane (Rosa Chambergo Tissue Bank/National Institute of Child Health-IPEN, Lima, Peru or collagen Membrane (Dentium Co, Seoul, Korea. The second was left uncovered (NC. The rabbits were killed after 3 and 5 weeks (3 rabbits/period. The results showed a high bone density and repair of the defect by new bone. The tomographic study revealed that the bone density of the defects treated with LAM at 3 weeks was equivalent to the density obtained with CM and higher density compared with NC (p0.05. The results show that lyophilizated amniotic membrane provides bone density equal or higher to the collagen membrane.

Use of collagen gel as an alternative extracellular matrix for the in vitro and in vivo growth of murine small intestinal epithelium.

Science.gov (United States)

Jabaji, Ziyad; Sears, Connie M; Brinkley, Garrett J; Lei, Nan Ye; Joshi, Vaidehi S; Wang, Jiafang; Lewis, Michael; Stelzner, Matthias; Martín, Martín G; Dunn, James C Y

2013-12-01

Methods for the in vitro culture of primary small intestinal epithelium have improved greatly in recent years. A critical barrier for the translation of this methodology to the patient's bedside is the ability to grow intestinal stem cells using a well-defined extracellular matrix. Current methods rely on the use of Matrigel(™), a proprietary basement membrane-enriched extracellular matrix gel produced in mice that is not approved for clinical use. We demonstrate for the first time the capacity to support the long-term in vitro growth of murine intestinal epithelium in monoculture, using type I collagen. We further demonstrate successful in vivo engraftment of enteroids co-cultured with intestinal subepithelial myofibroblasts in collagen gel. Small intestinal crypts were isolated from 6 to 10 week old transgenic enhanced green fluorescent protein (eGFP+) mice and suspended within either Matrigel or collagen gel; cultures were supported using previously reported media and growth factors. After 1 week, cultures were either lysed for DNA or RNA extraction or were implanted subcutaneously in syngeneic host mice. Quantitative real-time polymerase chain reaction (qPCR) was performed to determine expansion of the transgenic eGFP-DNA and to determine the mRNA gene expression profile. Immunohistochemistry was performed on in vitro cultures and recovered in vivo explants. Small intestinal crypts reliably expanded to form enteroids in either Matrigel or collagen in both mono- and co-cultures as confirmed by microscopy and eGFP-DNA qPCR quantification. Collagen-based cultures yielded a distinct morphology with smooth enteroids and epithelial monolayer growth at the gel surface; both enteroid and monolayer cells demonstrated reactivity to Cdx2, E-cadherin, CD10, Periodic Acid-Schiff, and lysozyme. Collagen-based enteroids were successfully subcultured in vitro, whereas pure monolayer epithelial sheets did not survive passaging. Reverse transcriptase-polymerase chain reaction

Slow Muscle Precursors Lay Down a Collagen XV Matrix Fingerprint to Guide Motor Axon Navigation.

Science.gov (United States)

Guillon, Emilie; Bretaud, Sandrine; Ruggiero, Florence

2016-03-02

The extracellular matrix (ECM) provides local positional information to guide motoneuron axons toward their muscle target. Collagen XV is a basement membrane component mainly expressed in skeletal muscle. We have identified two zebrafish paralogs of the human COL15A1 gene, col15a1a and col15a1b, which display distinct expression patterns. Here we show that col15a1b is expressed and deposited in the motor path ECM by slow muscle precursors also called adaxial cells. We further demonstrate that collagen XV-B deposition is both temporally and spatially regulated before motor axon extension from the spinal cord in such a way that it remains in this region after the adaxial cells have migrated toward the periphery of the myotome. Loss- and gain-of-function experiments in zebrafish embryos demonstrate that col15a1b expression and subsequent collagen XV-B deposition and organization in the motor path ECM depend on a previously undescribed two-step mechanism involving Hedgehog/Gli and unplugged/MuSK signaling pathways. In silico analysis predicts a putative Gli binding site in the col15a1b proximal promoter. Using col15a1b promoter-reporter constructs, we demonstrate that col15a1b participates in the slow muscle genetic program as a direct target of Hedgehog/Gli signaling. Loss and gain of col15a1b function provoke pathfinding errors in primary and secondary motoneuron axons both at and beyond the choice point where axon pathway selection takes place. These defects result in muscle atrophy and compromised swimming behavior, a phenotype partially rescued by injection of a smyhc1:col15a1b construct. These reveal an unexpected and novel role for collagen XV in motor axon pathfinding and neuromuscular development. In addition to the archetypal axon guidance cues, the extracellular matrix provides local information that guides motor axons from the spinal cord to their muscle targets. Many of the proteins involved are unknown. Using the zebrafish model, we identified an

A “Mini-Epidemic” of anti-glomerular basement membrane disease: Clinical and epidemiological study

Directory of Open Access Journals (Sweden)

Umesh Lingaraj

2017-01-01

Full Text Available Acute glomerulonephritis due to anti-glomerular basement membrane (anti-GBM antibody disease is rare, estimated to occur in fewer than one case per million population and accounts for less than 20% of rapidly progressive glomerulonephritis. The prevalence among patients evaluated for potential glomerular disease is lower. It accounts for fewer than 3% of all kidney biopsies done with crescentic glomerulonephritis. Cases of anti-GBM disease occurring in a cluster have rarely been reported. All biopsy proven anti-GBM disease cases were collected from January 2015 to March 2015 at our Institute. All cases were analyzed for demographic and clinical profile, pathological findings, treatment received and for any common environmental antigenic source. A total of 11 new biopsy proven anti-GBM cases were seen within a span of three months. Age group varied from 17–80 years. Seven were males and four were females. All were dialysis dependent at presentation. Seven had active cellular crescents, and four had fibrocellular. Only one patient was a smoker and none had a history of exposure to any forms of hydrocarbons. The peak seen from January 2015 to March 2015 does not correlate with any of seasonal occurrence of infections in southern India. Although there was clustering of cases to southern territories of Karnataka state, no common etiological agents could be identified. No patient had any previous urological surgeries. All patients received methylprednisolone with plasmapheresis 5–7 sessions and cyclophosphamide. All 11 patients were dialysis dependent at the end of three months. We conclude anti-GBM disease cannot be regarded as a rare cause of renal failure and lung hemorrhage. The occurrence of such epidemic within a short period suggests a possible unidentified environmental factor like infection or occupational agents as inciting agents. Identification of such inciting agents could help us in instituting appropriate preventing measures.

Potential Development of Hydrocarbon in Basement Reservoirs In Indonesia

Directory of Open Access Journals (Sweden)

D. Sunarjanto

2014-07-01

Full Text Available DOI: 10.17014/ijog.v8i3.165Basement rocks, in particular igneous and metamorphic rocks are known to have porosity and permeability which should not be ignored. Primary porosity of basement rocks occurs as the result of rock formation. The porosity increases by the presence of cracks occurring as the result of tectonic processes (secondary porosity. Various efforts have been carried out to explore hydrocarbon in basement rocks. Some oil and gas fields proved that the basement rocks are as reservoirs which so far have provided oil and gas in significant amount. A review using previous research data, new data, and observation of igneous rocks in some fields has been done to see the development of exploration and basement reservoirs in Indonesia. A review on terminology of basement rock up till the identification of oil and gas exploration in basement rocks need to be based on the latest technology. An environmental approach is suggested to be applied as an alternative in analyzing the policy on oil and gas exploration development, especially in basement reservoirs.

Hydrogels for lung tissue engineering: Biomechanical properties of thin collagen-elastin constructs.

Science.gov (United States)

Dunphy, Siobhán E; Bratt, Jessica A J; Akram, Khondoker M; Forsyth, Nicholas R; El Haj, Alicia J

2014-10-01

In this study, collagen-elastin constructs were prepared with the aim of producing a material capable of mimicking the mechanical properties of a single alveolar wall. Collagen has been used in a wide range of tissue engineering applications; however, due to its low mechanical properties its use is limited to non load-bearing applications without further manipulation using methods such as cross-linking or mechanical compression. Here, it was hypothesised that the addition of soluble elastin to a collagen hydrogel could improve its mechanical properties. Hydrogels made from collagen only and collagen plus varying amounts elastin were prepared. Young׳s modulus of each membrane was measured using the combination of a non-destructive indentation and a theoretical model previously described. An increase in Young׳s modulus was observed with increasing concentration of elastin. The use of non-destructive indentation allowed for online monitoring of the elastic moduli of cell-seeded constructs over 8 days. The addition of lung fibroblasts into the membrane increased the stiffness of the hydrogels further and cell-seeded collagen hydrogels were found to have a stiffness equal to the theoretical value for a single alveolar wall (≈5kPa). Through provision of some of the native extracellular matrix components of the lung parenchyma these scaffolds may be able to provide an initial building block toward the regeneration of new functional lung tissue. Copyright © 2014 Elsevier Ltd. All rights reserved.

Cellular Interaction of Integrin α3β1 with Laminin 5 Promotes Gap Junctional Communication

Science.gov (United States)

Lampe, Paul D.; Nguyen, Beth P.; Gil, Susana; Usui, Marcia; Olerud, John; Takada, Yoshikazu; Carter, William G.

1998-01-01

Wounding of skin activates epidermal cell migration over exposed dermal collagen and fibronectin and over laminin 5 secreted into the provisional basement membrane. Gap junctional intercellular communication (GJIC) has been proposed to integrate the individual motile cells into a synchronized colony. We found that outgrowths of human keratinocytes in wounds or epibole cultures display parallel changes in the expression of laminin 5, integrin α3β1, E-cadherin, and the gap junctional protein connexin 43. Adhesion of keratinocytes on laminin 5, collagen, and fibronectin was found to differentially regulate GJIC. When keratinocytes were adhered on laminin 5, both structural (assembly of connexin 43 in gap junctions) and functional (dye transfer) assays showed a two- to threefold increase compared with collagen and five- to eightfold over fibronectin. Based on studies with immobilized integrin antibody and integrin-transfected Chinese hamster ovary cells, the interaction of integrin α3β1 with laminin 5 was sufficient to promote GJIC. Mapping of intermediate steps in the pathway linking α3β1–laminin 5 interactions to GJIC indicated that protein trafficking and Rho signaling were both required. We suggest that adhesion of epithelial cells to laminin 5 in the basement membrane via α3β1 promotes GJIC that integrates individual cells into synchronized epiboles. PMID:9852164

Mutation analysis of COL4A3 and COL4A4 genes in a Chinese

Indian Academy of Sciences (India)

Autosomal dominant Alport syndrome (ADAS) accounts for 5% of all cases of Alport syndrome (AS), a primary basement membrane disorder arising from mutations in genes encoding the type IV collagen protein family.Mutationsin COL4A3 and COL4A4 genes were reported to be associated with ADAS. In this study, clinical ...

Ultrastructural immunocytochemical localization of chondroitin sulfate proteoglycan in Bruch's membrane of the rat

DEFF Research Database (Denmark)

Lin, W L; Essner, E; McCarthy, K J

1992-01-01

Two monoclonal antibodies (Mab 4D5 and 2D6) raised against the core protein of a basement membrane chondroitin sulfate proteoglycan from Reichert's membrane of the rat, were used for ultrastructural immunoperoxidase localization of this protein in Bruch's membrane of the rat. Immunoreactivity...

Corneal collagen crosslinking for keratoconus. A review

Directory of Open Access Journals (Sweden)

M. M. Bikbov

2014-10-01

Full Text Available Photochemical crosslinking is widely applied in ophthalmology. Its biochemical effect is due to the release of singlet oxygen that promotes anaerobic photochemical reaction. Keratoconus is one of the most common corneal ectasia affecting 1 in 250 to 250 000 persons. Currently, the rate of iatrogenic ectasia following eximer laser refractive surgery increases due to biomechanical weakening of the cornea. Morphologically and biochemically, ectasia is characterized by corneal layers thinning, contact between the stroma and epithelium resulting from Bowman’s membrane rupture, chromatin fragmentation in keratocyte nuclei, phagocytosis, abnormal staining and arrangement of collagen fibers, enzyme system disorders, and keratocyte apoptosis. In corneal ectasia, altered enzymatic processes result in the synthesis of abnormal collagen. Collagen packing is determined by the activity of various extracellular matrix enzymes which bind amines and aldehydes of collagen fiber amino acids. In the late stage, morphological changes of Descemet’s membrane (i.e., rupture and detachment develop. Abnormal hexagonal-shaped keratocytes and their apoptosis are the signs of endothelial dystrophy. The lack of analogs in domestic ophthalmology encouraged the scientists of Ufa Eye Research Institute to develop a device for corneal collagen crosslinking. The parameters of ultraviolet (i.e., wavelength, exposure time, power to achieve the desired effect were identified. The specifics of some photosensitizers in the course of the procedure were studied. UFalink, a device for UV irradiation of cornea, and photosensitizer Dextralink were developed and adopted. Due to the high risk of endothelial damage, this treatment is contraindicated in severe keratoconus (CCT less than 400 microns. Major effects of corneal collagen crosslinking are the following: Young’s modulus (modulus of elasticity increase by 328.9 % (on average, temperature tolerance increase by 5�

Acid-Mediated Tumor Proteolysis: Contribution of Cysteine Cathepsins

Directory of Open Access Journals (Sweden)

Jennifer M Rothberg

2013-10-01

Full Text Available One of the noncellular microenvironmental factors that contribute to malignancy of solid tumors is acidic peritumoral pH. We have previously demonstrated that extracellular acidosis leads to localization of the cysteine pro-tease cathepsin B on the tumor cell membrane and its secretion. The objective of the present study was to determine if an acidic extracellular pH such as that observed in vivo (i.e., pHe 6.8 affects the activity of proteases, e.g., cathepsin B, that contribute to degradation of collagen IV by tumor cells when grown in biologically relevant three-dimensional (3D cultures. For these studies, we used 1 3D reconstituted basement membrane overlay cultures of human carcinomas, 2 live cell imaging assays to assess proteolysis, and 3 in vivo imaging of active tumor proteases. At pHe 6.8, there were increases in pericellular active cysteine cathepsins and in degradation of dye-quenched collagen IV, which was partially blocked by a cathepsin B inhibitor. Imaging probes for active cysteine cathepsins localized to tumors in vivo. The amount of bound probe decreased in tumors in bicarbonate-treated mice, a treatment previously shown to increase peritumoral pHe and reduce local invasion of the tumors. Our results are consistent with the acid-mediated invasion hypothesis and with a role for cathepsin B in promoting degradation of a basement membrane protein substrate, i.e., type IV collagen, in an acidic peritumoral environment.

Reconstruction of irradiated bone segmental defects with a biomaterial associating MBCP+(R), microstructured collagen membrane and total bone marrow grafting: an experimental study in rabbits.

Science.gov (United States)

Jégoux, Franck; Goyenvalle, Eric; Cognet, Ronan; Malard, Olivier; Moreau, Francoise; Daculsi, Guy; Aguado, Eric

2009-12-15

The bone tissue engineering models used today are still a long way from any oncologic application as immediate postimplantation irradiation would decrease their osteoinductive potential. The aim of this study was to reconstruct a segmental critical size defect in a weight-bearing bone irradiated after implantation. Six white New Zealand rabbits were immediately implanted with a biomaterial associating resorbable collagen membrane EZ(R) filled and micro-macroporous biphasic calcium phosphate granules (MBCP+(R)). After a daily schedule of radiation delivery, and within 4 weeks, a total autologous bone marrow (BM) graft was injected percutaneously into the center of the implant. All the animals were sacrificed at 16 weeks. Successful osseous colonization was found to have bridged the entire length of the defects. Identical distribution of bone ingrowth and residual ceramics at the different levels of the implant suggests that the BM graft plays an osteoinductive role in the center of the defect. Periosteum-like formation was observed at the periphery, with the collagen membrane most likely playing a role. This model succeeded in bridging a large segmental defect in weight-bearing bone with immediate postimplantation fractionated radiation delivery. This has significant implications for the bone tissue engineering approach to patients with cancer-related bone defects.

Skin Basement Membrane: The Foundation of Epidermal Integrity—BM Functions and Diverse Roles of Bridging Molecules Nidogen and Perlecan

Directory of Open Access Journals (Sweden)

Dirk Breitkreutz

2013-01-01

Full Text Available The epidermis functions in skin as first defense line or barrier against environmental impacts, resting on extracellular matrix (ECM of the dermis underneath. Both compartments are connected by the basement membrane (BM, composed of a set of distinct glycoproteins and proteoglycans. Herein we are reviewing molecular aspects of BM structure, composition, and function regarding not only (i the dermoepidermal interface but also (ii the resident microvasculature, primarily focusing on the per se nonscaffold forming components perlecan and nidogen-1 and nidogen-2. Depletion or functional deficiencies of any BM component are lethal at some stage of development or around birth, though BM defects vary between organs and tissues. Lethality problems were overcome by developmental stage- and skin-specific gene targeting or by cell grafting and organotypic (3D cocultures of normal or defective cells, which allows recapitulating BM formation de novo. Thus, evidence is accumulating that BM assembly and turnover rely on mechanical properties and composition of the adjacent ECM and the dynamics of molecular assembly, including further “minor” local components, nidogens largely functioning as catalysts or molecular adaptors and perlecan as bridging stabilizer. Collectively, orchestration of BM assembly, remodeling, and the role of individual players herein are determined by the developmental, tissue-specific, or functional context.

Ten cases of severe oral lichen planus showing granular C3 deposition in oral mucosal basement membrane zone.

Science.gov (United States)

Hashimoto, Takashi; Fukuda, Aoi; Himejima, Akio; Morita, Shosuke; Tsuruta, Daisuke; Koga, Hiroshi; Krol, Rafal P; Ishii, Norito

2015-01-01

Oral lichen planus (OLP) may show depositions of immunoglobulins and complement components in oral mucosal basement membrane zone (BMZ) in direct immunofluorescence, although these finding are not frequently seen. We collected and examined ten cases of severe OLP showing granular C3 deposition in BMZ. In addition to clinical, histopathological and direct immunofluorescence assessments, we performed various immune-serological tests, including indirect immunofluorescence of normal human skin and 1M NaCl-split skin, immunoblotting of normal human epidermal and dermal extracts, recombinant proteins of BP180 NC16a and C-terminal domains, concentrated culture supernatant of HaCaT cells and purified human laminin-332, and enzyme-linked immunosorbent assays for BP230 and BP180. Direct immunofluorescence showed C3 deposition in BMZ exclusively of granular pattern in 7 cases and of both granular and linear patterns in 3 cases. The 10 cases showed no positive reactivity for either IgG or IgA antibodies in any immuno-serological tests. Detailed analyses of clinical, histopathological and immunological findings revealed striking female prevalence, although other parameters were in general characteristic of OLP. Granular C3 deposition in oral BMZ may be one of the characteristic features of severe OLP, although mechanisms for C3 deposition and its pathogenic role in OLP are currently unknown.

Proteolysis breaks tolerance toward intact α345(IV) collagen, eliciting novel anti-GBM autoantibodies specific for α345NC1 hexamers

Science.gov (United States)

Olaru, Florina; Wang, Xu-Ping; Luo, Wentian; Ge, Linna; Miner, Jeffrey H; Kleinau, Sandra; Geiger, Xochiquetzal J.; Wasiluk, Andrew; Heidet, Laurence; Kitching, A. Richard; Borza, Dorin-Bogdan

2012-01-01

Goodpasture disease is an autoimmune kidney disease mediated by autoAbs against NC1 monomers of α3(IV) collagen that bind to the glomerular basement membrane (GBM), usually causing rapidly progressive glomerulonephritis. We identified a novel type of human IgG4-restricted anti-GBM autoAbs associated with mild non-progressive glomerulonephritis, which specifically targeted α345NC1 hexamers but not α3NC1 monomers. The mechanisms eliciting these anti-GBM autoAbs were investigated in mouse models recapitulating this phenotype. Wild type and FcγRIIB−/− mice immunized with autologous murine GBM NC1 hexamers produced mouse IgG1-restricted autoAbs specific for α345NC1 hexamers, which bound to the GBM in vivo but did not cause glomerulonephritis. In these mice, intact collagen IV from murine GBM was not immunogenic. However, in Col4a3−/− Alport mice, both intact collagen IV and NC1 hexamers from murine GBM elicited IgG antibodies specific for α3α4α5NC1 hexamers, which were not subclass restricted. As heterologous antigen in COL4A3-humanized mice, murine GBM NC1 hexamers elicited mouse IgG1, IgG2a and IgG2b autoAbs specific for α345NC1 hexamers and induced anti-GBM Ab glomerulonephritis. These findings indicate that tolerance toward autologous intact α3α4α5(IV) collagen is established in hosts expressing this antigen, even though autoreactive B cells specific for α345NC1 hexamers are not purged from their repertoire. Proteolysis selectively breaches this tolerance by generating autoimmunogenic α3α4α5NC1 hexamers. This provides a mechanism eliciting autoAbs specific for α345NC1 hexamers, which are restricted to non-inflammatory IgG subclasses and non-nephritogenic. In Alport syndrome, lack of tolerance toward α3α4α5(IV) collagen promotes production of alloantibodies to α345NC1 hexamers, including pro-inflammatory IgG subclasses which mediate post-transplant anti-GBM nephritis. PMID:23303673

Membranous nephropathy (bubbling appearance and spike formation) without immunoglobulin deposition in a patient with systemic lupus erythematosus.

Science.gov (United States)

Miura, Naoto; Mori, Yuki; Yoshino, Masabumi; Suga, Norihiro; Kitagawa, Wataru; Yamada, Harutaka; Nishikawa, Kazuhiro; Imai, Hirokazu

2008-12-01

A 53-year-old Japanese man with systemic lupus erythematosus developed proteinuria and hematuria after a urinary stone episode. A light microscopic study of a kidney biopsy specimen demonstrated a bubbling appearance and spike formation of the basement membrane. Immunofluorescent studies revealed that there were no significant depositions of immunoglobulins, such as IgG (-), IgA (-), IgM (+/-), kappa light chain (+/-), lambda light chain (+/-), or C3 (-) in the glomerular capillary wall, though C1q was present as one-plus positive staining in mesangial areas. Electron microscopic studies showed that the thickness of the basement membrane varied from thin to thick without electron dense deposits, and that the cellular components of the podocyte were irregularly present in the basement membrane. Urinary protein decreased after the usage of prednisolone and mizoribine; however, proteinuria aggravated after an episode of urinary stone during the same treatment.

Chondrogenic differentiation of mesenchymal stem cells in a leakproof collagen sponge

International Nuclear Information System (INIS)

Chen Guoping; Akahane, Daisuke; Kawazoe, Naoki; Yamamoto, Katsuyuki; Tateishi, Tetsuya

2008-01-01

A three-dimensional culture of mesenchymal stem cells (MSCs) in a porous scaffold has been developed as a promising strategy for cartilage tissue engineering. The chondrogenic differentiation of MSCs derived from human bone marrow was studied by culturing the cells in a novel scaffold constructed of leakproof collagen sponge. All the surfaces of the collagen sponge except the top were wrapped with a membrane that has pores smaller than the cells to protect against cell leakage during cell seeding. The cells adhered to the collagen, distributed evenly, and proliferated to fill the spaces in the sponge. Cell seeding efficiency was greater than 95%. The MSCs cultured in the collagen sponge in the presence of TGF-β3 and BMP6 expressed a high level of genes encoding type II and type X collagen, sox9, and aggrecan. Histological examination by HE staining indicated that the differentiated cells showed a round morphology. The extracellular matrices were positively stained by safranin O and toluidine blue. Immunostaining with anti-type II collagen and anti-cartilage proteoglycan showed that type II collagen and cartilage proteoglycan were detected around the cells. These results suggest the chondrogenic differentiation of MSCs when cultured in the collagen sponge in the presence of TGF-β3 and BMP6

Epithelial growth by rat vibrissae follicles in vitro requires mesenchymal contact via native extracellular matrix

International Nuclear Information System (INIS)

Link, R.E.; Paus, R.; Stenn, K.S.; Kuklinska, E.; Moellmann, G.

1990-01-01

An in vitro assay utilizing the rat vibrissa anagen follicle as a model for studying the epithelial-mesenchymal interactions (EMI) in hair growth is described. Through selective disruption of the epithelial-mesenchymal interface, we investigate whether the specialized extracellular matrix (ECM) of the dermal papilla and basement membrane zone (BMZ) serves a crucial function in hair follicle EMI. Epithelial bulbs incubated intact within their follicular sheaths incorporate thymidine primarily into cells of the hair matrix and outer root sheath, as shown by autoradiography. However, after removal of its mesenchymal associations (dermal papilla and extrabulbar connective tissue), the epithelial bulb showed no incorporation. Neither externally added collagen (type I or IV) nor the basement membrane components in Matrigel could substitute for the growth supporting influence of native surrounding stroma. Mechanical separation of the bulb from the dermal papilla in the basement membrane zone inhibited thymidine incorporation by the epithelium even though mesenchyme was still in close proximity. Enzymatic digestion of the dermal papilla ECM and the basal lamina by Dispase, a fibronectinase and type IV collagenase, also inhibited bulb growth without evidence of cytotoxicity. These experiments suggest that direct epithelial to mesenchymal contact is required for the support of follicular epithelial growth in vitro and that specific ECM components, possibly fibronectin and/or type IV collagen, rather than diffusable factors alone, play a crucial role in the mechanism of hair follicle EMI. The in vitro system described here provides an alternative to developmental EMI models and may serve as a valuable tool for studying EMI in the adult mammalian organism

Morphometric Changes of the Socket after Site Preservation Using Nanobone and Collagen Membrane or Stypro Versus Extraction Alone

Directory of Open Access Journals (Sweden)

Salahi S

2015-06-01

Full Text Available Statement of Problem: The long-term success of a dental implant relies on implant osseointegration into native and viable bone, implant placement in an ideal position, and optimal hard and soft tissue contour. This requires the presence of sufficient alveolar bone volume, good alveolar ridge (Practically with no sign of atrophy and good surgical technique. Objectives: The aim of this randomized controlled clinical study was to evaluate morphometric changes after different alveolar ridge preservation procedures. Materials and Methods: In this study, 33 patients who had single-rooted premolar, which needed to be extracted, were recruited. Patients were randomly divided into 3 groups and after tooth extraction the following treatments were administered: in group A: NanoBone and a collagen membrane; in group B: NanoBone and Stypro; and in group C: natural healing. The following clinical parameters were evaluated at baseline and 6 months after the extraction: buccolingual width, midbuccal height (with the use of a custom made stent and width of keratinized gingiva. For data analysis, Paired t-test,one-way ANOVA and Tukey’s tests were used. Results: The average reduction in the buccolingual width, midbuccal height and keratinized gingiva was as follows: group A: 1.18±0.6, 0.64±0.92 and 3.45±1.75 mm; group B: 2.18±0.75, 0.73±0.78 and 4.73±0.9 mm; and group C: 1±0.89, 2.36±1.21 and 5±0.63 mm, respectively. Moreover, a significantly reduced resorption was found in both the buccolingual width and the width of keratinized gingiva in group A as compared to groups B and C (p<0.05. Conclusions: This study showed that the use of collagen membrane+Nano bone (group A can significantly reduce the horizontal resorption of the alveolar ridge and keratinized tissue more effectively than stypro+Nano bone (group B and blood clot alone and natural healing (group C.

Differential actions of the endocytic collagen receptor uPARAP/Endo180 and the collagenase MMP-2 in bone homeostasis

DEFF Research Database (Denmark)

Madsen, Daniel H; Jürgensen, Henrik J; Ingvarsen, Signe

2013-01-01

A well-coordinated remodeling of uncalcified collagen matrices is a pre-requisite for bone development and homeostasis. Collagen turnover proceeds through different pathways, either involving extracellular reactions exclusively, or being dependent on endocytic processes. Extracellular collagen...... degradation requires the action of secreted or membrane attached collagenolytic proteases, whereas the alternative collagen degradation pathway proceeds intracellularly after receptor-mediated uptake and delivery to the lysosomes. In this study we have examined the functional interplay between...

Air exchange rates and migration of VOCs in basements and residences.

Science.gov (United States)

Du, L; Batterman, S; Godwin, C; Rowe, Z; Chin, J-Y

2015-12-01

Basements can influence indoor air quality by affecting air exchange rates (AERs) and by the presence of emission sources of volatile organic compounds (VOCs) and other pollutants. We characterized VOC levels, AERs, and interzonal flows between basements and occupied spaces in 74 residences in Detroit, Michigan. Flows were measured using a steady-state multitracer system, and 7-day VOC measurements were collected using passive samplers in both living areas and basements. A walk-through survey/inspection was conducted in each residence. AERs in residences and basements averaged 0.51 and 1.52/h, respectively, and had strong and opposite seasonal trends, for example, AERs were highest in residences during the summer, and highest in basements during the winter. Airflows from basements to occupied spaces also varied seasonally. VOC concentration distributions were right-skewed, for example, 90th percentile benzene, toluene, naphthalene, and limonene concentrations were 4.0, 19.1, 20.3, and 51.0 μg/m(3), respectively; maximum concentrations were 54, 888, 1117, and 134 μg/m(3). Identified VOC sources in basements included solvents, household cleaners, air fresheners, smoking, and gasoline-powered equipment. The number and type of potential VOC sources found in basements are significant and problematic, and may warrant advisories regarding the storage and use of potentially strong VOCs sources in basements. Few IAQ studies have examined basements. A sizable volume of air can flow between the basement and living area, and AERs in these two zones can differ considerably. In many residences, the basement contains significant emission sources and contributes a large fraction of VOC concentrations found in the living area. Exposures can be lowered by removing VOC sources from the basement; other exposure management options, such as local ventilation or isolation, are unlikely to be practical. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

A two-dimensional model of the colonic crypt accounting for the role of the basement membrane and pericryptal fibroblast sheath.

Directory of Open Access Journals (Sweden)

Sara-Jane Dunn

Full Text Available The role of the basement membrane is vital in maintaining the integrity and structure of an epithelial layer, acting as both a mechanical support and forming the physical interface between epithelial cells and the surrounding connective tissue. The function of this membrane is explored here in the context of the epithelial monolayer that lines the colonic crypt, test-tube shaped invaginations that punctuate the lining of the intestine and coordinate a regular turnover of cells to replenish the epithelial layer every few days. To investigate the consequence of genetic mutations that perturb the system dynamics and can lead to colorectal cancer, it must be possible to track the emerging tissue level changes that arise in the crypt. To that end, a theoretical crypt model with a realistic, deformable geometry is required. A new discrete crypt model is presented, which focuses on the interaction between cell- and tissue-level behaviour, while incorporating key subcellular components. The model contains a novel description of the role of the surrounding tissue and musculature, based upon experimental observations of the tissue structure of the crypt, which are also reported. A two-dimensional (2D cross-sectional geometry is considered, and the shape of the crypt is allowed to evolve and deform. Simulation results reveal how the shape of the crypt may contribute mechanically to the asymmetric division events typically associated with the stem cells at the base. The model predicts that epithelial cell migration may arise due to feedback between cell loss at the crypt collar and density-dependent cell division, an hypothesis which can be investigated in a wet lab. This work forms the basis for investigation of the deformation of the crypt structure that can occur due to proliferation of cells exhibiting mutant phenotypes, experiments that would not be possible in vivo or in vitro.

A co-cultured skin model based on cell support membranes

International Nuclear Information System (INIS)

Dai, N.-T.; Yeh, M.-K.; Liu, Demeral David; Adams, E.F.; Chiang, C.-H.; Yen, C.-Y.; Shih, C.-M.; Sytwu, H.-K.; Chen, Tim-Mo; Wang, H.-J.; Williamson, M.R.; Coombes, A.G.A.

2005-01-01

Tissue engineering of skin based on collagen: PCL biocomposites using a designed co-culture system is reported. The collagen: PCL biocomposites having collagen: PCL (w/w) ratios of 1:4, 1:8, and 1:20 have been proven to be biocompatible materials to support both adult normal human epidermal Keratinocyte (NHEK) and mouse 3T3 fibroblast growth in cell culture, respectively, by Dai, Coombes, et al. in 2004. Films of collagen: PCL biocomposites were prepared using non-crosslinking method by impregnation of lyophilized collagen mats with PCL/dichloromethane solutions followed by solvent evaporation. To mimic the dermal/epidermal structure of skin, the 1:20 collagen: PCL biocomposites were selected for a feasibility study of a designed co-culture technique that would subsequently be used for preparing fibroblast/biocomposite/keratinocyte skin models. A 55.3% increase in cell number was measured in the designed co-culture system when fibroblasts were seeded on both sides of a biocomposite film compared with cell culture on one surface of the biocomposite in the feasibility study. The co-culture of human keratinocytes and 3T3 fibroblasts on each side of the membrane was therefore studied using the same co-culture system by growing keratinocytes on the top surface of membrane for 3 days and 3T3 fibroblasts underneath the membrane for 6 days. Scanning electron microscopy (SEM) and immunohistochemistry assay revealed good cell attachment and proliferation of both human keratinocytes and 3T3 fibroblasts with these two types of cells isolated well on each side of the membrane. Using a modified co-culture technique, a co-cultured skin model presenting a confluent epidermal sheet on one side of the biocomposite film and fibroblasts populated on the other side of the film was developed successfully in co-culture system for 28 days under investigations by SEM and immunohistochemistry assay. Thus, the design of a co-culture system based on 1:20 (w/w) collagen: PCL biocomposite

Evaluation of autogenous PRGF+β-TCP with or without a collagen membrane on bone formation and implant osseointegration in large size bone defects. A preclinical in vivo study.

Science.gov (United States)

Batas, Leonidas; Stavropoulos, Andreas; Papadimitriou, Serafim; Nyengaard, Jens R; Konstantinidis, Antonios

2016-08-01

The aim of this study was to evaluate whether the adjunctive use of a collagen membrane enhances bone formation and implant osseointegration in non-contained defects grafted with chair-side prepared autologous platelet-rich growth factor (PRGF) adsorbed on a β-TCP particulate carrier. Large box-type defects (10 × 6 mm; W × D) were prepared in the edentulated and completely healed mandibles of six Beagles dogs. An implant with moderately rough surface was placed in the center of each defect leaving the coronal 6 mm of the implant not covered with bone. The remaining defect space was then filled out with chair-side prepared autologous PRGF adsorbed on β-TCP particles and either covered with a collagen membrane (PRGF/β-TCP+CM) (6 defects) or left without a membrane (PRGF/β-TCP) (5 defects). Histology 4 months post-op showed new lamellar and woven bone formation encompassing almost entirely the defect and limited residual β-TCP particles. Extent of osseointegration of the previously exposed portion of the implants varied, but in general was limited. Within the defect, new mineralized bone (%) averaged 43.2 ± 9.86 vs. 39.9 ± 13.7 in the PRGF/β-TCP+CM and PRGF/β-TCP group (P = 0.22) and relative mineralized bone-to-implant contact (%) averaged 26.2 ± 16.45 vs. 35.91 ± 24.45, respectively (P = 0.5). First, bone-to-implant contact from the implant top was 4.1 ± 1.5 and 3.2 ± 2.3 (P = 0.9), in the PRGF/β-TCP+CM and PRGF/β-TCP group, respectively. Implantation of chair-side prepared autologous PRGF adsorbed on a β-TCP carrier in non-contained peri-implant defects resulted in large amounts of bone regeneration, but osseointegration was limited. Provisions for GBR with a collagen membrane did not significantly enhance bone regeneration or implant osseointegration. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Frictional Behavior of Altered Basement Approaching the Nankai Trough

Science.gov (United States)

Saffer, D. M.; Ikari, M.; Rooney, T. O.; Marone, C.

2017-12-01

The frictional behavior of basement rocks plays an important role in subduction zone faulting and seismicity. This includes earthquakes seaward of the trench, large megathrust earthquakes where seamounts are subducting, or where the plate interface steps down to basement. In exhumed subduction zone rocks such as the Shimanto complex in Japan, slivers of basalt are entrained in mélange which is evidence of basement involvement in the fault system. Scientific drilling during the Nankai Trough Seismogenic Zone Experiment (NanTroSEIZE) recovered basement rock from two reference sites (C0011 and C0012) located seaward of the trench offshore the Kii Peninsula during Integrated Ocean Discovery Program (IODP) Expeditions 322 and 333. The basement rocks are pillow basalts that appear to be heterogeneously altered, resulting in contrasting dense blue material and more vesicular gray material. Major element geochemistry shows differences in silica, calcium oxides and loss-on-ignition between the two types of samples. Minor element geochemistry reveals significant differences in vanadium, chromium, and barium. X-ray diffraction on a bulk sample powder representing an average composition shows a phyllosilicate content of 20%, most of which is expandable clays. We performed laboratory friction experiments in a biaxial testing apparatus as either intact sample blocks, or as gouge powders. We combine these experiments with measurements of Pennsylvania slate for comparison, including a mixed-lithology intact block experiment. Intact Nankai basement blocks exhibit a coefficient of sliding friction of 0.73; for Nankai basement powder, slate powder, slate blocks and slate-on-basement blocks the coefficient of sliding friction ranges from 0.44 to 0.57. At slip rates ranging from 3x10-8 to 3x10-4 m/s we observe predominantly velocity-strengthening frictional behavior, indicating a tendency for stable slip. At rates of < 1x10-6 m/s some velocity-weakening was observed, specifically in

Perlecan and the Blood-Brain Barrier: Beneficial Proteolysis?

Directory of Open Access Journals (Sweden)

Jill eRoberts

2012-08-01

Full Text Available The cerebral microvasculature is important for maintaining brain homeostasis. This is achieved via the blood-brain barrier (BBB, composed of endothelial cells with specialized tight junctions, astrocytes and a basement membrane. Prominent components of the basement membrane extracellular matrix (ECM include fibronectin, laminin, collagen IV and perlecan, all of which regulate cellular processes via signal transduction through various cell membrane bound ECM receptors. Expression and proteolysis of these ECM components can be rapidly altered during pathological states of the central nervous system. In particular, proteolysis of perlecan, a heparan sulfate proteoglycan, occurs within hours following ischemia induced by experimental stroke. Proteolysis of ECM components following stroke results in the degradation of the basement membrane and further disruption of the BBB. While it is clear that such proteolysis has negative consequences for the BBB, we propose that it also may lead to generation of ECM protein fragments, including the C-terminal domain V (DV of perlecan, that potentially have a positive influence on other aspects of CNS health. Indeed, perlecan DV has been shown to be persistently generated after stroke and beneficial as a neuroprotective molecule and promoter of post-stroke brain repair. This mini-review will discuss beneficial roles of perlecan protein fragment generation within the brain during stroke.

Magnetic Basement Depth Inversion in the Space Domain

Science.gov (United States)

Nunes, Tiago Mane; Barbosa, Valéria Cristina F.; Silva, João Batista C.

2008-10-01

We present a total-field anomaly inversion method to determine both the basement relief and the magnetization direction (inclination and declination) of a 2D sedimentary basin presuming negligible sediment magnetization. Our method assumes that the magnetic intensity contrast is constant and known. We use a nonspectral approach based on approximating the vertical cross section of the sedimentary basin by a polygon, whose uppermost vertices are forced to coincide with the basin outcrop, which are presumably known. For fixed values of the x coordinates our method estimates the z coordinates of the unknown polygon vertices. To obtain the magnetization direction we assume that besides the total-field anomaly, information about the basement’s outcrops at the basin borders and the basement depths at a few points is available. To obtain stable depth-to-basement estimates we impose overall smoothness and positivity constraints on the parameter estimates. Tests on synthetic data showed that the simultaneous estimation of the irregular basement relief and the magnetization direction yields good estimates for the relief despite the mild instability in the magnetization direction. The inversion of aeromagnetic data from the onshore Almada Basin, Brazil, revealed a shallow, eastward-dipping basement basin.

Geoelectrical characterisation of basement aquifers: the case of Iberekodo, southwestern Nigeria

Science.gov (United States)

Aizebeokhai, Ahzegbobor P.; Oyeyemi, Kehinde D.

2018-03-01

Basement aquifers, which occur within the weathered and fractured zones of crystalline bedrocks, are important groundwater resources in tropical and subtropical regions. The development of basement aquifers is complex owing to their high spatial variability. Geophysical techniques are used to obtain information about the hydrologic characteristics of the weathered and fractured zones of the crystalline basement rocks, which relates to the occurrence of groundwater in the zones. The spatial distributions of these hydrologic characteristics are then used to map the spatial variability of the basement aquifers. Thus, knowledge of the spatial variability of basement aquifers is useful in siting wells and boreholes for optimal and perennial yield. Geoelectrical resistivity is one of the most widely used geophysical methods for assessing the spatial variability of the weathered and fractured zones in groundwater exploration efforts in basement complex terrains. The presented study focuses on combining vertical electrical sounding with two-dimensional (2D) geoelectrical resistivity imaging to characterise the weathered and fractured zones in a crystalline basement complex terrain in southwestern Nigeria. The basement aquifer was delineated, and the nature, extent and spatial variability of the delineated basement aquifer were assessed based on the spatial variability of the weathered and fractured zones. The study shows that a multiple-gradient array for 2D resistivity imaging is sensitive to vertical and near-surface stratigraphic features, which have hydrological implications. The integration of resistivity sounding with 2D geoelectrical resistivity imaging is efficient and enhances near-surface characterisation in basement complex terrain.

Membrane culture and reduced oxygen tension enhances cartilage matrix formation from equine cord blood mesenchymal stromal cells in vitro.

Science.gov (United States)

Co, C; Vickaryous, M K; Koch, T G

2014-03-01

Ongoing research is aimed at increasing cartilage tissue yield and quality from multipotent mesenchymal stromal cells (MSC) for the purpose of treating cartilage damage in horses. Low oxygen culture has been shown to enhance chondrogenesis, and novel membrane culture has been proposed to increase tissue yield and homogeneity. The objective of this study was to evaluate and compare the effect of reduced oxygen and membrane culture during in vitro chondrogenesis of equine cord blood (CB) MSC. CB-MSC (n = 5 foals) were expanded at 21% oxygen prior to 3-week differentiation in membrane or pellet culture at 5% and 21% oxygen. Assessment included histological examination (H&E, toluidine Blue, immunohistochemistry (IHC) for collagen type I and II), protein quantification by hydroxyproline assay and dimethylmethylene assay, and mRNA analysis for collagen IA1, collagen IIA1, collagen XA1, HIF1α and Sox9. Among treatment groups, 5% membrane culture produced neocartilage most closely resembling hyaline cartilage. Membrane culture resulted in increased wet mass, homogenous matrix morphology and an increase in total collagen content, while 5% oxygen culture resulted in higher GAG and type II collagen content. No significant differences were observed for mRNA analysis. Membrane culture at 5% oxygen produces a comparatively larger amount of higher quality neocartilage. Matrix homogeneity is attributed to a uniform diffusion gradient and reduced surface tension. Membrane culture holds promise for scale-up for therapeutic purposes, for cellular preconditioning prior to cytotherapeutic applications, and for modeling system for gas-dependent chondrogenic differentiation studies. Crown Copyright © 2014. Published by Elsevier Ltd. All rights reserved.

Geochemistry of the Puna Austral and Cordillera Oriental basement

International Nuclear Information System (INIS)

Becchio, Raul; Lucassen, Friedrich; Franz, Gerhard; Kasemann, Simone

1998-01-01

Major and trace elements, rare earths, and 143 Nd/ 147 Nd and, 147 Sm/ 144 Nd isotope ratios have been determined in the Puna Austral and Cordillera Oriental basement. The basement is formed by high temperature amphibolite facies rocks ranulites (750-550 degrees C) and green schists. They are represented by schists, paragneiss, orthogneiss, migmatites, few metabasites, marbles and chalcosilicatic banks. Hypotheses on the formation and evolution of the basement are presented

Nonlinear optical microscopy reveals invading endothelial cells anisotropically alter three-dimensional collagen matrices

International Nuclear Information System (INIS)

Lee, P.-F.; Yeh, Alvin T.; Bayless, Kayla J.

2009-01-01

The interactions between endothelial cells (ECs) and the extracellular matrix (ECM) are fundamental in mediating various steps of angiogenesis, including cell adhesion, migration and sprout formation. Here, we used a noninvasive and non-destructive nonlinear optical microscopy (NLOM) technique to optically image endothelial sprouting morphogenesis in three-dimensional (3D) collagen matrices. We simultaneously captured signals from collagen fibers and endothelial cells using second harmonic generation (SHG) and two-photon excited fluorescence (TPF), respectively. Dynamic 3D imaging revealed EC interactions with collagen fibers along with quantifiable alterations in collagen matrix density elicited by EC movement through and morphogenesis within the matrix. Specifically, we observed increased collagen density in the area between bifurcation points of sprouting structures and anisotropic increases in collagen density around the perimeter of lumenal structures, but not advancing sprout tips. Proteinase inhibition studies revealed membrane-associated matrix metalloproteinase were utilized for sprout advancement and lumen expansion. Rho-associated kinase (p160ROCK) inhibition demonstrated that the generation of cell tension increased collagen matrix alterations. This study followed sprouting ECs within a 3D matrix and revealed that the advancing structures recognize and significantly alter their extracellular environment at the periphery of lumens as they progress

NATURAL BASEMENT VENTILATION AS A RADON MITIGATION TECHNIQUE

Science.gov (United States)

The report documents a study of natural basement ventilation in two research houses during both the summer cooling season and the winter heating season. NOTE: Natural basement ventilation has always been recommended as a way to reduce radon levels in houses. However, its efficacy...

Basement domain map of the conterminous United States and Alaska

Science.gov (United States)

Lund, Karen; Box, Stephen E.; Holm-Denoma, Christopher S.; San Juan, Carma A.; Blakely, Richard J.; Saltus, Richard W.; Anderson, Eric D.; DeWitt, Ed

2015-01-01

The basement-domain map is a compilation of basement domains in the conterminous United States and Alaska designed to be used at 1:5,000,000-scale, particularly as a base layer for national-scale mineral resource assessments. Seventy-seven basement domains are represented as eighty-three polygons on the map. The domains are based on interpretations of basement composition, origin, and architecture and developed from a variety of sources. Analysis of previously published basement, lithotectonic, and terrane maps as well as models of planetary development were used to formulate the concept of basement and the methodology of defining domains that spanned the ages of Archean to present but formed through different processes. The preliminary compilations for the study areas utilized these maps, national-scale gravity and aeromagnetic data, published and limited new age and isotopic data, limited new field investigations, and conventional geologic maps. Citation of the relevant source data for compilations and the source and types of original interpretation, as derived from different types of data, are provided in supporting descriptive text and tables.

PF-1355, a mechanism-based myeloperoxidase inhibitor, prevents immune complex vasculitis and anti-glomerular basement membrane glomerulonephritis.

Science.gov (United States)

Zheng, Wei; Warner, Roscoe; Ruggeri, Roger; Su, Chunyan; Cortes, Christian; Skoura, Athanasia; Ward, Jessica; Ahn, Kay; Kalgutkar, Amit; Sun, Dexue; Maurer, Tristan S; Bonin, Paul D; Okerberg, Carlin; Bobrowski, Walter; Kawabe, Thomas; Zhang, Yanwei; Coskran, Timothy; Bell, Sammy; Kapoor, Bhupesh; Johnson, Kent; Buckbinder, Leonard

2015-05-01

Small vessel vasculitis is a life-threatening condition and patients typically present with renal and pulmonary injury. Disease pathogenesis is associated with neutrophil accumulation, activation, and oxidative damage, the latter being driven in large part by myeloperoxidase (MPO), which generates hypochlorous acid among other oxidants. MPO has been associated with vasculitis, disseminated vascular inflammation typically involving pulmonary and renal microvasculature and often resulting in critical consequences. MPO contributes to vascular injury by 1) catabolizing nitric oxide, impairing vasomotor function; 2) causing oxidative damage to lipoproteins and endothelial cells, leading to atherosclerosis; and 3) stimulating formation of neutrophil extracellular traps, resulting in vessel occlusion and thrombosis. Here we report a selective 2-thiouracil mechanism-based MPO inhibitor (PF-1355 [2-(6-(2,5-dimethoxyphenyl)-4-oxo-2-thioxo-3,4-dihydropyrimidin-1(2H)-yl)acetamide) and demonstrate that MPO is a critical mediator of vasculitis in mouse disease models. A pharmacokinetic/pharmacodynamic response model of PF-1355 exposure in relation with MPO activity was derived from mouse peritonitis. The contribution of MPO activity to vasculitis was then examined in an immune complex model of pulmonary disease. Oral administration of PF-1355 reduced plasma MPO activity, vascular edema, neutrophil recruitment, and elevated circulating cytokines. In a model of anti-glomerular basement membrane disease, formerly known as Goodpasture disease, albuminuria and chronic renal dysfunction were completely suppressed by PF-1355 treatment. This study shows that MPO activity is critical in driving immune complex vasculitis and provides confidence in testing the hypothesis that MPO inhibition will provide benefit in treating human vasculitic diseases. Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.

Different assembly of type IV collagen on hydrophilic and hydrophobic substrata alters endothelial cells interaction

Directory of Open Access Journals (Sweden)

NM Coelho

2010-06-01

Full Text Available Considering the structural role of type IV collagen (Col IV in the assembly of the basement membrane (BM and the perspective of mimicking its organization for vascular tissue engineering purposes, we studied the adsorption pattern of this protein on model hydrophilic (clean glass and hydrophobic trichloro(octadecylsilane (ODS surfaces known to strongly affect the behavior of other matrix proteins. The amount of fluorescently labeled Col IV was quantified showing saturation of the surface for concentration of the adsorbing solution of about 50μg/ml, but with approximately twice more adsorbed protein on ODS. AFM studies revealed a fine – nearly single molecular size – network arrangement of Col IV on hydrophilic glass, which turns into a prominent and growing polygonal network consisting of molecular aggregates on hydrophobic ODS. The protein layer forms within minutes in a concentration-dependent manner. We further found that human umbilical vein endothelial cells (HUVEC attach less efficiently to the aggregated Col IV (on ODS, as judged by the significantly altered cell spreading, focal adhesions formation and the development of actin cytoskeleton. Conversely, the immunofluorescence studies for integrins revealed that the fine Col IV network formed on hydrophilic substrata is better recognized by the cells via both α1 and α2 heterodimers which support cellular interaction, apart from these on hydrophobic ODS where almost no clustering of integrins was observed.

Radon mitigation experience in houses with basements and adjoining crawl spaces

International Nuclear Information System (INIS)

Messing, M.; Henschel, D.B.

1990-01-01

Active soil depressurization systems were installed in four basement houses with adjoining crawl spaces in Maryland. In addition, existing soil depressurization systems were modified in two additional basement-plus-crawl-space houses. These six houses were selected to include both good and poor communication beneath the basement slab, and different degrees of importance of the crawl space as a source of the indoor radon. The radon reduction effectiveness was compared for: depressurization only under the basement slab; depressurization only under a polyethylene liner over the unpaved crawl-space floor; and simultaneous depressurization under both the basement slab and the crawl-space liner. The objective of this paper is to identify under what conditions treatment of the basement alone might provide sufficient radon reductions in houses of this substructure, and what incremental benefits might be achieved by also treating the crawl space

Collagen fibrillogenesis: fibronectin, integrins, and minor collagens as organizers and nucleators.

Science.gov (United States)

Kadler, Karl E; Hill, Adele; Canty-Laird, Elizabeth G

2008-10-01

Collagens are triple helical proteins that occur in the extracellular matrix (ECM) and at the cell-ECM interface. There are more than 30 collagens and collagen-related proteins but the most abundant are collagens I and II that exist as D-periodic (where D = 67 nm) fibrils. The fibrils are of broad biomedical importance and have central roles in embryogenesis, arthritis, tissue repair, fibrosis, tumor invasion, and cardiovascular disease. Collagens I and II spontaneously form fibrils in vitro, which shows that collagen fibrillogenesis is a selfassembly process. However, the situation in vivo is not that simple; collagen I-containing fibrils do not form in the absence of fibronectin, fibronectin-binding and collagen-binding integrins, and collagen V. Likewise, the thin collagen II-containing fibrils in cartilage do not form in the absence of collagen XI. Thus, in vivo, cellular mechanisms are in place to control what is otherwise a protein self-assembly process. This review puts forward a working hypothesis for how fibronectin and integrins (the organizers) determine the site of fibril assembly, and collagens V and XI (the nucleators) initiate collagen fibrillogenesis.

Crucial Role of Mesangial Cell-derived Connective Tissue Growth Factor in a Mouse Model of Anti-Glomerular Basement Membrane Glomerulonephritis.

Science.gov (United States)

Toda, Naohiro; Mori, Kiyoshi; Kasahara, Masato; Ishii, Akira; Koga, Kenichi; Ohno, Shoko; Mori, Keita P; Kato, Yukiko; Osaki, Keisuke; Kuwabara, Takashige; Kojima, Katsutoshi; Taura, Daisuke; Sone, Masakatsu; Matsusaka, Taiji; Nakao, Kazuwa; Mukoyama, Masashi; Yanagita, Motoko; Yokoi, Hideki

2017-02-13

Connective tissue growth factor (CTGF) coordinates the signaling of growth factors and promotes fibrosis. Neonatal death of systemic CTGF knockout (KO) mice has hampered analysis of CTGF in adult renal diseases. We established 3 types of CTGF conditional KO (cKO) mice to investigate a role and source of CTGF in anti-glomerular basement membrane (GBM) glomerulonephritis. Tamoxifen-inducible systemic CTGF (Rosa-CTGF) cKO mice exhibited reduced proteinuria with ameliorated crescent formation and mesangial expansion in anti-GBM nephritis after induction. Although CTGF is expressed by podocytes at basal levels, podocyte-specific CTGF (pod-CTGF) cKO mice showed no improvement in renal injury. In contrast, PDGFRα promoter-driven CTGF (Pdgfra-CTGF) cKO mice, which predominantly lack CTGF expression by mesangial cells, exhibited reduced proteinuria with ameliorated histological changes. Glomerular macrophage accumulation, expression of Adgre1 and Ccl2, and ratio of M1/M2 macrophages were all reduced both in Rosa-CTGF cKO and Pdgfra-CTGF cKO mice, but not in pod-CTGF cKO mice. TGF-β1-stimulated Ccl2 upregulation in mesangial cells and macrophage adhesion to activated mesangial cells were decreased by reduction of CTGF. These results reveal a novel mechanism of macrophage migration into glomeruli with nephritis mediated by CTGF derived from mesangial cells, implicating the therapeutic potential of CTGF inhibition in glomerulonephritis.

Abnormalities in the basement membrane structure promote basal keratinocytes in the epidermis of hypertrophic scars to adopt a proliferative phenotype.

Science.gov (United States)

Yang, Shaowei; Sun, Yexiao; Geng, Zhijun; Ma, Kui; Sun, Xiaoyan; Fu, Xiaobing

2016-05-01

The majority of studies on scar formation have mainly focused on the dermis and little is known of the involvement of the epidermis. Previous research has demonstrated that the scar tissue-derived keratinocytes are different from normal cells at both the genetic and cell biological levels; however, the mechanisms responsible for the fundamental abnormalities in keratinocytes during scar development remain elusive. For this purpose, in this study, we used normal, wound edge and hypertrophic scar tissue to examine the morphological changes which occur during epidermal regeneration as part of the wound healing process and found that the histological structure of hypertrophic scar tissues differed from that of normal skin, with a significant increase in epidermal thickness. Notably, staining of the basement membrane (BM) appeared to be absent in the scar tissues. Moreover, immunofluorescence staining for cytokeratin (CK)10, CK14, CK5, CK19 and integrin-β1 indicated the differential expression of cell markers in the epidermal keratinocytes among the normal, wound edge and hypertrophic scar tissues, which corresponded with the altered BM structures. By using a panel of proteins associated with BM components, we validated our hypothesis that the BM plays a significant role in regulating the cell fate decision of epidermal keratinocytes during skin wound healing. Alterations in the structure of the BM promote basal keratinocytes to adopt a proliferative phenotype both in vivo and in vitro.

Matrix metalloproteinase-2 and its correlation with basal membrane components laminin-5 and collagen type IV in paediatric burn patients measured with Surface Plasmon Resonance Imaging (SPRI) biosensors.

Science.gov (United States)

Weremijewicz, Artur; Matuszczak, Ewa; Sankiewicz, Anna; Tylicka, Marzena; Komarowska, Marta; Tokarzewicz, Anna; Debek, Wojciech; Gorodkiewicz, Ewa; Hermanowicz, Adam

2018-01-30

The purpose of this study was the determination of matrix metalloproteinase-2 and its correlation with basal membrane components laminin-5 and collagen type IV in the blood plasma of burn patients measured with Surface Plasmon Resonance Imaging (SPRI) biosensors. 31 children scalded by hot water who were managed at the Department of Paediatric Surgery between 2014-2015, after primarily presenting with burns in 4-20% TBSA were included into the study (age 9 months up to 14 years, mean age 2,5+1 years). There were 10 girls and 21 boys. Venous blood samples were drawn 2-6h, and 12-16h after the thermal injury, and on the subsequent days 3, 5 and 7. The matrix metalloproteinase-2, collagen type IV and laminin-5 concentrations were assessed using Surface Plasmon Resonance Imaging by the investigators blinded to the other data. The MMP-2, laminin-5 and collagen type IV concentrations in the blood plasma of patients with burns, were highest 12-16h after thermal injury, the difference was statistically significant. The MMP-2, laminin-5 and collagen type IV concentrations measured 3 days, 5 days and 7 days after the thermal injury, slowly decreased over time, and on the 7th day reached the normal range, when compared with the concentration measured in controls. Current work is the first follow-up study regarding MMP-2 in burns. MMP-2, laminin-5 and collagen type IV levels were elevated early after burn injury in the plasma of studied patients, and were highest 12-16h after the injury. MMP-2, laminin-5 and collagen type IV levels were not proportional to the severity of the burn. We believe in the possibility that the gradual decrease of MMP-2, collagen type IV and laminin-5 concentrations could be connected with the process of healing, but to prove it, more investigation is needed in this area. The SPR imaging biosensor is a good diagnostic tool for determination of MMP-2, laminin-5 and collagen type IV in blood plasma of patients with burns. Copyright © 2017 Elsevier Ltd

Extreme Descemet's membrane rupture with hydrops in keratoconus: Clinical and histological manifestations

Directory of Open Access Journals (Sweden)

I-Ping Loh

2018-06-01

Full Text Available Purpose: To study the clinical and histological manifestations of an extreme Descemet's membrane rupture as a result of keratoconus. Observations: Using Periodic acid-Schiff assay to study a keratoconic cornea with an extreme rupture showed that the ruptured Descemet's membrane had retracted and folded into scrolls and ridges. The dimensions of the rupture were estimated to be 3.7mm2, and the central cornea was extremely thinned with a thickness of only 260μm. Stromal scarring and loosely packed lamellae were present anterior to the scrolls and ridges. Antibodies targetting the major components of Descemet's membrane, Laminin and type IV collagen, displayed intense labelling adjacent to the scrolls where the stroma was denuded and differential expression patterns lined the ridges. Environmental scanning electron microscopy showed possible collagen deposition at the site of rupture. Conclusions and importance: The specific staining patterns of laminin and type IV collagen suggest these components have an important role in re-endothelisation of the cornea. This is the first known report of spatial resolution of the topography of the Descemet's membrane rupture established by environmental scanning electron microscopic image montage. Keywords: Keratoconus, Descemet's membrane, Descemet's tear, Hydrops, Corneae, Histology

Basement depressurization using dwelling mechanical exhaust ventilation system

International Nuclear Information System (INIS)

Collignan, B.; O'Kelly, P.; Pilch, E.

2004-01-01

The mechanical ventilation exhaust system is commonly used in France to generate air renewal into building and especially into dwelling. It consists of a permanent mechanical air extraction from technical rooms (kitchen, bathrooms and toilets) using a unique fan connected to exhaust ducts. Natural air inlets in living room and bed rooms ensure an air flow from living spaces towards technical rooms. To fight against radon into building, the most recognised efficient technique is the Soil Depressurization System (S.D.S.) consisting in depressurizing the house basement. The aim of this study is to test the ability of the dwelling mechanical ventilation system to depressurize the basement in conjunction with air renewal of a house. For that purpose, a S.D.S. has been installed in an experimental house at CSTB during its construction. At first, tests undertaken with a variable velocity fan connected to the S.D.S. have characterised the permeability of the basement. It is shown that basement can be depressurized adequately with a relatively low air flow rate. At a second stage, S.D.S. has been connected to the exhaust ventilation fan used for the mechanical ventilation of the house. Results obtained show the ability of such ventilation system to generate sufficient depressurization in the basement and to ensure simultaneously adequate air change rate in the dwelling. (author)

Highly concentrated collagen solutions leading to transparent scaffolds of controlled three-dimensional organizations for corneal epithelial cell colonization.

Science.gov (United States)

Tidu, Aurélien; Ghoubay-Benallaoua, Djida; Teulon, Claire; Asnacios, Sophie; Grieve, Kate; Portier, François; Schanne-Klein, Marie-Claire; Borderie, Vincent; Mosser, Gervaise

2018-05-29

This study aimed at controlling both the organization and the transparency of dense collagen scaffolds making use of the lyotropic mesogen properties of collagen. Cholesteric or plywood-like liquid crystal phases were achieved using mixtures of acetic and hydrochloric acids as solvents. The critical pH at which the switch between the two phases occurred was around pH = 3. The use of the two acids led to fibrillated collagen I scaffolds, whose visual aspect ranged from opaque to transparent. Rheological investigations showed that viscoelastic properties of the plywood-like solutions were optimized for molding due to faster recovery. They also confirmed the correlation between the elastic modulus and the diameter of collagen fibrils obtained after fibrillogenesis under ammonia vapor. Human corneal epithelial cells, grown from donor limbal explants, were cultured both on transparent plywood-like matrices and on human amniotic membranes for 14 days. The development of corneal epithelium and the preservation of epithelial stem cells were checked by optical microscopy, colony formation assay, immuno-fluorescence and quantitative polymerase chain reaction. A higher level of amplification of limbal stem cells was obtained with collagen matrices compared with amniotic membranes, showing the high biocompatibility of our scaffolds. We therefore suggest that collagen solutions presenting both plywood-like organization and transparency might be of interest for biomedical applications in ophthalmology.

Grenville age of basement rocks in Cape May NJ well: New evidence for Laurentian crust in U.S. Atlantic Coastal Plain basement Chesapeake terrane

Science.gov (United States)

Sheridan, R.E.; Maguire, T.J.; Feigenson, M.D.; Patino, L.C.; Volkert, R.A.

1999-01-01

The Chesapeake terrane of the U.S. mid-Atlantic Coastal Plain basement is bounded on the northwest by the Salisbury positive gravity and magnetic anomaly and extends to the southeast as far as the Atlantic coast. It underlies the Coastal Plain of Virginia, Maryland, Delaware and southern New Jersey. Rubidium/Strontium dating of the Chesapeake terrane basement yields an age of 1.025 ?? 0.036 Ga. This age is typical of Grenville province rocks of the Middle to Late Proterozoic Laurentian continent. The basement lithologies are similar to some exposed Grenville-age rocks of the Appalachians. The TiO2 and Zr/P2O5 composition of the metagabbro from the Chesapeake terrane basement is overlapped by those of the Proterozoic mafic dikes in the New Jersey Highlands. These new findings support the interpretation that Laurentian basement extends southeast as far as the continental shelf in the U.S. mid-Atlantic region. The subcrop of Laurentian crust under the mid-Atlantic Coastal Plain implies unroofing by erosion of the younger Carolina (Avalon) supracrustal terrane. Dextral-transpression fault duplexes may have caused excessive uplift in the Salisbury Embayment area during the Alleghanian orogeny. This extra uplift in the Salisbury area may have caused the subsequent greater subsidence of the Coastal Plain basement in the embayment.

Development of the zebrafish myoseptum with emphasis on the myotendinous junction.

Science.gov (United States)

Charvet, Benjamin; Malbouyres, Marilyne; Pagnon-Minot, Aurélie; Ruggiero, Florence; Le Guellec, Dominique

2011-12-01

Zebrafish myosepta connect two adjacent muscle cells and transmit muscular forces to axial structures during swimming via the myotendinous junction (MTJ). The MTJ establishes transmembrane linkages system consisting of extracellular matrix molecules (ECM) surrounding the basement membrane, cytoskeletal elements anchored to sarcolema, and all intermediate proteins that link ECM to actin filaments. Using a series of zebrafish specimens aged between 24 h post-fertilization and 2 years old, the present paper describes at the transmission electron microscope level the development of extracellular and intracellular elements of the MTJ. The transverse myoseptum development starts during the segmentation period by deposition of sparse and loosely organized collagen fibrils. During the hatching period, a link between actin filaments and sarcolemma is established. The basal lamina underlining sarcolemma is well differentiated. Later, collagen fibrils display an orthogonal orientation and fibroblast-like cells invade the myoseptal stroma. A dense network of collagen fibrils is progressively formed that both anchor myoseptal fibroblasts and sarcolemmal basement membrane. The differentiation of a functional MTJ is achieved when sarcolemma interacts with both cytoskeletal filaments and extracellular components. This solid structural link between contractile apparatus and ECM leads to sarcolemma deformations resulting in the formation of regular invaginations, and allows force transmission during muscle contraction. This paper presents the first ultrastructural atlas of the zebrafish MTJ development, which represents an useful tool to analyse the mechanisms of the myotendinous system formation and their disruption in muscle disorders.

Radionuclide distribution in TMI-2 reactor building basement liquids and solids

International Nuclear Information System (INIS)

Horan, J.T.; McIsaac, C.V.; Keefer, D.G.

1984-01-01

As a result of the TMI-2 accident, approximately 2.46 x 10 6 L of contaminated water were released to the Reactor Building basement. The principal fission product release pathway from the damaged core was through the reactor coolant system (RCS) to the pressurizer, through the pressure-operated relief valve (PORV) on the pressurizer to the Reactor Coolant Drain Tank (RCDT), and then through the RCDT rupture disk to the Reactor Building basement. Since August 1979, a number of efforts have been made to determine the location, quantity, and composition of fission products released to the Reactor Building basement. These efforts have included sampling of the basement water and solids, the basement sump pump recirculation line, the RCDT, and visual surveys using a closed circuit television (CCTV) system. The analysis of basement samples has provided data on the physical and radioisotopic characteristics of the liquids and solids. This paper describes the sample collection techniques and discusses radiochemical analyses results

Collagenous sprue

DEFF Research Database (Denmark)

Soendergaard, Christoffer; Riis, Lene Buhl; Nielsen, Ole Haagen

2014-01-01

Collagenous sprue is a rare clinicopathological condition of the small bowel. It is characterised by abnormal subepithelial collagen deposition and is typically associated with malabsorption, diarrhoea and weight loss. The clinical features of collagenous sprue often resemble those of coeliac...... disease and together with frequent histological findings like mucosal thinning and intraepithelial lymphocytosis the diagnosis may be hard to reach without awareness of this condition. While coeliac disease is treated using gluten restriction, collagenous sprue is, however, not improved...... by this intervention. In cases of diet-refractory 'coeliac disease' it is therefore essential to consider collagenous sprue to initiate treatment at an early stage to prevent the fibrotic progression. Here, we report a case of a 78-year-old man with collagenous sprue and present the clinical and histological...

Collagencin, an antibacterial peptide from fish collagen: Activity, structure and interaction dynamics with membrane

International Nuclear Information System (INIS)

Ennaas, Nadia; Hammami, Riadh; Gomaa, Ahmed; Bédard, François; Biron, Éric; Subirade, Muriel; Beaulieu, Lucie; Fliss, Ismail

2016-01-01

In this study, we first report characterization of collagencin, an antimicrobial peptide identified from fish collagen hydrolysate. The peptide completely inhibited the growth of Staphylococcus aureus at 1.88 mM. Although non-toxic up to 470 μM, collagencin was hemolytic at higher concentrations. The secondary structure of collagencin was mainly composed by β-sheet and β-turn as determined by CD measurements and molecular dynamics. The peptide is likely to form β-sheet structure under hydrophobic environments and interacts with both anionic (phosphatidylglycerol) and zwitterionic (phosphoethanolamine and phosphatidylcholine) lipids as shown with CD spectroscopy and molecular dynamics. The peptide formed several hydrogen bonds with both POPG and POPE lipids and remained at membrane–water interface, suggesting that collagencin antibacterial action follows a carpet mechanism. Collagenous fish wastes could be processed by enzymatic hydrolysis and transformed into products of high value having functional or biological properties. Marine collagens are a promising source of antimicrobial peptides with new implications in food safety and human health. - Highlights: • Collagencin, an antibacterial (G+ & G-) peptide identified from fish collagen hydrolysate. • The peptide completely inhibited the growth of S. aureus at 1.88 mM and non-toxic at 470 μM. • The secondary structure was mainly composed by β-sheet and turn as determined by CD and MD. • Collagencin interacts with both anionic and zwitterionic lipids as shown with CD and MD. • Collagencin antibacterial action probably follows a carpet mechanism.

Collagencin, an antibacterial peptide from fish collagen: Activity, structure and interaction dynamics with membrane

Energy Technology Data Exchange (ETDEWEB)

Ennaas, Nadia [STELA Dairy Research Centre, Institute of Nutrition and Functional Foods, Université Laval, G1V 0A6 Québec, QC (Canada); Hammami, Riadh, E-mail: riadh.hammami@fsaa.ulaval.ca [STELA Dairy Research Centre, Institute of Nutrition and Functional Foods, Université Laval, G1V 0A6 Québec, QC (Canada); Gomaa, Ahmed [STELA Dairy Research Centre, Institute of Nutrition and Functional Foods, Université Laval, G1V 0A6 Québec, QC (Canada); Bédard, François; Biron, Éric [Faculty of Pharmacy, Université Laval and Laboratory of Medicinal Chemistry, CHU de Québec Research Centre, G1V 4G2 Québec, QC (Canada); Subirade, Muriel [STELA Dairy Research Centre, Institute of Nutrition and Functional Foods, Université Laval, G1V 0A6 Québec, QC (Canada); Beaulieu, Lucie, E-mail: lucie.beaulieu@fsaa.ulaval.ca [STELA Dairy Research Centre, Institute of Nutrition and Functional Foods, Université Laval, G1V 0A6 Québec, QC (Canada); Department of Biology, Chemistry and Geography, Université du Québec à Rimouski (UQAR), 300 Allée des Ursulines, Rimouski, QC G5L 3A1 (Canada); Fliss, Ismail, E-mail: ismail.fliss@fsaa.ulaval.ca [STELA Dairy Research Centre, Institute of Nutrition and Functional Foods, Université Laval, G1V 0A6 Québec, QC (Canada)

2016-04-29

In this study, we first report characterization of collagencin, an antimicrobial peptide identified from fish collagen hydrolysate. The peptide completely inhibited the growth of Staphylococcus aureus at 1.88 mM. Although non-toxic up to 470 μM, collagencin was hemolytic at higher concentrations. The secondary structure of collagencin was mainly composed by β-sheet and β-turn as determined by CD measurements and molecular dynamics. The peptide is likely to form β-sheet structure under hydrophobic environments and interacts with both anionic (phosphatidylglycerol) and zwitterionic (phosphoethanolamine and phosphatidylcholine) lipids as shown with CD spectroscopy and molecular dynamics. The peptide formed several hydrogen bonds with both POPG and POPE lipids and remained at membrane–water interface, suggesting that collagencin antibacterial action follows a carpet mechanism. Collagenous fish wastes could be processed by enzymatic hydrolysis and transformed into products of high value having functional or biological properties. Marine collagens are a promising source of antimicrobial peptides with new implications in food safety and human health. - Highlights: • Collagencin, an antibacterial (G+ & G-) peptide identified from fish collagen hydrolysate. • The peptide completely inhibited the growth of S. aureus at 1.88 mM and non-toxic at 470 μM. • The secondary structure was mainly composed by β-sheet and turn as determined by CD and MD. • Collagencin interacts with both anionic and zwitterionic lipids as shown with CD and MD. • Collagencin antibacterial action probably follows a carpet mechanism.

RESEARCH ON REDUCING PREMATURITY RUPTURE OF MEMBRANE

Directory of Open Access Journals (Sweden)

Maria URSACHI (BOLOTA

2016-12-01

Full Text Available The membranes surrounding the amniotic cavity are composed from amnion and chorion, tightly adherent layers which are composed of several cell types, including epithelial cells, trophoblasts cells and mesenchyme cells, embedded in a collagenous matrix. They retain amniotic fluid, secret substances into the amniotic fluid, as well as to the uterus and protect the fetus against upward infections from urogenital tract. Normally, the membranes it breaks during labor. Premature rupture of the amniotic sac (PRAS is defined as rupture of membranes before the onset of labor. Premature rupture of the fetal membrane, which occurs before 37 weeks of gestation, usually, refers to preterm premature rupture of membranes. Despite advances in the care period, premature rupture of membranes and premature rupture of membranes preterm continue to be regarded as serious obstetric complications. On the term 8% - 10% of pregnant women have premature rupture of membranes; these women are at increased risk of intrauterine infections, where the interval between membrane rupture and expulsion is rolled-over. Premature rupture of membranes preterm occurs in approximately 1% of all pregnancies and is associated with 30% -40% of preterm births. Thus, it is important to identify the cause of pre-term birth (after less than 37 completed weeks of "gestation" and its complications, including respiratory distress syndrome, neonatal infection and intraventricular hemorrhage. Objectives: the development of the protocol of the clinical trial on patients with impending preterm birth, study clinical and statistical on the socio-demographic characteristics of patients with imminent preterm birth; clinical condition of patients and selection of cases that could benefit from the application of interventional therapy; preclinical investigation (biological and imaging of patients with imminent preterm birth; the modality therapy; clinical investigation of the effectiveness of short

Depth-To-Basement Mapping Using Fractal Technique: Application ...

African Journals Online (AJOL)

... and can thus be obtained at source level. Application to aeromagnetic data from the Chad basin north eastern Nigeria produced a basement relief which range from depths of 2.47 km to 5.40 km with an average of 3.92 +- 0.66 km. Keywords: Fractal, depth, basement, spectra, aeromagnetic. Nigerian Journal of Physics Vol ...

60 Review Article PRETERM RUPTURE OF MEMBRANES: THE ...

African Journals Online (AJOL)

Preterm prelabour rupture of membranes (PPROM) is one of the major factors that have been found to ... is because of the multiple risk factors that have been associated with PROM and also the non-uniformity in ... collagen within the extracellular matrix of the chorioamniotic membrane induced by an increased expression.

Measure Guideline: Basement Insulation Basics

Energy Technology Data Exchange (ETDEWEB)

Aldrich, R.; Mantha, P.; Puttagunta, S.

2012-10-01

This guideline is intended to describe good practices for insulating basements in new and existing homes, and is intended to be a practical resources for building contractors, designers, and also to homeowners.

Ultrastructural and biochemical characterization of mechanically adaptable collagenous structures in the edible sea urchin Paracentrotus lividus.

Science.gov (United States)

Barbaglio, Alice; Tricarico, Serena; Ribeiro, Ana R; Di Benedetto, Cristiano; Barbato, Marta; Dessì, Desirèe; Fugnanesi, Valeria; Magni, Stefano; Mosca, Fabio; Sugni, Michela; Bonasoro, Francesco; Barbosa, Mario A; Wilkie, Iain C; Candia Carnevali, M Daniela

2015-06-01

The viscoelastic properties of vertebrate connective tissues rarely undergo significant changes within physiological timescales, the only major exception being the reversible destiffening of the mammalian uterine cervix at the end of pregnancy. In contrast to this, the connective tissues of echinoderms (sea urchins, starfish, sea cucumbers, etc.) can switch reversibly between stiff and compliant conditions in timescales of around a second to minutes. Elucidation of the molecular mechanism underlying such mutability has implications for the zoological, ecological and evolutionary field. Important information could also arise for veterinary and biomedical sciences, particularly regarding the pathological plasticization or stiffening of connective tissue structures. In the present investigation we analyzed aspects of the ultrastructure and biochemistry in two representative models, the compass depressor ligament and the peristomial membrane of the edible sea urchin Paracentrotus lividus, compared in three different mechanical states. The results provide further evidence that the mechanical adaptability of echinoderm connective tissues does not necessarily imply changes in the collagen fibrils themselves. The higher glycosaminoglycan (GAG) content registered in the peristomial membrane with respect to the compass depressor ligament suggests a diverse role of these molecules in the two mutable collagenous tissues. The possible involvement of GAG in the mutability phenomenon will need further clarification. During the shift from a compliant to a standard condition, significant changes in GAG content were detected only in the compass depressor ligament. Similarities in terms of ultrastructure (collagen fibrillar assembling) and biochemistry (two alpha chains) were found between the two models and mammalian collagen. Nevertheless, differences in collagen immunoreactivity, alpha chain migration on SDS-PAGE and BLAST alignment highlighted the uniqueness of sea urchin

Use of Clotted Human Plasma and Aprotinin in Skin Tissue Engineering: A Novel Approach to Engineering Composite Skin on a Porous Scaffold.

Science.gov (United States)

Paul, Michelle; Kaur, Pritinder; Herson, Marisa; Cheshire, Perdita; Cleland, Heather; Akbarzadeh, Shiva

2015-10-01

Tissue-engineered composite skin is a promising therapy for the treatment of chronic and acute wounds, including burns. Providing the wound bed with a dermal scaffold populated by autologous dermal and epidermal cellular components can further entice host cell infiltration and vascularization to achieve permanent wound closure in a single stage. However, the high porosity and the lack of a supportive basement membrane in most commercially available dermal scaffolds hinders organized keratinocyte proliferation and stratification in vitro and may delay re-epithelization in vivo. The objective of this study was to develop a method to enable the in vitro production of a human skin equivalent (HSE) that included a porous scaffold and dermal and epidermal cells expanded ex vivo, with the potential to be used for definitive treatment of skin defects in a single procedure. A collagen-glycosaminoglycan dermal scaffold (Integra(®)) was populated with adult fibroblasts. A near-normal skin architecture was achieved by the addition of coagulated human plasma to the fibroblast-populated scaffold before seeding cultured keratinocytes. This resulted in reducing scaffold pore size and improving contact surfaces. Skin architecture and basement membrane formation was further improved by the addition of aprotinin (a serine protease inhibitor) to the culture media to inhibit premature clot digestion. Histological assessment of the novel HSE revealed expression of keratin 14 and keratin 10 similar to native skin, with a multilayered neoepidermis morphologically comparable to human skin. Furthermore, deposition of collagen IV and laminin-511 were detected by immunofluorescence, indicating the formation of a continuous basement membrane at the dermal-epidermal junction. The proposed method was efficient in producing an in vitro near native HSE using the chosen off-the-shelf porous scaffold (Integra). The same principles and promising outcomes should be applicable to other biodegradable

The preosteoblast response of electrospinning PLGA/PCL nanofibers: effects of biomimetic architecture and collagen I

Directory of Open Access Journals (Sweden)

Qian YZ

2016-08-01

Full Text Available Yunzhu Qian,1,2 Hanbang Chen,1 Yang Xu,1 Jianxin Yang,2 Xuefeng Zhou,3 Feimin Zhang,1 Ning Gu3 1Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University, Nanjing, 2Center of Stomatology, The Second Affiliated Hospital of Soochow University, Suzhou, 3School of Biological Science and Medical Engineering, Southeast University, Nanjing, People’s Republic of China Abstract: Constructing biomimetic structure and incorporating bioactive molecules is an effective strategy to achieve a more favorable cell response. To explore the effect of electrospinning (ES nanofibrous architecture and collagen I (COL I-incorporated modification on tuning osteoblast response, a resorbable membrane composed of poly(lactic-co-glycolic acid/poly(caprolactone (PLGA/PCL; 7:3 w/w was developed via ES. COL I was blended into PLGA/PCL solution to prepare composite ES membrane. Notably, relatively better cell response was delivered by the bioactive ES-based membrane which was fabricated by modification of 3,4-dihydroxyphenylalanine and COL I. After investigation by field emission scanning electron microscopy, Fourier transform infrared spectroscopy, contact angle measurement, and mechanical test, polyporous three-dimensional nanofibrous structure with low tensile force and the successful integration of COL I was obtained by the ES method. Compared with traditional PLGA/PCL membrane, the surface hydrophilicity of collagen-incorporated membranes was largely enhanced. The behavior of mouse preosteoblast MC3T3-E1 cell infiltration and proliferation on membranes was studied at 24 and 48 hours. The negative control was fabricated by solvent casting. Evaluation of cell adhesion and morphology demonstrated that all the ES membranes were more favorable for promoting the cell adhesion and spreading than the casting membrane. Cell Counting Kit-8 assays revealed that biomimetic architecture, surface topography, and bioactive properties of membranes were favorable for cell

Nonparenchymal cells cultivated from explants of fibrotic liver resemble endothelial and smooth muscle cells from blood vessel walls

International Nuclear Information System (INIS)

Voss, B.; Rauterberg, J.; Pott, G.; Brehmer, U.; Allam, S.; Lehmann, R.; von Bassewitz, D.B.

1982-01-01

Tissue specimens from human fibrotic liver obtained by needle biopsy were cultured. Two cell types emerged from the tissue explants. From their morphology and biosynthetic products they resembled smooth muscle cells and endothelial cells from blood vessel walls. In the endothelial cells, factor VIII-associated protein was demonstrated by indirect immunofluorescence. Synthesis of collagen types I and III, basement membrane collagen types IV and V, and fibronectin by both cell types was observed by immunofluorescence microscopy. Homogeneous cultures of smooth muscle cells were observed in subcultures. After incubation with [ 14 C]glycine, collagen was isolated and characterized by CM cellulose chromatography, and consisted mainly of types I and III. These data suggest involvement of mesenchymal cells in hepatic fibrosis; they presumably originate from blood vessel or sinusoidal walls

Transmembrane collagen XVII modulates integrin dependent keratinocyte migration via PI3K/Rac1 signaling.

Directory of Open Access Journals (Sweden)

Stefanie Löffek

Full Text Available The hemidesmosomal transmembrane component collagen XVII (ColXVII plays an important role in the anchorage of the epidermis to the underlying basement membrane. However, this adhesion protein seems to be also involved in the regulation of keratinocyte migration, since its expression in these cells is strongly elevated during reepithelialization of acute wounds and in the invasive front of squamous cell carcinoma, while its absence in ColXVII-deficient keratinocytes leads to altered cell motility. Using a genetic model of murine Col17a1⁻/⁻ keratinocytes we elucidated ColXVII mediated signaling pathways in cell adhesion and migration. Col17a1⁻/⁻ keratinocytes exhibited increased spreading on laminin 332 and accelerated, but less directed cell motility. These effects were accompanied by increased expression of the integrin subunits β4 and β1. The migratory phenotype, as evidenced by formation of multiple unstable lamellipodia, was associated with enhanced phosphoinositide 3-kinase (PI3K activity. Dissection of the signaling pathway uncovered enhanced phosphorylation of the β4 integrin subunit and the focal adhesion kinase (FAK as activators of PI3K. This resulted in elevated Rac1 activity as a downstream consequence. These results provide mechanistic evidence that ColXVII coordinates keratinocyte adhesion and directed motility by interfering integrin dependent PI3K activation and by stabilizing lamellipodia at the leading edge of reepithelializing wounds and in invasive squamous cell carcinoma.

Effects of 3D-Printed Polycaprolactone/β-Tricalcium Phosphate Membranes on Guided Bone Regeneration.

Science.gov (United States)

Shim, Jin-Hyung; Won, Joo-Yun; Park, Jung-Hyung; Bae, Ji-Hyeon; Ahn, Geunseon; Kim, Chang-Hwan; Lim, Dong-Hyuk; Cho, Dong-Woo; Yun, Won-Soo; Bae, Eun-Bin; Jeong, Chang-Mo; Huh, Jung-Bo

2017-04-25

This study was conducted to compare 3D-printed polycaprolactone (PCL) and polycaprolactone/β-tricalcium phosphate (PCL/β-TCP) membranes with a conventional commercial collagen membrane in terms of their abilities to facilitate guided bone regeneration (GBR). Fabricated membranes were tested for dry and wet mechanical properties. Fibroblasts and preosteoblasts were seeded into the membranes and rates and patterns of proliferation were analyzed using a kit-8 assay and by scanning electron microscopy. Osteogenic differentiation was verified by alizarin red S and alkaline phosphatase (ALP) staining. An in vivo experiment was performed using an alveolar bone defect beagle model, in which defects in three dogs were covered with different membranes. CT and histological analyses at eight weeks after surgery revealed that 3D-printed PCL/β-TCP membranes were more effective than 3D-printed PCL, and substantially better than conventional collagen membranes in terms of biocompatibility and bone regeneration and, thus, at facilitating GBR.

Effects of 3D-Printed Polycaprolactone/β-Tricalcium Phosphate Membranes on Guided Bone Regeneration

Directory of Open Access Journals (Sweden)

Jin-Hyung Shim

2017-04-01

Full Text Available This study was conducted to compare 3D-printed polycaprolactone (PCL and polycaprolactone/β-tricalcium phosphate (PCL/β-TCP membranes with a conventional commercial collagen membrane in terms of their abilities to facilitate guided bone regeneration (GBR. Fabricated membranes were tested for dry and wet mechanical properties. Fibroblasts and preosteoblasts were seeded into the membranes and rates and patterns of proliferation were analyzed using a kit-8 assay and by scanning electron microscopy. Osteogenic differentiation was verified by alizarin red S and alkaline phosphatase (ALP staining. An in vivo experiment was performed using an alveolar bone defect beagle model, in which defects in three dogs were covered with different membranes. CT and histological analyses at eight weeks after surgery revealed that 3D-printed PCL/β-TCP membranes were more effective than 3D-printed PCL, and substantially better than conventional collagen membranes in terms of biocompatibility and bone regeneration and, thus, at facilitating GBR.

Guided bone regeneration in rat mandibular defects using resorbable poly(trimethylene carbonate) barrier membranes

NARCIS (Netherlands)

van Leeuwen, A. C.; Huddleston Slater, J. J. R.; Gielkens, P. F. M.; de Jong, J. R.; Grijpma, D. W.; Bos, R. R. M.

The present study evaluates a new synthetic degradable barrier membrane based on poly(trimethylene carbonate) (PTMC) for use in guided bone regeneration. A collagen membrane and an expanded polytetrafluoroethylene (e-PTFE) membrane served as reference materials. In 192 male Sprague-Dawley rats, a

Guided bone regeneration in rat mandibular defects using resorbable poly(trimethylene carbonate) barrier membranes

NARCIS (Netherlands)

van Leeuwen, A.C.; Huddelston Slater, J.J.R.; Gielkens, P.F.M.; de Jong, J.R.; Grijpma, Dirk W.; Bos, R.R.M.

2012-01-01

The present study evaluates a new synthetic degradable barrier membrane based on poly(trimethylene carbonate) (PTMC) for use in guided bone regeneration. A collagen membrane and an expanded polytetrafluoroethylene (e-PTFE) membrane served as reference materials. In 192 male Sprague–Dawley rats, a

Creation of an acellular vaginal matrix for potential vaginal augmentation and cloacal repair.

Science.gov (United States)

Greco, K V; Jones, L G; Obiri-Yeboa, I; Ansari, T

2018-05-21

our aim was to use porcine vagina to create a vaginal matrix and test its cellular biocompatibility. vagina was harvested from pigs and de-cellularised (DC) using a combination of detergents (Triton x-100 and sodium deoxycholate) and enzymes (DNAse/RNAse). the presence of cellular material, collagen structural integrity and basement membrane proteins were assessed histologically. To address cytocompatibility, porcine adipose derived-mesenchymal stem cells (AD-MSC) were harvested from abdominal fat together with vaginal epithelial cells (VEC) and seeded onto the mucosal aspect of the vaginal scaffold. Both cells populations were seeded individually and assessed histologically at days 3 and 10. MAIN OUTCOMES/RESULTS: the combination of enzymes and detergents resulted in a totally acellular matrix with very low DNA amount (control= 97.5ng/μl ± 10.8 vs DC= 40.1 ng/μl ±0.33 p=0.02). The extra cellular matrix (ECM) showed retention of collagen fibres and elastin and a 50% retention in glycosaminoglycan content; (control= 1.18μg/mg ± 0.28 DC = 1.35μg/mg ± 0.1 p=0.03) and an intact basement membrane (positive for both laminin and collagen IV). Seeded scaffolds showed cell attachment with both AD-MSC and VEC at days 3 and 10. it is possible to generate an acellular porcine vaginal matrix capable of supporting cells to reconstruct the vagina for future pre-clinical testing, and holds promise for creating clinically relevant sized tissue for human application. Copyright © 2018. Published by Elsevier Inc.

Patch esophagoplasty using an in-body-tissue-engineered collagenous connective tissue membrane.

Science.gov (United States)

Okuyama, Hiroomi; Umeda, Satoshi; Takama, Yuichi; Terasawa, Takeshi; Nakayama, Yasuhide

2018-02-01

Although many approaches to esophageal replacement have been investigated, these efforts have thus far only met limited success. In-body-tissue-engineered connective tissue tubes have been reported to be effective as vascular replacement grafts. The aim of this study was to investigate the usefulness of an In-body-tissue-engineered collagenous connective tissue membrane, "Biosheet", as a novel esophageal scaffold in a beagle model. We prepared Biosheets by embedding specially designed molds into subcutaneous pouches in beagles. After 1-2months, the molds, which were filled with ingrown connective tissues, were harvested. Rectangular-shaped Biosheets (10×20mm) were then implanted to replace defects of the same size that had been created in the cervical esophagus of the beagle. An endoscopic evaluation was performed at 4 and 12weeks after implantation. The esophagus was harvested and subjected to a histological evaluation at 4 (n=2) and 12weeks (n=2) after implantation. The animal study protocols were approved by the National Cerebral and Cardiovascular Centre Research Institute Committee (No. 16048). The Biosheets showed sufficient strength and flexibility to replace the esophagus defect. All animals survived with full oral feeding during the study period. No anastomotic leakage was observed. An endoscopic study at 4 and 12weeks after implantation revealed that the anastomotic sites and the internal surface of the Biosheets were smooth, without stenosis. A histological analysis at 4weeks after implantation demonstrated that stratified squamous epithelium was regenerated on the internal surface of the Biosheets. A histological analysis at 12weeks after implantation showed the regeneration of muscle tissue in the implanted Biosheets. The long-term results of patch esophagoplasty using Biosheets showed regeneration of stratified squamous epithelium and muscular tissues in the implanted sheets. These results suggest that Biosheets may be useful as a novel esophageal

Poliomyelitis in MuLV-infected ICR-SCID mice after injection of basement membrane matrix contaminated with lactate dehydrogenase-elevating virus.

Science.gov (United States)

Carlson Scholz, Jodi A; Garg, Rohit; Compton, Susan R; Allore, Heather G; Zeiss, Caroline J; Uchio, Edward M

2011-10-01

The arterivirus lactate dehydrogenase-elevating virus (LDV) causes life-long viremia in mice. Although LDV infection generally does not cause disease, infected mice that are homozygous for the Fv1(n) allele are prone to develop poliomyelitis when immunosuppressed, a condition known as age-dependent poliomyelitis. The development of age-dependent poliomyelitis requires coinfection with endogenous murine leukemia virus. Even though LDV is a common contaminant of transplantable tumors, clinical signs of poliomyelitis after inadvertent exposure to LDV have not been described in recent literature. In addition, LDV-induced poliomyelitis has not been reported in SCID or ICR mice. Here we describe the occurrence of poliomyelitis in ICR-SCID mice resulting from injection of LDV-contaminated basement membrane matrix. After exposure to LDV, a subset of mice presented with clinical signs including paresis, which was associated with atrophy of the hindlimb musculature, and tachypnea; in addition, some mice died suddenly with or without premonitory signs. Mice presenting within the first 6 mo after infection had regions of spongiosis, neuronal necrosis and astrocytosis of the ventral spinal cord, and less commonly, brainstem. Axonal degeneration of ventral roots prevailed in more chronically infected mice. LDV was identified by RT-PCR in 12 of 15 mice with typical neuropathology; positive antiLDV immunolabeling was identified in all PCR-positive animals (n = 7) tested. Three of 8 mice with neuropathology but no clinical signs were LDV negative by RT-PCR. RT-PCR yielded murine leukemia virus in spinal cords of all mice tested, regardless of clinical presentation or neuropathology.

Anti-nucleosome antibodies complexed to nucleosomal antigens show anti-DNA reactivity and bind to rat glomerular basement membrane in vivo.

Science.gov (United States)

Kramers, C; Hylkema, M N; van Bruggen, M C; van de Lagemaat, R; Dijkman, H B; Assmann, K J; Smeenk, R J; Berden, J H

1994-01-01

Histones can mediate the binding of DNA and anti-DNA to the glomerular basement membrane (GBM). In ELISA histone/DNA/anti-DNA complexes are able to bind to heparan sulfate (HS), an intrinsic constituent of the GBM. We questioned whether histone containing immune complexes are able to bind to the GBM, and if so, whether the ligand in the GBM is HS. Monoclonal antibodies (mAbs) complexed to nucleosomal antigens and noncomplexed mAbs were isolated from culture supernatants of four IgG anti-nuclear mAbs. All noncomplexed mAbs showed strong anti-nucleosome reactivity in ELISA. One of them showed in addition anti-DNA reactivity in noncomplexed form. The other three mAbs only showed anti-DNA reactivity when they were complexed to nucleosomal antigens. After renal perfusion a fine granular binding of complexed mAbs to the glomerular capillary wall and activation of complement was observed in immunofluorescence, whereas noncomplexed mAbs did not bind. Immuno-electron microscopy showed binding of complexes to the whole width of the GBM. When HS in the GBM was removed by renal heparinase perfusion the binding of complexed mAb decreased, but did not disappear completely. We conclude that anti-nucleosome mAbs, which do not bind DNA, become DNA reactive once complexed to nucleosomal antigens. These complexed mAbs can bind to the GBM. The binding ligand in the GBM is partly, but not solely, HS. Binding to the GBM of immune complexes containing nucleosomal material might be an important event in the pathogenesis of lupus nephritis. Images PMID:8040312

Experiments on pollutant transport from soil into residential basements by pressure-driven airflow

International Nuclear Information System (INIS)

Nazaroff, W.W.; Lewis, S.R.; Doyle, S.M.; Moed, B.A.; Nero, A.V.

1987-01-01

At two residences in Portland, OR, they have investigated (1) the coupling between residential basements and the air in nearby soil and (2) the influence of basement depressurization on the migration of air in soil. With the basements depressurized 25-50 Pa relative to outdoor air, underpressures as great as 20-40% of those in the basement were observed at sampling points in the soil. Sulfur hexafluoride was injected into the soil near the houses and its concentration monitored in soil air and in the house over time, both with and without basement depressurization. Depressurization was seen to have a substantial effect on the migration of the tracer within the soil. For basement depressurizations of 25-50 Pa, effective transport velocities through the soil and into the houses were observed to exceed 1 m h -1 . Airborne 222 Rn concentration was monitored in the basement of one house during the 6-day investigation and was seen to increase substantially on each of the seven occasions that the house was depressurized. The techniques employed are applicable to the study of problems of excessive radon entry into buildings and the migration of toxic vapors from waste dumps and landfills

Increasing extracellular matrix collagen level and MMP activity induces cyst development in polycystic kidney disease.

Science.gov (United States)

Liu, Bin; Li, Chenghai; Liu, Zijuan; Dai, Zonghan; Tao, Yunxia

2012-09-11

Polycystic Kidney Disease (PKD) kidneys exhibit increased extracellular matrix (ECM) collagen expression and metalloproteinases (MMPs) activity. We investigated the role of these increases on cystic disease progression in PKD kidneys. We examined the role of type I collagen (collagen I) and membrane bound type 1 MMP (MT1-MMP) on cyst development using both in vitro 3 dimensional (3D) collagen gel culture and in vivo PCK rat model of PKD. We found that collagen concentration is critical in controlling the morphogenesis of MDCK cells cultured in 3D gels. MDCK cells did not form 3D structures at collagen I concentrations lower than 1 mg/ml but began forming tubules when the concentration reaches 1 mg/ml. Significantly, these cells began to form cyst when collagen I concentration reached to 1.2 mg/ml, and the ratios of cyst to tubule structures increased as the collagen I concentration increased. These cells exclusively formed cyst structures at a collagen I concentration of 1.8 mg/ml or higher. Overexpression of MT1-MMP in MDCK cells significantly induced cyst growth in 3D collagen gel culture. Conversely, inhibition of MMPs activity with doxycycline, a FDA approved pan-MMPs inhibitor, dramatically slowed cyst growth. More importantly, the treatment of PCK rats with doxycycline significantly decreased renal tubule cell proliferation and markedly inhibited the cystic disease progression. Our data suggest that increased collagen expression and MMP activity in PKD kidneys may induce cyst formation and expansion. Our findings also suggest that MMPs may serve as a therapeutic target for the treatment of human PKD.

Increasing extracellular matrix collagen level and MMP activity induces cyst development in polycystic kidney disease

Directory of Open Access Journals (Sweden)

Liu Bin

2012-09-01

Full Text Available Abstract Background Polycystic Kidney Disease (PKD kidneys exhibit increased extracellular matrix (ECM collagen expression and metalloproteinases (MMPs activity. We investigated the role of these increases on cystic disease progression in PKD kidneys. Methods We examined the role of type I collagen (collagen I and membrane bound type 1 MMP (MT1-MMP on cyst development using both in vitro 3 dimensional (3D collagen gel culture and in vivo PCK rat model of PKD. Results We found that collagen concentration is critical in controlling the morphogenesis of MDCK cells cultured in 3D gels. MDCK cells did not form 3D structures at collagen I concentrations lower than 1 mg/ml but began forming tubules when the concentration reaches 1 mg/ml. Significantly, these cells began to form cyst when collagen I concentration reached to 1.2 mg/ml, and the ratios of cyst to tubule structures increased as the collagen I concentration increased. These cells exclusively formed cyst structures at a collagen I concentration of 1.8 mg/ml or higher. Overexpression of MT1-MMP in MDCK cells significantly induced cyst growth in 3D collagen gel culture. Conversely, inhibition of MMPs activity with doxycycline, a FDA approved pan-MMPs inhibitor, dramatically slowed cyst growth. More importantly, the treatment of PCK rats with doxycycline significantly decreased renal tubule cell proliferation and markedly inhibited the cystic disease progression. Conclusions Our data suggest that increased collagen expression and MMP activity in PKD kidneys may induce cyst formation and expansion. Our findings also suggest that MMPs may serve as a therapeutic target for the treatment of human PKD.

A collagen-binding EGFR antibody fragment targeting tumors with a collagen-rich extracellular matrix

OpenAIRE

Hui Liang; Xiaoran Li; Bin Wang; Bing Chen; Yannan Zhao; Jie Sun; Yan Zhuang; Jiajia Shi; He Shen; Zhijun Zhang; Jianwu Dai

2016-01-01

Many tumors over-express collagen, which constitutes the physical scaffold of tumor microenvironment. Collagen has been considered to be a target for cancer therapy. The collagen-binding domain (CBD) is a short peptide, which could bind to collagen and achieve the sustained release of CBD-fused proteins in collagen scaffold. Here, a collagen-binding EGFR antibody fragment was designed and expressed for targeting the collagen-rich extracellular matrix in tumors. The antibody fragment (Fab) of ...

Demineralized Freeze-Dried Bovine Cortical Bone: Its Potential for Guided Bone Regeneration Membrane

Directory of Open Access Journals (Sweden)

David B. Kamadjaja

2017-01-01

Full Text Available Background. Bovine pericardium collagen membrane (BPCM had been widely used in guided bone regeneration (GBR whose manufacturing process usually required chemical cross-linking to prolong its biodegradation. However, cross-linking of collagen fibrils was associated with poorer tissue integration and delayed vascular invasion. Objective. This study evaluated the potential of bovine cortical bone collagen membrane for GBR by evaluating its antigenicity potential, cytotoxicity, immune and tissue response, and biodegradation behaviors. Material and Methods. Antigenicity potential of demineralized freeze-dried bovine cortical bone membrane (DFDBCBM was done with histology-based anticellularity evaluation, while cytotoxicity was analyzed using MTT Assay. Evaluation of immune response, tissue response, and biodegradation was done by randomly implanting DFDBCBM and BPCM in rat’s subcutaneous dorsum. Samples were collected at 2, 5, and 7 days and 7, 14, 21, and 28 days for biocompatibility and tissue response-biodegradation study, respectively. Result. DFDBCBM, histologically, showed no retained cells; however, it showed some level of in vitro cytotoxicity. In vivo study exhibited increased immune response to DFDBCBM in early healing phase; however, normal tissue response and degradation rate were observed up to 4 weeks after DFDBCBM implantation. Conclusion. Demineralized freeze-dried bovine cortical bone membrane showed potential for clinical application; however, it needs to be optimized in its biocompatibility to fulfill all requirements for GBR membrane.

Persistence of collagen type II-specific T-cell clones in the synovial membrane of a patient with rheumatoid arthritis

International Nuclear Information System (INIS)

Londei, M.; Savill, C.M.; Verhoef, A.; Brennan, F.; Leech, Z.A.; Feldmann, M.; Duance, V.; Maini, R.N.

1989-01-01

Rheumatoid arthritis is an autoimmune disease characterized by T-cell infiltration of the synovium of joints. Analysis of the phenotype and antigen specificity of the infiltrating cells may thus provide insight into the pathogenesis of rheumatoid arthritis. T cells were cloned with interleukin 2, a procedure that selects for in vivo-activated cells. All clones had the CD4 CDW29 phenotype. Their antigen specificity was tested by using a panel of candidate joint autoantigens. Four of 17 reacted against autologous blood mononuclear cells. Two clones proliferated in response to collagen type II. After 21 months, another set of clones was derived from synovial tissue of the same joint. One of eight clones tested showed a strong proliferative response against collagen type II. The uncloned synovial T cells of a third operation from another joint also responded to collagen type II. The persistence of collagen type II-specific T cells in active rheumatoid joints over a period of 3 years suggests that collagen type II could be one of the autoantigens involved in perpetuating the inflammatory process in rheumatoid arthritis

Discussion on the basement topography and its relation with the uranium mineralization in Xiangshan basin

International Nuclear Information System (INIS)

Long Qihua; Liu Qingcheng

2002-01-01

The depth of the basement and the relation between the basement relief shape and uranium mineralization are discussed by forward and inverse computation for large-scale gravity data in Xiangshan basin. The difference of basement topography result in the inhomogeneous distribution of uranium mineralization. The margin of the basement upheaval section and the variation place of basement topography are the favorable place for uranium mineralization. It's helpful to prospect deep and blind uranium deposit in Xiangshan basin

Proximal collagenous gastroenteritides:

DEFF Research Database (Denmark)

Nielsen, Ole Haagen; Riis, Lene Buhl; Danese, Silvio

2014-01-01

AIM: While collagenous colitis represents the most common form of the collagenous gastroenteritides, the collagenous entities affecting the proximal part of the gastrointestinal tract are much less recognized and possibly overlooked. The aim was to summarize the latest information through a syste...

Endocytic collagen degradation

DEFF Research Database (Denmark)

Madsen, Daniel H.; Jürgensen, Henrik J.; Ingvarsen, Signe Ziir

2012-01-01

it crucially important to understand both the collagen synthesis and turnover mechanisms in this condition. Here we show that the endocytic collagen receptor, uPARAP/Endo180, is a major determinant in governing the balance between collagen deposition and degradation. Cirrhotic human livers displayed a marked...... up-regulation of uPARAP/Endo180 in activated fibroblasts and hepatic stellate cells located close to the collagen deposits. In a hepatic stellate cell line, uPARAP/Endo180 was shown to be active in, and required for, the uptake and intracellular degradation of collagen. To evaluate the functional...... groups of mice clearly revealed a fibrosis protective role of uPARAP/Endo180. This effect appeared to directly reflect the activity of the collagen receptor, since no compensatory events were noted when comparing the mRNA expression profiles of the two groups of mice in an array system focused on matrix-degrading...

Examination of the Basement of Historic Buildings in Investment Activity

Directory of Open Access Journals (Sweden)

Ulybin Aleksey

2016-01-01

Full Text Available The process and methodology of the survey of basements rarely mentioned in the various construction rules and regulations. Basically describes the procedure of conducting a detailed survey of some of the individual elements. These surveys are fundamental in nature, include a large number of estimates and require significant financial and time costs. Usually the purpose of these surveys is to check the state of the building as a whole, it’s safe operation or before starting of reconstruction. In the process of selecting areas of investment activity such large-scale survey is not possible. Needed a quick and inexpensive method intended for decision about investment in a particular object. At the same time, the survey should cover all the elements of the basement significantly affect the cost of reconstruction of the basement associated with his penetration. The article presents the general conception of conducting a rapid survey. The described methods and technologies applicable to the examination for the purpose of making decisions about investments in reconstruction of a basement level rooms. The composition of the works and their sequence. A comparison of the advantages and disadvantages of different methods. The practical examples. Scheme of conducting a rapid survey of the basement. The article analyzes the materials used in the construction of historic buildings in St. Petersburg.

Stimulation of MMP-11 (stromelysin-3) expression in mouse fibroblasts by cytokines, collagen and co-culture with human breast cancer cell lines

International Nuclear Information System (INIS)

Selvey, Saxon; Haupt, Larisa M; Thompson, Erik W; Matthaei, Klaus I; Irving, Michael G; Griffiths, Lyn R

2004-01-01

Matrix metalloproteinases (MMPs) are central to degradation of the extracellular matrix and basement membrane during both normal and carcinogenic tissue remodeling. MT1-MMP (MMP-14) and stromelysin-3 (MMP-11) are two members of the MMP family of proteolytic enzymes that have been specifically implicated in breast cancer progression. Expressed in stromal fibroblasts adjacent to epithelial tumour cells, the mechanism of MT1-MMP and MMP-11 induction remains unknown. To investigate possible mechanisms of induction, we examined the effects of a number of plausible regulatory agents and treatments that may physiologically influence MMP expression during tumour progression. Thus NIH3T3 and primary mouse embryonic fibroblasts (MEFs) were: a) treated with the cytokines IL-1β, IL-2, IL-6, IL-8 and TGF-β for 3, 6, 12, 24, and 48 hours; b) grown on collagens I, IV and V; c) treated with fibronectin, con-A and matrigel; and d) co-cultured with a range of HBC (human breast cancer) cell lines of varied invasive and metastatic potential. Competitive quantitative RT-PCR indicated that MMP-11 expression was stimulated to a level greater than 100%, by 48 hour treatments of IL-1β, IL-2, TGF-β, fibronectin and collagen V. No other substantial changes in expression of MMP-11 or MT1-MMP in either tested fibroblast culture, under any treatment conditions, were observed. We have demonstrated significant MMP-11 stimulation in mouse fibroblasts using cytokines, matrix constituents and HBC cell lines, and also some inhibition of MT1-MMP. Our data suggest that the regulation of these genes in the complex stromal-epithelial interactions that occur in human breast carcinoma, is influenced by several mechanisms

Stimulation of MMP-11 (stromelysin-3 expression in mouse fibroblasts by cytokines, collagen and co-culture with human breast cancer cell lines

Directory of Open Access Journals (Sweden)

Matthaei Klaus I

2004-07-01

Full Text Available Abstract Background Matrix metalloproteinases (MMPs are central to degradation of the extracellular matrix and basement membrane during both normal and carcinogenic tissue remodeling. MT1-MMP (MMP-14 and stromelysin-3 (MMP-11 are two members of the MMP family of proteolytic enzymes that have been specifically implicated in breast cancer progression. Expressed in stromal fibroblasts adjacent to epithelial tumour cells, the mechanism of MT1-MMP and MMP-11 induction remains unknown. Methods To investigate possible mechanisms of induction, we examined the effects of a number of plausible regulatory agents and treatments that may physiologically influence MMP expression during tumour progression. Thus NIH3T3 and primary mouse embryonic fibroblasts (MEFs were: a treated with the cytokines IL-1β, IL-2, IL-6, IL-8 and TGF-β for 3, 6, 12, 24, and 48 hours; b grown on collagens I, IV and V; c treated with fibronectin, con-A and matrigel; and d co-cultured with a range of HBC (human breast cancer cell lines of varied invasive and metastatic potential. Results Competitive quantitative RT-PCR indicated that MMP-11 expression was stimulated to a level greater than 100%, by 48 hour treatments of IL-1β, IL-2, TGF-β, fibronectin and collagen V. No other substantial changes in expression of MMP-11 or MT1-MMP in either tested fibroblast culture, under any treatment conditions, were observed. Conclusion We have demonstrated significant MMP-11 stimulation in mouse fibroblasts using cytokines, matrix constituents and HBC cell lines, and also some inhibition of MT1-MMP. Our data suggest that the regulation of these genes in the complex stromal-epithelial interactions that occur in human breast carcinoma, is influenced by several mechanisms.

Investigation of the subsurface features of the basement complex of ...

African Journals Online (AJOL)

3D seismic reflection survey was recently carried out within the Zaria area of the basement complex of northern Nigeria, in order to investigate the complexity of the subsurface features within the basement. The geology of the survey area was characterized by gneisses and low grade meta-sedimentary rocks that form the ...

Human Bone Derived Collagen for the Development of an Artificial Corneal Endothelial Graft. In Vivo Results in a Rabbit Model.

Directory of Open Access Journals (Sweden)

Natalia Vázquez

Full Text Available Corneal keratoplasty (penetrating or lamellar using cadaveric human tissue, is nowadays the main treatment for corneal endotelial dysfunctions. However, there is a worldwide shortage of donor corneas available for transplantation and about 53% of the world's population have no access to corneal transplantation. Generating a complete cornea by tissue engineering is still a tough goal, but an endothelial lamellar graft might be an easier task. In this study, we developed a tissue engineered corneal endothelium by culturing human corneal endothelial cells on a human purified type I collagen membrane. Human corneal endothelial cells were cultured from corneal rims after corneal penetrating keratoplasty and type I collagen was isolated from remnant cancellous bone chips. Isolated type I collagen was analyzed by western blot, liquid chromatography -mass spectrometry and quantified using the exponentially modified protein abundance index. Later on, collagen solution was casted at room temperature obtaining an optically transparent and mechanically manageable membrane that supports the growth of human and rabbit corneal endothelial cells which expressed characteristic markers of corneal endothelium: zonula ocluddens-1 and Na+/K+ ATPase. To evaluate the therapeutic efficiency of our artificial endothelial grafts, human purified type I collagen membranes cultured with rabbit corneal endothelial cells were transplanted in New Zealand white rabbits that were kept under a minimal immunosuppression regimen. Transplanted corneas maintained transparency for as long as 6 weeks without obvious edema or immune rejection and maintaining the same endothelial markers that in a healthy cornea. In conclusion, it is possible to develop an artificial human corneal endothelial graft using remnant tissues that are not employed in transplant procedures. This artificial endothelial graft can restore the integrality of corneal endothelium in an experimental model of

RESEARCH ON REDUCING PREMATURITY RUPTURE OF MEMBRANE

OpenAIRE

Maria URSACHI (BOLOTA); Emil ANTON; Sorana Caterina ANTON

2016-01-01

The membranes surrounding the amniotic cavity are composed from amnion and chorion, tightly adherent layers which are composed of several cell types, including epithelial cells, trophoblasts cells and mesenchyme cells, embedded in a collagenous matrix. They retain amniotic fluid, secret substances into the amniotic fluid, as well as to the uterus and protect the fetus against upward infections from urogenital tract. Normally, the membranes it breaks during labor. Premature rupture of the amn...

Basement Surface Faulting and Topography for Savannah River Site and Vicinity

International Nuclear Information System (INIS)

Cumbest, R.J.

1998-01-01

This report integrates the data from more than 60 basement borings and over 100 miles of seismic reflection profiling acquired on the Savannah River Site to map the topography of the basement (unweathered rock) surface and faulting recorded on this surface

Physiological type I collagen organization induces the formation of a novel class of linear invadosomes

Science.gov (United States)

Juin, Amélie; Billottet, Clotilde; Moreau, Violaine; Destaing, Olivier; Albiges-Rizo, Corinne; Rosenbaum, Jean; Génot, Elisabeth; Saltel, Frédéric

2012-01-01

Invadosomes are F-actin structures capable of degrading the matrix through the activation of matrix metalloproteases. As fibrillar type I collagen promotes pro-matrix metalloproteinase 2 activation by membrane type 1 matrix metalloproteinase, we aimed at investigating the functional relationships between collagen I organization and invadosome induction. We found that fibrillar collagen I induced linear F-actin structures, distributed along the fibrils, on endothelial cells, macrophages, fibroblasts, and tumor cells. These structures share features with conventional invadosomes, as they express cortactin and N-WASP and accumulate the scaffold protein Tks5, which proved essential for their formation. On the basis of their ability to degrade extracellular matrix elements and their original architecture, we named these structures “linear invadosomes.” Interestingly, podosomes or invadopodia were replaced by linear invadosomes upon contact of the cells with fibrillar collagen I. However, linear invadosomes clearly differ from classical invadosomes, as they do not contain paxillin, vinculin, and β1/β3 integrins. Using knockout mouse embryonic fibroblasts and RGD peptide, we demonstrate that linear invadosome formation and activity are independent of β1 and β3 integrins. Finally, linear invadosomes also formed in a three-dimensional collagen matrix. This study demonstrates that fibrillar collagen I is the physiological inducer of a novel class of invadosomes. PMID:22114353

Collagen Quantification in Tissue Specimens.

Science.gov (United States)

Coentro, João Quintas; Capella-Monsonís, Héctor; Graceffa, Valeria; Wu, Zhuning; Mullen, Anne Maria; Raghunath, Michael; Zeugolis, Dimitrios I

2017-01-01

Collagen is the major extracellular protein in mammals. Accurate quantification of collagen is essential in the biomaterials (e.g., reproducible collagen scaffold fabrication), drug discovery (e.g., assessment of collagen in pathophysiologies, such as fibrosis), and tissue engineering (e.g., quantification of cell-synthesized collagen) fields. Although measuring hydroxyproline content is the most widely used method to quantify collagen in biological specimens, the process is very laborious. To this end, the Sircol™ Collagen Assay is widely used due to its inherent simplicity and convenience. However, this method leads to overestimation of collagen content due to the interaction of Sirius red with basic amino acids of non-collagenous proteins. Herein, we describe the addition of an ultrafiltration purification step in the process to accurately determine collagen content in tissues.

Release of antibiotics from collagen dressing.

Science.gov (United States)

Grzybowski, J; Antos-Bielska, M; Ołdak, E; Trafny, E A

1997-01-01

Our new collagen dressing has been developed recently. Three types (A, B, and C) of the dressing were prepared in this study. Each type contained bacitracin, neomycin or colistin. The antibiotic was input into: i. collagen sponge (CS)--type A, ii. layer of limited hydrophobicity (LLH)--type B, and iii. into both CS and LLH layers--type C. The final concentration of the antibiotic that resulted from the loading level was 2 mg/cm2 for the dressings of type A and B and 4 mg/cm2 for the dressing of type C. The antibiotics were then extracted from the pieces of dressings for two days through dialysis membrane. Susceptibility of 54 bacterial strains (S. aureus, P. aeruginosa, and Acinetobacter) isolated from burn wounds were tested to the three antibiotics used for preparation of the dressings. The results of the study evidenced that efficiency of released of antibiotics into the extracts depended on the kind of antibiotic and on the type of dressing. The concentration of the antibiotics proved to be much higher than MIC90 values of the bacterial isolates tested in respect to their susceptibility. The dressing containing mixture of the three antibiotics in two layers--CS and LLH is now considered as potentially effective for care of infected wounds. It may be useful for the treatment of infected wounds or for profilaxis of contaminated wounds, ensuring: i. sufficient antimicrobial activity in wound, and ii. optimal wound environment for the presence of collagenic biomaterial on the damaged tissue.

Anti-glomerular basement membrane (anti-GBM) disease accompanied by vasculitis that was not positive for antineutrophil cytoplasmic antibodies to myeloperoxidase and proteinase 3: a report of two cases and the incidence of anti-GBM disease at one institution.

Science.gov (United States)

Nakabayashi, Kimimasa; Fujioka, Yasunori; Arimura, Yoshihiro; Fukuoka, Toshihito; Marumo, Tomohumi; Umino, Michiru; Kamiya, Yasushi; Okai, Takahiro; Tsurumaki, Shigeru; Nagasawa, Toshihiko; Yamada, Akira

2011-08-01

Anti-glomerular basement membrane (anti-GBM) disease is thought to be distinct from vasculitis. In contrast, there have been several papers suggesting the presence of angiitis in cases that were positive for anti-GBM antibody (Ab), as well as for either myeloperoxidase (MPO)- or proteinase 3 (PR3)-anti-neutrophil cytoplasmic antibody (ANCA) (Group I). We experienced four patients who had anti-GBM Abs, but not MPO- and PR3-ANCA (Group II), and two of these patients were found to have vasculitis. Therefore, we performed an in-depth study on these two patients. The patients with anti-GBM disease were isolated from 578 cases whose renal tissues were examined, and they were categorized into two groups. We have already published the data about Group I. We then proceeded to study two vasculitic patients in Group II clinically, pathologically, and serologically. The anti-GBM Ab and ANCA levels were detected by enzyme-linked immunosorbent assays. Renal specimens were studied by routine staining as well as immunohistochemical investigations of CD31 and type IV collagen. The total number of patients with anti-GBM disease was 7 (7/578 = 1.2%), with 3 patients belonging to Group I and 4 patients belonging to Group II. Two patients in Group II were diagnosed to have vasculitis, but the remaining 2 patients did not. One vasculitic patient was complicated by pulmonary hemorrhage, while the other vasculitic patient displayed peripheral neuropathy as well as a small cavity lesion in the lung. The latter patient was found to be positive for perinuclear (p)-ANCA, but not for any other ANCA subsets. The renal pathology in the two vasculitic patients showed crescentic glomerulonephritis (CSGN) and immunoglobulin (Ig) G linear deposits along the glomerular capillary loops. The former patient showed fibrinoid angiitis in an afferent arteriole as well as peritubular capillaritis. The latter patient demonstrated peritubular capillaritis. These peritubular capillaritides were diagnosed by

Membranous glomerulonephritis in a child asymptomatic for hepatitis B virus. Concomitant seropositivity for HBsAG and anti-HBs.

Science.gov (United States)

Hirsch, H Z; Ainsworth, S K; DeBeukelaer, M; Brissie, R M; Hennigar, G R

1981-04-01

The presence of hepatitis B surface antigen (HBsAg) in association with immunoglobulins and complement components within the glomerular basement membranes of adults having chronic active hepatitis has been well documented. In addition, investigators in Poland have demonstrated HBsAg immune complexes in glomeruli of children who did not have clinical evidence of hepatitis. More recently, a single case of childhood membranous glomerulonephritis in an asymptomatic carrier of hepatitis B virus was cited by observers in Canada. Reported here is the deposition of HBsAg immune complexes in the glomerular basement membranes of a 13-year-old black boy who had membranous glomerulopathy but not clinical evidence of hepatitis. This may be the first reported case in the United States of HbsAg-associated membranous glomerulonephritis in a child asymptomatic for hepatitis B virus, and only the second such case in North America. However, unlike previous studies of childhood glomerulopathy in association with hepatitis B virus, this patient is seropositive for both HBsAg and anti-HBs (antibody for hepatitis B surface antigen). Similar "rare" serologic findings were found for the patient's eldest male sib.

Geology of the Pan-African basement Complex in Ube-Wulko area ...

African Journals Online (AJOL)

The Ube-Wulko area of southeast Akwanga falls within the Pan-African remobilized Basement Complex of northcentral Nigeria. It consists of intensely multi-deformed high grade polymetamorphic basement rocks, predominantly composed of migmatitic gneisses and schists and subordinate quartzites, marbles, and ...

Collagen Accumulation in Osteosarcoma Cells lacking GLT25D1 Collagen Galactosyltransferase.

Science.gov (United States)

Baumann, Stephan; Hennet, Thierry

2016-08-26

Collagen is post-translationally modified by prolyl and lysyl hydroxylation and subsequently by glycosylation of hydroxylysine. Despite the widespread occurrence of the glycan structure Glc(α1-2)Gal linked to hydroxylysine in animals, the functional significance of collagen glycosylation remains elusive. To address the role of glycosylation in collagen expression, folding, and secretion, we used the CRISPR/Cas9 system to inactivate the collagen galactosyltransferase GLT25D1 and GLT25D2 genes in osteosarcoma cells. Loss of GLT25D1 led to increased expression and intracellular accumulation of collagen type I, whereas loss of GLT25D2 had no effect on collagen secretion. Inactivation of the GLT25D1 gene resulted in a compensatory induction of GLT25D2 expression. Loss of GLT25D1 decreased collagen glycosylation by up to 60% but did not alter collagen folding and thermal stability. Whereas cells harboring individually inactivated GLT25D1 and GLT25D2 genes could be recovered and maintained in culture, cell clones with simultaneously inactive GLT25D1 and GLT25D2 genes could be not grown and studied, suggesting that a complete loss of collagen glycosylation impairs osteosarcoma cell proliferation and viability. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Estrogen and progesterone receptors have distinct roles in the establishment of the hyperplastic phenotype in PR-A transgenic mice

Energy Technology Data Exchange (ETDEWEB)

Simian, Marina; Bissell, Mina J.; Barcellos-Hoff, Mary Helen; Shyamala, Gopalan

2009-05-11

Expression of the A and B forms of progesterone receptor (PR) in an appropriate ratio is critical for mammary development. Mammary glands of PR-A transgenic mice, carrying an additional A form of PR as a transgene, exhibit morphological features associated with the development of mammary tumors. Our objective was to determine the roles of estrogen (E) and progesterone (P) in the genesis of mammary hyperplasias/preneoplasias in PR-A transgenics. We subjected PR-A mice to hormonal treatments and analyzed mammary glands for the presence of hyperplasias and used BrdU incorporation to measure proliferation. Quantitative image analysis was carried out to compare levels of latency-associated peptide and transforming growth factor beta 1 (TGF{beta}1) between PR-A and PR-B transgenics. Basement membrane disruption was examined by immunofluorescence and proteolytic activity by zymography. The hyperplastic phenotype of PR-A transgenics is inhibited by ovariectomy, and is reversed by treatment with E + P. Studies using the antiestrogen ICI 182,780 or antiprogestins RU486 or ZK 98,299 show that the increase in proliferation requires signaling through E/estrogen receptor alpha but is not sufficient to give rise to hyperplasias, whereas signaling through P/PR has little impact on proliferation but is essential for the manifestation of hyperplasias. Increased proliferation is correlated with decreased TGF{beta}1 activation in the PR-A transgenics. Analysis of basement membrane integrity showed loss of laminin-5, collagen III and collagen IV in mammary glands of PR-A mice, which is restored by ovariectomy. Examination of matrix metalloproteases (MMPs) showed that total levels of MMP-2 correlate with the steady-state levels of PR, and that areas of laminin-5 loss coincide with those of activation of MMP-2 in PR-A transgenics. Activation of MMP-2 is dependent on treatment with E and P in ovariectomized wild-type mice, but is achieved only by treatment with P in PR-A mice. These data

Co-effect of silk and amniotic membrane for tendon repair.

Science.gov (United States)

Seo, Young-Kwon; Kim, Jun-Hyung; Eo, Su-Rak

2016-08-01

The objective of the present study was to determine the feasibility and biocompatibility of a silk scaffold and a composite silk scaffold in terms of new tendon generation using a rabbit Achilles tendon model. The silk scaffold was constructed using a weaving machine, then soaked in a 1% collagen-hyaluronan (HA) solution and air-dried, whereas the composite silk scaffold was composed of a silk scaffold containing a lyophilized collagen-HA substrate. Tenocytes were cultured in vitro to compare cell populations in the two groups. The cellular densities on composite silk scaffolds were 40% higher on average than those on silk scaffolds in 30-day tenocyte cultures. The tendon scaffolds had implanted into Achilles tendon defects in 16 white New Zealand rabbits. Rabbits were randomly divided into the following three groups: group I, silk scaffold alone; group II, composite silk scaffold; and group III, composite silk scaffold wrapped by an amniotic membrane. Implants were harvested 2, 8, and 12 weeks post-implantation. Histological examinations were conducted using hematoxylin-eosin (H&E), Masson's trichrome, and by performing immunohistochemical staining for CD34. After 12 weeks, the three groups were distinguishable based on gross examination. The histological examination revealed more organized collagen fibrils in groups III, which showed a dense, parallel, linear organization of collagen bundles. CD34 staining revealed neoangiogenesis in groups III. The results of this research showed that collagen-HA substrates with amniotic membrane accelerate cellular migration and angiogenesis in neotendons.

development in a basement terrain

African Journals Online (AJOL)

2004-05-17

May 17, 2004 ... especially as a reconnaissance tool (de Jong et al., 1981; de. Rooy et al., 1986; ... ful naval radio transmitters in the very low frequency range. (15-25kHz). ... involved partial curve matching and computer iteration tech- niques. .... result predicted depth to fresh basement bedrock to be 46m whilst drilling ...

Depth of magnetic basement in Iran based on fractal spectral analysis of aeromagnetic data

Science.gov (United States)

Teknik, Vahid; Ghods, Abdolreza

2017-06-01

To estimate the shape of sedimentary basins, a critical parameter in hydrocarbon exploration, we calculated the depth of magnetic basement by applying a fractal spectral method to the aeromagnetic map of Iran. The depth of magnetic basement is a close proxy for the shape of sedimentary basins provided that igneous basement is strongly magnetized relative to the overlying sediments and there is no interbedding magnetic layer in the sediments. The shape of the power spectrum of magnetic anomalies is sensitive to the depth of magnetic basement, the thickness of the magnetic layer, the fractal parameter of magnetization and the size of the window used for the calculation of the power spectrum. Using a suite of synthetic tests, we have shown that the estimation of the depth of magnetic basement of up to 20 km is not very sensitive to the often unknown fractal parameter and thus the spectral method is a reliable tool to calculate the depth of magnetic basement. The depth of magnetic basement is in the range of 7-16 km in the Zagros, east Alborz, Tabas, Jazmurian and Makran regions, showing a close correlation with depths estimated from the maximum thickness of stratigraphic columns. We have also found new sedimentary basins in Bostan Abad, Bijar and south of Orumiyeh Lake. The significant depth of the magnetic basement in the Makran, Jazmurain depression, southeast Caspian Sea, Tabas, Great Kavir, south of Orumiyeh Lake, Bostan Abad and Bijar sedimentary basins makes them future prospects for hydrocarbon explorations. The depth of magnetic basement is strongly reduced over the Neyriz and Kermanshah Ophiolites, but it does not show any meaningful correlation with other outcrops of ophiolitic rocks in Iran.

Effects of 3D-Printed Polycaprolactone/?-Tricalcium Phosphate Membranes on Guided Bone Regeneration

OpenAIRE

Shim, Jin-Hyung; Won, Joo-Yun; Park, Jung-Hyung; Bae, Ji-Hyeon; Ahn, Geunseon; Kim, Chang-Hwan; Lim, Dong-Hyuk; Cho, Dong-Woo; Yun, Won-Soo; Bae, Eun-Bin; Jeong, Chang-Mo; Huh, Jung-Bo

2017-01-01

This study was conducted to compare 3D-printed polycaprolactone (PCL) and polycaprolactone/β-tricalcium phosphate (PCL/β-TCP) membranes with a conventional commercial collagen membrane in terms of their abilities to facilitate guided bone regeneration (GBR). Fabricated membranes were tested for dry and wet mechanical properties. Fibroblasts and preosteoblasts were seeded into the membranes and rates and patterns of proliferation were analyzed using a kit-8 assay and by scanning electron micro...

Quantitative N-linked Glycoproteomics of Myocardial Ischemia and Reperfusion Injury Reveals Early Remodeling in the Extracellular Environment

DEFF Research Database (Denmark)

Parker, Benjamin L; Palmisano, Giuseppe; Edwards, Alistair V G

2011-01-01

, while dimethyl labeling confirmed 46 of these and revealed an additional 62 significant changes. These were mainly from predicted extracellular matrix and basement membrane proteins that are implicated in cardiac remodeling. Analysis of N-glycans released from myocardial proteins suggest...... that the observed changes were not due to significant alterations in N-glycan structures. Altered proteins included the collagen-laminin-integrin complexes and collagen assembly enzymes, cadherins, mast cell proteases, proliferation-associated secreted protein acidic and rich in cysteine, and microfibril......Extracellular and cell surface proteins are generally modified with N-linked glycans and glycopeptide enrichment is an attractive tool to analyze these proteins. The role of N-linked glycoproteins in cardiovascular disease, particularly ischemia and reperfusion injury, is poorly understood...

A BMP responsive transcriptional region in the chicken type X collagen gene.

Science.gov (United States)

Volk, S W; Luvalle, P; Leask, T; Leboy, P S

1998-10-01

Bone morphogenetic proteins (BMPs) were originally identified by their ability to induce ectopic bone formation and have been shown to promote both chondrogenesis and chondrocyte hypertrophy. BMPs have recently been found to activate a membrane serine/threonine kinase signaling mechanism in a variety of cell types, but the downstream effectors of BMP signaling in chondrocyte differentiation remain unidentified. We have previously reported that BMP-2 markedly stimulates type X collagen expression in prehypertrophic chick sternal chondrocytes, and that type X collagen mRNA levels in chondrocytes cultured under serum-free (SF) conditions are elevated 3- to 5-fold within 24 h. To better define the molecular mechanisms of induction of chondrocyte hypertrophy by BMPs, we examined the effect of BMPs on type X collagen production by 15-day chick embryo sternal chondrocytes cultured under SF conditions in the presence or absence of 30 ng/ml BMP-2, BMP-4, or BMP-7. Two populations of chondrocytes were used: one representing resting cartilage isolated from the caudal third of the sterna and the second representing prehypertrophic cartilage from the cephalic third of the sterna. BMP-2, BMP-4, and BMP-7 all effectively promoted chondrocyte maturation of cephalic sternal chondrocytes as measured by high levels of alkaline phosphatase, diminished levels of type II collagen, and induction of the hypertrophic chondrocyte-specific marker, type X collagen. To test whether BMP control of type X collagen expression occurs at the transcriptional level, we utilized plasmid constructs containing the chicken collagen X promoter and 5' flanking regions fused to a reporter gene. Constructs were transiently transfected into sternal chondrocytes cultured under SF conditions in the presence or absence of 30 ng/ml BMP-2, BMP-4, or BMP-7. A 533 bp region located 2.4-2.9 kb upstream from the type X collagen transcriptional start site was both necessary and sufficient for strong BMP responsiveness

Differential regulation of collagen secretion by kinin receptors in cardiac fibroblast and myofibroblast

Energy Technology Data Exchange (ETDEWEB)

Catalán, Mabel; Smolic, Christian [Centro de estudios moleculares de la célula, Facultad de Ciencias Químicas y Farmacéuticas, Universidad de Chile (Chile); Contreras, Ariel [Instituto Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile (Chile); Ayala, Pedro; Olmedo, Ivonne; Copaja, Miguel; Boza, Pía; Vivar, Raúl; Avalos, Yennifer [Centro de estudios moleculares de la célula, Facultad de Ciencias Químicas y Farmacéuticas, Universidad de Chile (Chile); Lavandero, Sergio [Centro de estudios moleculares de la célula, Facultad de Ciencias Químicas y Farmacéuticas, Universidad de Chile (Chile); Instituto Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile (Chile); Department of Internal Medicine (Cardiology Division), University of Texas Southwestern Medical Center, Dallas, TX (United States); Velarde, Victoria [Departamento de Ciencias Fisiológicas, Facultad de Ciencias Biológicas, Pontificia Universidad Católica de Chile, Santiago (Chile); Díaz-Araya, Guillermo, E-mail: gadiaz@ciq.uchile.cl [Centro de estudios moleculares de la célula, Facultad de Ciencias Químicas y Farmacéuticas, Universidad de Chile (Chile)

2012-06-15

Kinins mediate their cellular effects through B1 (B1R) and B2 (B2R) receptors, and the activation of B2R reduces collagen synthesis in cardiac fibroblasts (CF). However, the question of whether B1R and/or B2R have a role in cardiac myofibroblasts remains unanswered. Methods: CF were isolated from neonate rats and myofibroblasts were generated by an 84 h treatment with TGF-β1 (CMF). B1R was evaluated by western blot, immunocytochemistry and radioligand assay; B2R, inducible nitric oxide synthase (iNOS), endothelial nitric oxide synthase (eNOS), and cyclooxygenases 1and 2 (COX-1, and COX-2) were evaluated by western blot; intracellular Ca{sup +2} levels were evaluated with Fluo-4AM; collagen secretion was measured in the culture media using the picrosirius red assay kit. Results: B2R, iNOS, COX-1 and low levels of B1R but not eNOS, were detected by western blot in CF. Also, B1R, B2R, and COX-2 but not iNOS, eNOS or COX-1, were detected by western blot in CMF. By immunocytochemistry, our results showed lower intracellular B1R levels in CF and higher B1R levels in CMF, mainly localized on the cell membrane. Additionally, we found B1R only in CMF cellular membrane through radioligand displacement assay. Bradykinin (BK) B2R agonist increased intracellular Ca{sup 2+} levels and reduced collagen secretion both in CF and CMF. These effects were blocked by HOE-140, and inhibited by L-NAME, 1400W and indomethacin. Des-Arg-kallidin (DAKD) B1R agonist did not increase intracellular Ca{sup 2+} levels in CF; however, after preincubation for 1 h with DAKD and re-stimulation with the same agonist, we found a low increase in intracellular Ca{sup 2+} levels. Finally, DAKD increased intracellular Ca{sup 2+} levels and decreased collagen secretion in CMF, being this effect blocked by the B1R antagonist des-Arg9-Leu8-kallidin and indomethacin, but not by L-NAME or 1400 W. Conclusion: B1R, B2R, iNOS and COX-1 were expressed differently between CF and CMF, and collagen secretion was

Differential regulation of collagen secretion by kinin receptors in cardiac fibroblast and myofibroblast

International Nuclear Information System (INIS)

Catalán, Mabel; Smolic, Christian; Contreras, Ariel; Ayala, Pedro; Olmedo, Ivonne; Copaja, Miguel; Boza, Pía; Vivar, Raúl; Avalos, Yennifer; Lavandero, Sergio; Velarde, Victoria; Díaz-Araya, Guillermo

2012-01-01

Kinins mediate their cellular effects through B1 (B1R) and B2 (B2R) receptors, and the activation of B2R reduces collagen synthesis in cardiac fibroblasts (CF). However, the question of whether B1R and/or B2R have a role in cardiac myofibroblasts remains unanswered. Methods: CF were isolated from neonate rats and myofibroblasts were generated by an 84 h treatment with TGF-β1 (CMF). B1R was evaluated by western blot, immunocytochemistry and radioligand assay; B2R, inducible nitric oxide synthase (iNOS), endothelial nitric oxide synthase (eNOS), and cyclooxygenases 1and 2 (COX-1, and COX-2) were evaluated by western blot; intracellular Ca +2 levels were evaluated with Fluo-4AM; collagen secretion was measured in the culture media using the picrosirius red assay kit. Results: B2R, iNOS, COX-1 and low levels of B1R but not eNOS, were detected by western blot in CF. Also, B1R, B2R, and COX-2 but not iNOS, eNOS or COX-1, were detected by western blot in CMF. By immunocytochemistry, our results showed lower intracellular B1R levels in CF and higher B1R levels in CMF, mainly localized on the cell membrane. Additionally, we found B1R only in CMF cellular membrane through radioligand displacement assay. Bradykinin (BK) B2R agonist increased intracellular Ca 2+ levels and reduced collagen secretion both in CF and CMF. These effects were blocked by HOE-140, and inhibited by L-NAME, 1400W and indomethacin. Des-Arg-kallidin (DAKD) B1R agonist did not increase intracellular Ca 2+ levels in CF; however, after preincubation for 1 h with DAKD and re-stimulation with the same agonist, we found a low increase in intracellular Ca 2+ levels. Finally, DAKD increased intracellular Ca 2+ levels and decreased collagen secretion in CMF, being this effect blocked by the B1R antagonist des-Arg9-Leu8-kallidin and indomethacin, but not by L-NAME or 1400 W. Conclusion: B1R, B2R, iNOS and COX-1 were expressed differently between CF and CMF, and collagen secretion was regulated differentially by

A clinical stydy on the effectiveness of slow - resorbing collagen membrane barrier therapy to guide regeneration in mandibular class II furcations in human

Directory of Open Access Journals (Sweden)

Abolfazli N

1998-09-01

Full Text Available The present clinical trial was designed to evaluate the regenerative potential of periodontal tissues in degree II"nfurcation defects at mandibular molars of human using a slow-resorbing collagen membrane and a surgical treatment"ntechnique based on the principles of guided tissue regeneration."nThe patient sampleinclude 8 subjects who had periodontal lessions in right and left mandibular molars regions, including moderate to advance periodonal destruction within the radicular area. Following a baseline examination including recording the clinical measurements (PD, Al, HC, F.G.M , the furcation- involved molars were randomly assigned in each patient to either a test or a control treatment procedure. Included the evevation of mucoperiosteal flaps, recording measurement from the cemento enamel junction (C.E.J directly coronal to the furcation area to the alveolar crest and to the base of the defect-Horizontal furcation measurements were also made using a William's probe, finally a collagen membrrane placed on the involved area to cover the entrance of the furcation and adjucent root surfaces as well as a portion of the alveolar bone apical to the crest. The flaps were repositioned and secured with interdental sutures. A procedure identical to the one used at the test teeth was Performed at the control teeth region with the exception of the placement of the collagen membrance. Following surgery all patients were placed on a plaque control regimen. All Patients received normal postsurgical care and at 6 month post-surgery were scheduled for re-entry surgery. Before re-entry surgery all clinical parameters recorded again. The re-entry mucoperiosteal flaps were designed to expose the furcation area for measurements, as describedabove. There was clinical improvement in all measurements made in both the test and control patients (especially in test group over the 6 month period. The horizontal and vertical furcation measurements did yield a

Contribution To The Geology Of Basement Rocks In The South Western Desert Of Egypt

International Nuclear Information System (INIS)

Sadek, M.F.; Khyamy, A.A.

2003-01-01

Three major Precambrian basement inliers are exposed in the South Western Desert of Egypt between Long. 29 degree E and the River Nile within the Uweinat-Bir Safsaf-Aswan E-W uplift system. These are Bir Safsaf, Gabal EI-Asr and Gabal Umm Shaghir areas. Smaller outcrops include Gabal EI-Gara El-Hamra and Gabal El-Gara EI-Soda, Gabal Siri, GabaI EI-Fantas and Aswan-Kalabsha area as well as the scattered outcrops around Darb El-Arbain road. Band ratios 5/7, 5/1, 4 of Landsat TM images were applied to delineate the borders, the lithologic units and structural features of low relief basement outcrops within the surrounding flat lying sedimentary rocks and sand plains. These basement rocks comprise ortho gneisses (assumed by many authors as related to old continent pre Pan-African rocks), G 1 tonalite-granodiorite, and G2 monzogranite-alkali feldspar granite intruded by variable dykes. The boundaries between the basement exposures and the sedimentary rocks are marked by nonconformity surfaces or sets of faults. Both basement and sedimentary rocks are intruded by Mesozoic syenite-G3 granites, rhyolite, trachytic plugs and Upper Cretaceous to Tertiary basalts. The basement exposures are structurally controlled by major E- W fault systems. Their vertical uplifting is overprinted by folding the overlying sedimentary rocks. This study revealed that, the different basement exposures in the SE of the Western Desert of Egypt are similar in appearance and field relations to the Pan-African basement rocks extending towards the east of the River Nile and exposed everywhere in the Eastern Desert of Egypt

Rapid Upregulation of Orai1 Abundance in the Plasma Membrane of Platelets Following Activation with Thrombin and Collagen Related Peptide

Directory of Open Access Journals (Sweden)

Guilai Liu

2015-11-01

Full Text Available Background: Blood platelets accomplish primary hemostasis following vascular injury and contribute to the orchestration of occlusive vascular disease. Platelets are activated by an increase of cytosolic Ca2+-activity ([Ca2+]i, which is accomplished by Ca2+-release from intracellular stores and subsequent store operated Ca2+ entry (SOCE through Ca2+ release activated Ca2+ channel moiety Orai1. Powerful activators of platelets include thrombin and collagen related peptide (CRP, which are in part effective by activation of small G- protein Rac1. The present study explored the influence of thrombin and CRP on Orai1 protein abundance and cytosolic Ca2+-activity ([Ca2+]i in platelets drawn from wild type mice. Methods: Orai1 protein surface abundance was quantified utilizing CF™488A conjugated antibodies, and [Ca2+]i was determined with Fluo3-fluorescence. Results: In resting platelets, Orai1 protein abundance and [Ca2+]i were low. Thrombin (0.02 U/ml and CRP (5ug/ml within 2 min increased [Ca2+]i and Orai1 protein abundance at the platelet surface. [Ca2+]i was further increased by Ca2+ ionophore ionomycin (1 µM and by store depletion with the sarcoendoplasmatic Ca2+ ATPase inhibitor thapsigargin (1 µM. However, Orai1 protein abundance at the platelet surface was not significantly affected by ionomycin and only slightly increased by thapsigargin. The effect of thrombin and CRP on Orai1 abundance and [Ca2+]i was significantly blunted by Rac1 inhibitor NSC23766 (50 µM. Conclusion: The increase of [Ca2+]i following stimulation of platelets with thrombin and collagen related peptide is potentiated by ultrarapid Rac1 sensitive translocation of Orai1 into the cell membrane.

Disruption of the ECM33 Gene in Candida albicans Prevents Biofilm Formation, Engineered Human Oral Mucosa Tissue Damage and Gingival Cell Necrosis/Apoptosis

Directory of Open Access Journals (Sweden)

Mahmoud Rouabhia

2012-01-01

Full Text Available In this study we demonstrated that ΔCaecm33 double mutant showed reduced biofilm formation and causes less damage to gingival mucosa tissues. This was confirmed by the reduced level of necrotic cells and Bax/Bcl2 gene expression as apoptotic markers. In contrast, parental and Caecm33 mutant strains decreased basement membrane protein production (laminin 5 and type IV collagen. We thus propose that ECM33 gene/protein represents a novel target for the prevention and treatment of infections caused by Candida.

Collagen-binding peptidoglycans inhibit MMP mediated collagen degradation and reduce dermal scarring.

Directory of Open Access Journals (Sweden)

Kate Stuart

Full Text Available Scarring of the skin is a large unmet clinical problem that is of high patient concern and impact. Wound healing is complex and involves numerous pathways that are highly orchestrated, leaving the skin sealed, but with abnormal organization and composition of tissue components, namely collagen and proteoglycans, that are then remodeled over time. To improve healing and reduce or eliminate scarring, more rapid restoration of healthy tissue composition and organization offers a unique approach for development of new therapeutics. A synthetic collagen-binding peptidoglycan has been developed that inhibits matrix metalloproteinase-1 and 13 (MMP-1 and MMP-13 mediated collagen degradation. We investigated the synthetic peptidoglycan in a rat incisional model in which a single dose was delivered in a hyaluronic acid (HA vehicle at the time of surgery prior to wound closure. The peptidoglycan treatment resulted in a significant reduction in scar tissue at 21 days as measured by histology and visual analysis. Improved collagen architecture of the treated wounds was demonstrated by increased tensile strength and transmission electron microscopy (TEM analysis of collagen fibril diameters compared to untreated and HA controls. The peptidoglycan's mechanism of action includes masking existing collagen and inhibiting MMP-mediated collagen degradation while modulating collagen organization. The peptidoglycan can be synthesized at low cost with unique design control, and together with demonstrated preclinical efficacy in reducing scarring, warrants further investigation for dermal wound healing.

Possible options for reducing occupational dose from the TMI-2 basement

International Nuclear Information System (INIS)

Munson, L.F.; Harty, R.

1985-11-01

The major sources of exposure in the basement include the enclosed stairwell/elevator shaft structure, water and sludge in the elevator shaft, cast concrete walls, concrete floor slab, water and sludge on the floor, and activity in the paint and loose surface contamination. The sources were identified using data obtained by the utility from water processing, water and solid samples, remote video inspections and radiation monitoring with a robot, and strings of thermoluminescent dosimeters lowered from upper elevations. The area dose rates in the basement range from approximately 4 R/hr (in the NE quadrant) to over 1100 R/hr (near the enclosed stairwell/elevator shaft structure). It is estimated that the basement contains between 11,000 and 21,000 curies of 137 Cs. Specific decontamination and cleanup techniques are discussed. These techniques include flushing with water, high-pressure water blasting, leaching, scabbling and chemical cleaning. The applicability of these techniques to the major sources of radiation are discussed, and possible approaches and work sequences for basement cleanup are given

Membrane-reinforced three-dimensional electrospun silk fibroin scaffolds for bone tissue engineering

International Nuclear Information System (INIS)

Yang, Sung Yeun; Hwang, Tae Heon; Ryu, WonHyoung; Che, Lihua; Oh, Jin Soo; Ha, Yoon

2015-01-01

Electrospun silk fibroin (SF) scaffolds have drawn much attention because of their resemblance to natural tissue architecture such as extracellular matrix, and the biocompatibility of SF as a candidate material to replace collagen. However, electrospun scaffolds lack the physical integrity of bone tissue scaffolds, which require resistance to mechanical loadings. In this work, we propose membrane-reinforced electrospun SF scaffolds by a serial process of electrospinning and freeze-drying of SF solutions in two different solvents: formic acid and water, respectively. After wet electrospinning followed by replacement of methanol with water, SF nanofibers dispersed in water were mixed with aqueous SF solution. Freeze-drying of the mixed solution resulted in 3D membrane-connected SF nanofibrous scaffolds (SF scaffolds) with a thickness of a few centimeters. We demonstrated that the SF concentration of aqueous SF solution controlled the degree of membrane reinforcement between nanofibers. It was also shown that both increase in degree of membrane reinforcement and inclusion of hydroxyapatite (HAP) nanoparticles resulted in higher resistance to compressive loadings of the SF scaffolds. Culture of human osteoblasts on collagen, SF, and SF-HAP scaffolds showed that both SF and SF-HAP scaffolds had biocompatibility and cell proliferation superior to that of the collagen scaffolds. SF-HAP scaffolds with and without BMP-2 were used for in vivo studies for 4 and 8 weeks, and they showed enhanced bone tissue formation in rat calvarial defect models. (paper)

A collagen-binding EGFR antibody fragment targeting tumors with a collagen-rich extracellular matrix.

Science.gov (United States)

Liang, Hui; Li, Xiaoran; Wang, Bin; Chen, Bing; Zhao, Yannan; Sun, Jie; Zhuang, Yan; Shi, Jiajia; Shen, He; Zhang, Zhijun; Dai, Jianwu

2016-02-17

Many tumors over-express collagen, which constitutes the physical scaffold of tumor microenvironment. Collagen has been considered to be a target for cancer therapy. The collagen-binding domain (CBD) is a short peptide, which could bind to collagen and achieve the sustained release of CBD-fused proteins in collagen scaffold. Here, a collagen-binding EGFR antibody fragment was designed and expressed for targeting the collagen-rich extracellular matrix in tumors. The antibody fragment (Fab) of cetuximab was fused with CBD (CBD-Fab) and expressed in Pichia pastoris. CBD-Fab maintained antigen binding and anti-tumor activity of cetuximab and obtained a collagen-binding ability in vitro. The results also showed CBD-Fab was mainly enriched in tumors and had longer retention time in tumors in A431 s.c. xenografts. Furthermore, CBD-Fab showed a similar therapeutic efficacy as cetuximab in A431 xenografts. Although CBD-Fab hasn't showed better therapeutic effects than cetuximab, its smaller molecular and special target may be applicable as antibody-drug conjugates (ADC) or immunotoxins.

Alpha1 and Alpha2 Integrins Mediate Invasive Activity of Mouse Mammary Carcinoma Cells through Regulation of Stromelysin-1 Expression

Energy Technology Data Exchange (ETDEWEB)

Lochter, Andre; Navre, Marc; Werb, Zena; Bissell, Mina J

1998-06-29

Tumor cell invasion relies on cell migration and extracellular matrix proteolysis. We investigated the contribution of different integrins to the invasive activity of mouse mammary carcinoma cells. Antibodies against integrin subunits {alpha}6 and {beta}1, but not against {alpha}1 and {alpha}2, inhibited cell locomotion on a reconstituted basement membrane in two-dimensional cell migration assays, whereas antibodies against {beta}1, but not against a6 or {alpha}2, interfered with cell adhesion to basement membrane constituents. Blocking antibodies against {alpha}1 integrins impaired only cell adhesion to type IV collagen. Antibodies against {alpha}1, {alpha}2, {alpha}6, and {beta}1, but not {alpha}5, integrin subunits reduced invasion of a reconstituted basement membrane. Integrins {alpha}1 and {alpha}2, which contributed only marginally to motility and adhesion, regulated proteinase production. Antibodies against {alpha}1 and {alpha}2, but not {alpha}6 and {beta}1, integrin subunits inhibited both transcription and protein expression of the matrix metalloproteinase stromelysin-1. Inhibition of tumor cell invasion by antibodies against {alpha}1 and {alpha}2 was reversed by addition of recombinant stromelysin-1. In contrast, stromelysin-1 could not rescue invasion inhibited by anti-{alpha}6 antibodies. Our data indicate that {alpha}1 and {alpha}2 integrins confer invasive behavior by regulating stromelysin-1 expression, whereas {alpha}6 integrins regulate cell motility. These results provide new insights into the specific functions of integrins during tumor cell invasion.

Detection of gelatinolytic activity in developing basement membranes of the mouse embryo head by combining sensitive in situ zymography with immunolabeling.

Science.gov (United States)

Gkantidis, Nikolaos; Katsaros, Christos; Chiquet, Matthias

2012-10-01

Genetic evidence indicates that the major gelatinases MMP-2 and MMP-9 are involved in mammalian craniofacial development. Since these matrix metalloproteinases are secreted as proenzymes that require activation, their tissue distribution does not necessarily reflect the sites of enzymatic activity. Information regarding the spatial and temporal expression of gelatinolytic activity in the head of the mammalian embryo is sparse. Sensitive in situ zymography with dye-quenched gelatin (DQ-gelatin) has been introduced recently; gelatinolytic activity results in a local increase in fluorescence. Using frontal sections of wild-type mouse embryo heads from embryonic day 14.5-15.5, we optimized and validated a simple double-labeling in situ technique for combining DQ-gelatin zymography with immunofluorescence staining. MMP inhibitors were tested to confirm the specificity of the reaction in situ, and results were compared to standard SDS-gel zymography of tissue extracts. Double-labeling was used to show the spatial relationship in situ between gelatinolytic activity and immunostaining for gelatinases MMP-2 and MMP-9, collagenase 3 (MMP-13) and MT1-MMP (MMP-14), a major activator of pro-gelatinases. Strong gelatinolytic activity, which partially overlapped with MMP proteins, was confirmed for Meckel's cartilage and developing mandibular bone. In addition, we combined in situ zymography with immunostaining for extracellular matrix proteins that are potential gelatinase substrates. Interestingly, gelatinolytic activity colocalized precisely with laminin-positive basement membranes at specific sites around growing epithelia in the developing mouse head, such as the ducts of salivary glands or the epithelial fold between tongue and lower jaw region. Thus, this sensitive method allows to associate, with high spatial resolution, gelatinolytic activity with epithelial morphogenesis in the embryo.

3D depth-to-basement and density contrast estimates using gravity and borehole data

Science.gov (United States)

Barbosa, V. C.; Martins, C. M.; Silva, J. B.

2009-05-01

We present a gravity inversion method for simultaneously estimating the 3D basement relief of a sedimentary basin and the parameters defining the parabolic decay of the density contrast with depth in a sedimentary pack assuming the prior knowledge about the basement depth at a few points. The sedimentary pack is approximated by a grid of 3D vertical prisms juxtaposed in both horizontal directions, x and y, of a right-handed coordinate system. The prisms' thicknesses represent the depths to the basement and are the parameters to be estimated from the gravity data. To produce stable depth-to-basement estimates we impose smoothness on the basement depths through minimization of the spatial derivatives of the parameters in the x and y directions. To estimate the parameters defining the parabolic decay of the density contrast with depth we mapped a functional containing prior information about the basement depths at a few points. We apply our method to synthetic data from a simulated complex 3D basement relief with two sedimentary sections having distinct parabolic laws describing the density contrast variation with depth. Our method retrieves the true parameters of the parabolic law of density contrast decay with depth and produces good estimates of the basement relief if the number and the distribution of boreholes are sufficient. We also applied our method to real gravity data from the onshore and part of the shallow offshore Almada Basin, on Brazil's northeastern coast. The estimated 3D Almada's basement shows geologic structures that cannot be easily inferred just from the inspection of the gravity anomaly. The estimated Almada relief presents steep borders evidencing the presence of gravity faults. Also, we note the existence of three terraces separating two local subbasins. These geologic features are consistent with Almada's geodynamic origin (the Mesozoic breakup of Gondwana and the opening of the South Atlantic Ocean) and they are important in understanding

Effects of ionizing radiation on glycerolated amniotic membranes as a substract for cultured human epithelium

International Nuclear Information System (INIS)

Paggiaro, Andre Oliveira

2011-01-01

The amniotic membrane (AM) is a biomaterial with biological properties that are beneficial to tissue repair. It has been used as a temporary coverage to threat burns and chronic wounds. Recently, it has been served as a substrate for keratinocytes culture to construct a living skin equivalent. However, MA is a biological material, and its transplantation could cause infectious disease for receptors. So, it must be preserved and sterilized before clinical use. The aim of this study was to evaluate the radiation effects on glycerol-preserved MA, considering its compatibility to support human keratinocytes culture. Four MA were stored in high concentrations of glycerol (> 85%) and half of them were radio sterilized with a dose of 25 kGy. Then, we established two groups: nonirradiated MA (MA-ni) and irradiated MA (MA-i). Both groups was deepithelialized by a standardized protocol and was investigated morphologically, immunohistochemical and ultrastructural. Subsequently human keratinocytes were cultivated immersed and in air-liquid interface on denuded surface of MA-i and MA-ni. The results were compared at 14 and 21 days of culture by light and electron microscopy. After epithelial denudation, analyses demonstrated the continuity of the basement membrane in MA-ni group, whereas in the irradiated group, there was no indication of the basement membrane’s presence on the surface of MA. The cell cultures showed that in the non-irradiated group, there was growth of a multi-layered and differentiated epithelium, with a stratum corneum’s formation in air-liquid interface. In the irradiated group, the epithelium had only two or three layer, little cell differentiation, with the same results immersed or air-liquid interface system. Glycerol-preserved MA was biocompatible with the growth of a cultivated epithelium, showing its potential as a skin substitute. Irradiation at 25 kGy cause structural damage to the tissue, making changes in basement membrane, that facilitates

Insight into podocyte differentiation from the study of human genetic disease: nail-patella syndrome and transcriptional regulation in podocytes.

Science.gov (United States)

Morello, Roy; Lee, Brendan

2002-05-01

In recent years, our understanding of the molecular basis of kidney development has benefited from the study of rare genetic diseases affecting renal function. This has especially been the case with the differentiation of the highly specialized podocyte in the pathogenesis of human disorders and mouse phenotypes affecting the renal filtration barrier. This filtration barrier represents the end product of a complex series of signaling events that produce a tripartite structure consisting of interdigitating podocyte foot processes with intervening slit diaphragms, the glomerular basement membrane, and the fenestrated endothelial cell. Dysregulation of unique cytoskeletal and extracellular matrix proteins in genetic forms of nephrotic syndrome has shown how specific structural proteins contribute to podocyte function and differentiation. However, much less is known about the transcriptional determinants that both specify and maintain this differentiated cell. Our studies of a skeletal malformation syndrome, nail-patella syndrome, have shown how the LIM homeodomain transcription factor, Lmx1b, contributes to transcriptional regulation of glomerular basement membrane collagen expression by podocytes. Moreover, they raise intriguing questions about more global transcriptional regulation of podocyte morphogenesis.

Mapping magnetic lineaments and subsurface basement beneath ...

Indian Academy of Sciences (India)

65

studied the basement structures beneath parts of the Lower Benue Trough (LBT). Anudu et .... order vertical derivatives can be calculated respectively using the relations below: 145. ( ) ... minerals as in the case of the FVD-RTP-TMI (Figure 6).

Reactor-building-basement radionuclide and source distribution studies. Volume 3

International Nuclear Information System (INIS)

Cox, T.E.; Horan, J.T.; Worku, G.

1983-06-01

The Three Mile Island Unit 2 (TMI-2) Reactor Building basement has been sampled several times since August 1979. This report compiles the analytical results and sample history for the liquid and solid samples obtained to date. In addition, basement radiation levels were also obtained using thermoluminescent dosimeters (TLDs). The data obtained will provide information to support ongoing mass balance and source term studies and will aid in characterizing the 282-ft elevation for decontamination planning and dose reduction

Disorganized collagen scaffold interferes with fibroblast mediated deposition of organized extracellular matrix in vitro.

Science.gov (United States)

Saeidi, Nima; Guo, Xiaoqing; Hutcheon, Audrey E K; Sander, Edward A; Bale, Shyam Sundar; Melotti, Suzanna A; Zieske, James D; Trinkaus-Randall, Vickery; Ruberti, Jeffrey W

2012-10-01

Many tissue engineering applications require the remodeling of a degradable scaffold either in vitro or in situ. Although inefficient remodeling or failure to fully remodel the temporary matrix can result in a poor clinical outcome, very few investigations have examined in detail, the interaction of regenerative cells with temporary scaffoldings. In a recent series of investigations, randomly oriented collagen gels were directly implanted into human corneal pockets and followed for 24 months. The resulting remodeling response exhibited a high degree of variability which likely reflects differing regenerative/synthetic capacity across patients. Given this variability, we hypothesize that a disorganized, degradable provisional scaffold could be disruptive to a uniform, organized reconstruction of stromal matrix. In this investigation, two established corneal stroma tissue engineering culture systems (collagen scaffold-based and scaffold-free) were compared to determine if the presence of the disorganized collagen gel influenced matrix production and organizational control exerted by primary human corneal fibroblast cells (PHCFCs). PHCFCs were cultured on thin disorganized reconstituted collagen substrate (RCS--five donors: average age 34.4) or on a bare polycarbonate membrane (five donors: average age 32.4 controls). The organization and morphology of the two culture systems were compared over the long-term at 4, 8, and 11/12 weeks. Construct thickness and extracellular matrix organization/alignment was tracked optically with bright field and differential interference contrast (DIC) microscopy. The details of cell/matrix morphology and cell/matrix interaction were examined with standard transmission, cuprolinic blue and quick-freeze/deep-etch electron microscopy. Both the scaffold-free and the collagen-based scaffold cultures produced organized arrays of collagen fibrils. However, at all time points, the amount of organized cell-derived matrix in the scaffold

The osteoinductive effect of chitosan-collagen composites around pure titanium implant surfaces in rats.

Science.gov (United States)

Kung, S; Devlin, H; Fu, E; Ho, K-Y; Liang, S-Y; Hsieh, Y-D

2011-02-01

The enhancing effects of chitosan on activation of platelets and differentiation of osteoprogenitor cells have been demonstrated in vitro. The purpose of this study was to evaluate the in vivo osteoinductive effect of chitosan-collagen composites around pure titanium implant surfaces. Chitosan-collagen composites containing chitosan of different molecular weights (450 and 750 kDa) were wrapped onto titanium implants and embedded into the subcutaneous area on the back of 15 Sprague-Dawley rats. The control consisted of implants wrapped with plain collagen type I membranes. Implants and surrounding tissues were retrieved 6 wks after surgery and identified by Alizarin red and Alcian blue whole mount staining. The newly formed structures in the test groups were further analyzed by Toluidine blue and Masson-Goldner trichrome staining, and immunohistochemical staining with osteopontin and alkaline phosphotase. The bone formation parameters of the new bone in the two test groups were measured and compared. New bone formed ectopically in both chitosan-collagen groups, whereas no bone induction occurred in the negative control group. These newly formed bone-like structures were further confirmed by immunohistochemical staining. Comparison of bone parameters of the newly induced bone revealed no statistically significant differences between the 450 and 750 kDa chitosan-collagen groups. Our results demonstrated that chitosan-collagen composites might induce in vivo new bone formation around pure titanium implant surfaces. Different molecular weights of chitosan did not show significantly different effects on the osteoinductive potential of the test materials. © 2010 John Wiley & Sons A/S.

Influence of modified polyester on the material properties of collagen-based biocomposites and in vitro evaluation of cytocompatibility

Energy Technology Data Exchange (ETDEWEB)

Wu, Chin-San, E-mail: t50008@cc.kyu.edu.tw

2015-03-01

The cytocompatibility of composite materials collagen (Col)/poly(hydroxyalkanoate) (PHA) and collagen/maleic anhydride-grafted PHA (PHA-g-MA) was investigated in this study. Col was homogeneously dispersed in the PHA-g-MA matrix as a result of condensation reactions. Mechanical characterisation indicated that the improved adhesion between Col and PHA-g-MA enhanced the tensile strength of the composite compared with that of PHA/Col. PHA-g-MA/Col composites were also more water-resistant than PHA/Col composites. Collagen and cell proliferation analysis indicated that PHA and PHA-g-MA and their composites were biocompatible with respect to FB proliferation. Cell-cycle and apoptosis assays by FBs on the PHA series composite samples were not affected by DNA content related to damage, i.e. rapid apoptosis/necrosis was not observed, demonstrating the potential of PHA/Col or PHA-g-MA/Col membranes for biomedical material applications. - Highlights: • Composites were prepared using polyester and collagen to explore their cytocompatibility. • The mechanical properties of the composite were significantly enhanced by the use of grafted polyester and collagen. • The polyester and collagen composites facilitated excellent cell viability and collagen production. • The cell cycle was not affected by DNA content related to damage, and it did not lead to rapid apoptosis or necrosis of the cells by the composite.

Influence of modified polyester on the material properties of collagen-based biocomposites and in vitro evaluation of cytocompatibility

International Nuclear Information System (INIS)

Wu, Chin-San

2015-01-01

The cytocompatibility of composite materials collagen (Col)/poly(hydroxyalkanoate) (PHA) and collagen/maleic anhydride-grafted PHA (PHA-g-MA) was investigated in this study. Col was homogeneously dispersed in the PHA-g-MA matrix as a result of condensation reactions. Mechanical characterisation indicated that the improved adhesion between Col and PHA-g-MA enhanced the tensile strength of the composite compared with that of PHA/Col. PHA-g-MA/Col composites were also more water-resistant than PHA/Col composites. Collagen and cell proliferation analysis indicated that PHA and PHA-g-MA and their composites were biocompatible with respect to FB proliferation. Cell-cycle and apoptosis assays by FBs on the PHA series composite samples were not affected by DNA content related to damage, i.e. rapid apoptosis/necrosis was not observed, demonstrating the potential of PHA/Col or PHA-g-MA/Col membranes for biomedical material applications. - Highlights: • Composites were prepared using polyester and collagen to explore their cytocompatibility. • The mechanical properties of the composite were significantly enhanced by the use of grafted polyester and collagen. • The polyester and collagen composites facilitated excellent cell viability and collagen production. • The cell cycle was not affected by DNA content related to damage, and it did not lead to rapid apoptosis or necrosis of the cells by the composite

Collagen turnover after tibial fractures

DEFF Research Database (Denmark)

Joerring, S; Krogsgaard, M; Wilbek, H

1994-01-01

Collagen turnover after tibial fractures was examined in 16 patients with fracture of the tibial diaphysis and in 8 patients with fracture in the tibial condyle area by measuring sequential changes in serological markers of turnover of types I and III collagen for up to 26 weeks after fracture....... The markers were the carboxy-terminal extension peptide of type I procollagen (PICP), the amino-terminal extension peptide of type III procollagen (PIIINP), and the pyridinoline cross-linked carboxy-terminal telopeptide of type I collagen (ICTP). The latter is a new serum marker of degradation of type I...... collagen. A group comparison showed characteristic sequential changes in the turnover of types I and III collagen in fractures of the tibial diaphysis and tibial condyles. The turnover of type III collagen reached a maximum after 2 weeks in both groups. The synthesis of type I collagen reached a maximum...

Enhanced stabilization of collagen by furfural.

Science.gov (United States)

Lakra, Rachita; Kiran, Manikantan Syamala; Usha, Ramamoorthy; Mohan, Ranganathan; Sundaresan, Raja; Korrapati, Purna Sai

2014-04-01

Furfural (2-furancarboxaldehyde), a product derived from plant pentosans, has been investigated for its interaction with collagen. Introduction of furfural during fibril formation enhanced the thermal and mechanical stability of collagen. Collagen films treated with furfural exhibited higher denaturation temperature (Td) (pFurfural and furfural treated collagen films did not have any cytotoxic effect. Rheological characterization showed an increase in shear stress and shear viscosity with increasing shear rate for treated collagen. Circular dichroism (CD) studies indicated that the furfural did not have any impact on triple helical structure of collagen. Scanning electron microscopy (SEM) of furfural treated collagen exhibited small sized porous structure in comparison with untreated collagen. Thus this study provides an alternate ecologically safe crosslinking agent for improving the stability of collagen for biomedical and industrial applications. Copyright © 2014 Elsevier B.V. All rights reserved.

Relationship of Basal laminar deposit and membranous debris to the clinical presentation of early age-related macular degeneration.

Science.gov (United States)

Sarks, Shirley; Cherepanoff, Svetlana; Killingsworth, Murray; Sarks, John

2007-03-01

To correlate basal laminar deposit (BLamD) and membranous debris, including basal linear deposit (BLinD), with the evolution of early age-related macular degeneration (AMD). A clinicopathologic collection of 132 eyes with a continuous layer of BLamD was reviewed. The thickness and type of BLamD and the sites of membranous debris deposition were correlated with the clinical progression of the disease. Two types of BLamD, termed early and late, were identified based on light microscopic appearance by using the picro-Mallory stain. The progressive accumulation of late type BLamD correlated well with increasing BLamD thickness, advancing RPE degeneration, poorer vision, increasing age, and clinically evident pigment changes. Membranous debris initially accumulated diffusely as BLinD, most eyes with BLinD and early BLamD remaining funduscopically normal. However, membranous debris also formed focal collections as basal mounds internal to the RPE basement membrane and as soft drusen external to the basement membrane. Eyes in which membranous debris remained confined to basal mounds belonged to older patients with poorer vision, whereas patients with soft drusen were younger and had better vision. The presence of BLinD and early BLamD define threshold AMD, which manifests clinically as a normal fundus. Although late BLamD correlates most closely with clinical pigment abnormalities, it is the quantity and sites of membranous debris accumulation that appear to determine whether the disease develops pigment changes only or follows the alternative pathway of soft drusen formation with its attendant greater risk of choroidal neovascularization (CNV).

Modeling radon entry into houses with basements: Model description and verification

International Nuclear Information System (INIS)

Revzan, K.L.; Fisk, W.J.; Gadgil, A.J.

1991-01-01

We model radon entry into basements using a previously developed three-dimensional steady-state finite difference model that has been modified in the following ways: first, cylindrical coordinates are used to take advantage of the symmetry of the problem in the horizontal plant; second, the configuration of the basement has been made more realistic by incorporating the concrete footer; third, a quadratic relationship between the pressure and flow in the L-shaped gap between slab, footer, and wall has been employed; fourth, the natural convection of the soil gas which follows from the heating of the basement in winter has been taken into account. The temperature field in the soil is determined from the equation of energy conservation, using the basement, surface, and deep-soil temperatures as boundary conditions. The pressure field is determined from Darcy's law and the equation of mass conservation (continuity), assuming that there is no flow across any boundary except the soil surface (atmospheric pressure) and the opening in the basement shell (fixed pressure). After the pressure and temperatures field have been obtained the velocity field is found from Darcy's law. Finally, the radon concentration field is found from the equation of mass-transport. The convective radon entry rate through the opening or openings is then calculated. In this paper we describe the modified model, compare the predicted radon entry rates with and without the consideration of thermal convection, and compare the predicted rates with determined from data from 7 houses in the Spokane River valley of Washington and Idaho. Although the predicted rate is much lower than the mean of the rates determined from measurements, errors in the measurement of soil permeability and variations in the permeability of the area immediately under the basement slab, which has a significant influence on the pressure field, can account for the range of entry rates inferred from the data. 25 refs., 8 figs

Histological Evaluation of Decellularized Skeletal Muscle Tissue Using Two Different Decellularization Agents

Directory of Open Access Journals (Sweden)

Hana Hrebíková

2017-02-01

Full Text Available The aim of the present study was to determine effect of two decellularized agents, sodium dodecyl sulphate (SDS and Triton X-100, to the skeletal muscle tissue. Final scaffold was evaluated by several histological techniques to analyse preservation of essential structures including collagen and elastic fibres, basement membranes, glycosaminoglycans and also to confirm elimination of nuclear and cytoplasmic components which are redundant in effectively prepared decellularized scaffolds. Comparison of tissue scaffolds processed with different detergents proved that SDS is superior to Triton X-100 as it can effectively decellularize muscle tissue.

The effect of gamma-ray and ethylene oxide sterilization on collagen-based wound-repair materials

International Nuclear Information System (INIS)

Gorham, S.D.; Scott, R.

1993-01-01

The effects of 2.5, 5.0, 7.5 and 10 Mrad gamma-irradiation and ethylene oxide sterilization on collagen-coated vicryl mesh, on collagen film and vicryl mesh were investigated. No cytotoxic effects were observed towards either L929 cells or human fibroblasts with any of the treatments, even at the highest doses of irradiation. Although the rate of biodegradation and tissue reaction towards the test materials appeared to be relatively unaffected by ethylene oxide, the rate of breakdown in the lumbar muscles of laboratory rats was considerably increased by irradiation. Dose-dependent irradiation damage to the vicryl mesh was confirmed in vitro using viscometry and by an increase in the rate of hydrolysis in phosphate-buffered saline (PBS) at 37 o C. Tensiometric studies on irradiated collagen films showed a dose-dependent reduction in both the break load and elongation before breaking. A small reduction in the break load and stretch to breaking was also observed following treatment with ethylene oxide. The findings presented favour ethylene oxide as a method of sterilization of both the composite membrane and its individual components. However, gamma-irradiation produced no toxic effects or adverse tissue reaction under the conditions described, and is more convenient to use. Hence, provided that the efficacy of the membrane would not be compromised by the higher rate of degradation, it could also be considered for sterilization of the material. (author)

High resolution imaging of collagen organisation and synthesis using a versatile collagen specific probe

NARCIS (Netherlands)

Boerboom, R.A.; Krahn - Nash, K.; Megens, R.T.A.; Zandvoort, van M.; Merkx, M.; Bouten, C.V.C.

2007-01-01

Collagen is the protein primarily responsible for the load-bearing properties of tissues and collagen architecture is one of the main determinants of the mechanical properties of tissues. Visualisation of changes in collagen three-dimensional structure is essential in order to improve our

Characteristics of the crystalline basement beneath the Ordos Basin: Constraint from aeromagnetic data

Directory of Open Access Journals (Sweden)

Zhentao Wang

2015-05-01

Full Text Available Aeromagnetic anomaly zonation of the Ordos Basin and adjacent areas was obtained by processing high-precision and large-scale aeromagnetic anomalies with an approach of reduction to the pole upward continuation. Comparative study on aeromagnetic and seismic tomography suggests that aeromagnetic anomalies in this area are influenced by both the magnetic property of the rock and the burial depth of the Precambrian crystalline basement. Basement depth might be the fundamental control factor for aeromagnetic anomalies because the positive and negative anomalies on the reduction to the pole-upward-continuation anomaly maps roughly coincide with the uplifts and depressions of the crystalline basement in the basin. The results, together with the latest understanding of basement faults, SHRIMP U-Pb zircon dating of metamorphic rock and granite, drilling data, detrital zircon ages, and gravity data interpretation, suggest that the Ordos block is not an entirety of Archean.

Changes in collagenous tissue microstructures and distributions of cathepsin L in body wall of autolytic sea cucumber (Stichopus japonicus).

Science.gov (United States)

Liu, Yu-Xin; Zhou, Da-Yong; Ma, Dong-Dong; Liu, Yan-Fei; Li, Dong-Mei; Dong, Xiu-Ping; Tan, Ming-Qian; Du, Ming; Zhu, Bei-Wei

2016-12-01

The autolysis of sea cucumber (Stichopus japonicus) was induced by ultraviolet (UV) irradiation, and the changes of microstructures of collagenous tissues and distributions of cathepsin L were investigated using histological and histochemical techniques. Intact collagen fibers in fresh S. japonicus dermis were disaggregated into collagen fibrils after UV stimuli. Cathepsin L was identified inside the surface of vacuoles in the fresh S. japonicus dermis cells. After the UV stimuli, the membranes of vacuoles and cells were fused together, and cathepsin L was released from cells and diffused into tissues. The density of cathepsin L was positively correlated with the speed and degree of autolysis in different layers of body wall. Our results revealed that lysosomal cathepsin L was released from cells in response to UV stimuli, which contacts and degrades the extracellular substrates such as collagen fibers, and thus participates in the autolysis of S. japonicus. Copyright © 2016 Elsevier Ltd. All rights reserved.

Prostate-specific membrane antigen (PSMA)-mediated laminin proteolysis generates a pro-angiogenic peptide

Czech Academy of Sciences Publication Activity Database

Conway, R. E.; Rojas, C.; Alt, J.; Nováková, Zora; Richardson, S. M.; Rodrick, T. C.; Fuentes, J. L.; Richardson, N. H.; Attalla, J.; Stewart, S.; Fahmy, B.; Bařinka, Cyril; Ghosh, M.; Shapiro, L. H.; Slusher, B. S.

2016-01-01

Roč. 19, č. 4 (2016), s. 487-500 ISSN 0969-6970 R&D Projects: GA ČR GAP301/12/1513; GA MŠk(CZ) ED1.1.00/02.0109 Institutional support: RVO:86652036 Keywords : TUMOR-ASSOCIATED NEOVASCULATURE * BASEMENT-MEMBRANE * DISTINCT ANTITUMOR PROPERTIES Subject RIV: FA - Cardiovascular Diseases incl. Cardiotharic Surgery Impact factor: 5.253, year: 2016

Dynamic culture of a thermosensitive collagen hydrogel as an extracellular matrix improves the construction of tissue-engineered peripheral nerve.

Science.gov (United States)

Huang, Lanfeng; Li, Rui; Liu, Wanguo; Dai, Jin; Du, Zhenwu; Wang, Xiaonan; Ma, Jianchao; Zhao, Jinsong

2014-07-15

Tissue engineering technologies offer new treatment strategies for the repair of peripheral nerve injury, but cell loss between seeding and adhesion to the scaffold remains inevitable. A thermosensitive collagen hydrogel was used as an extracellular matrix in this study and combined with bone marrow mesenchymal stem cells to construct tissue-engineered peripheral nerve composites in vitro. Dynamic culture was performed at an oscillating frequency of 0.5 Hz and 35° swing angle above and below the horizontal plane. The results demonstrated that bone marrow mesenchymal stem cells formed membrane-like structures around the poly-L-lactic acid scaffolds and exhibited regular alignment on the composite surface. Collagen was used to fill in the pores, and seeded cells adhered onto the poly-L-lactic acid fibers. The DNA content of the bone marrow mesenchymal stem cells was higher in the composites constructed with a thermosensitive collagen hydrogel compared with that in collagen I scaffold controls. The cellular DNA content was also higher in the thermosensitive collagen hydrogel composites constructed with the thermosensitive collagen hydrogel in dynamic culture than that in static culture. These results indicate that tissue-engineered composites formed with thermosensitive collagen hydrogel in dynamic culture can maintain larger numbers of seeded cells by avoiding cell loss during the initial adhesion stage. Moreover, seeded cells were distributed throughout the material.

Routes towards Novel Collagen-Like Biomaterials

Directory of Open Access Journals (Sweden)

Adrian V. Golser

2018-04-01

Full Text Available Collagen plays a major role in providing mechanical support within the extracellular matrix and thus has long been used for various biomedical purposes. Exemplary, it is able to replace damaged tissues without causing adverse reactions in the receiving patient. Today’s collagen grafts mostly are made of decellularized and otherwise processed animal tissue and therefore carry the risk of unwanted side effects and limited mechanical strength, which makes them unsuitable for some applications e.g., within tissue engineering. In order to improve collagen-based biomaterials, recent advances have been made to process soluble collagen through nature-inspired silk-like spinning processes and to overcome the difficulties in providing adequate amounts of source material by manufacturing collagen-like proteins through biotechnological methods and peptide synthesis. Since these methods also open up possibilities to incorporate additional functional domains into the collagen, we discuss one of the best-performing collagen-like type of proteins, which already have additional functional domains in the natural blueprint, the marine mussel byssus collagens, providing inspiration for novel biomaterials based on collagen-silk hybrid proteins.

PHAGOCYTOSIS AND REMODELING OF COLLAGEN MATRICES

OpenAIRE

Abraham, Leah C.; Dice, J Fred.; Lee, Kyongbum; Kaplan, David L.

2007-01-01

The biodegradation of collagen and the deposition of new collagen-based extracellular matrices are of central importance in tissue remodeling and function. Similarly, for collagen-based biomaterials used in tissue engineering, the degradation of collagen scaffolds with accompanying cellular infiltration and generation of new extracellular matrix is critical for integration of in vitro grown tissues in vivo. In earlier studies we observed significant impact of collagen structure on primary lun...

3D collagen fibrillar microstructure guides pancreatic cancer cell phenotype and serves as a critical design parameter for phenotypic models of EMT.

Directory of Open Access Journals (Sweden)

T J Puls

Full Text Available Pancreatic cancer, one of the deadliest cancers, is characterized by high rates of metastasis and intense desmoplasia, both of which are associated with changes in fibrillar type I collagen composition and microstructure. Epithelial to mesenchymal transition (EMT, a critical step of metastasis, also involves a change in extracellular matrix (ECM context as cells detach from basement membrane (BM and engage interstitial matrix (IM. The objective of this work was to develop and apply an in-vitro three-dimensional (3D tumor-ECM model to define how ECM composition and biophysical properties modulate pancreatic cancer EMT. Three established pancreatic ductal adenocarcinoma (PDAC lines were embedded within 3D matrices prepared with type I collagen Oligomer (IM at various fibril densities to control matrix stiffness or Oligomer and Matrigel combined at various ratios while maintaining constant matrix stiffness. Evaluation of cell morphology and protein expression at both the cellular- and population-levels revealed a spectrum of matrix-driven EMT phenotypes that were dependent on ECM composition and architecture as well as initial PDAC phenotype. In general, exposure to fibrillar IM was sufficient to drive EMT, with cells displaying spindle-shaped morphology and mesenchymal markers, and non-fibrillar BM promoted more epithelial behavior. When cultured within low density Oligomer, only a subpopulation of epithelial BxPC-3 cells displayed EMT while mesenchymal MiaPaCa-2 cells displayed more uniform spindle-shaped morphologies and mesenchymal marker expression. Interestingly, as IM fibril density increased, associated changes in spatial constraints and matrix stiffness resulted in all PDAC lines growing as tight clusters; however mesenchymal marker expression was maintained. Collectively, the comparison of these results to other in-vitro tumor models highlights the role of IM fibril microstructure in guiding EMT heterogeneity and showcases the potential

Collagen XII myopathy with rectus femoris atrophy and collagen XII retention in fibroblasts

DEFF Research Database (Denmark)

Witting, Nanna; Krag, Thomas; Werlauff, Ulla

2018-01-01

INTRODUCTION: Mutation in the collagen XII gene (COL12A1) was recently reported to induce Bethlem myopathy. We describe a family affected by collagen XII-related myopathy in 3 generations. METHODS: Systematic interview, clinical examination, skin biopsies, and MRI of muscle were used. RESULTS...... affection and abnormal collagen XII retention in fibroblasts. MRI disclosed a selective wasting of the rectus femoris muscle. DISCUSSION: COL12A1 mutations should be considered in patients with a mild Bethlem phenotype who present with selective wasting of the rectus femoris, absence of the outside......-in phenomenon on MRI, and abnormal collagen XII retention in fibroblasts. Muscle Nerve, 2018....

A novel method to immobilize collagen on polypropylene film as substrate for hepatocyte culture

International Nuclear Information System (INIS)

Zhang Yong; Wang Wenjie; Feng Qingling; Cui Fuzhai; Xu Yingxin

2006-01-01

To improve the biocompatibility of polypropylene membrane with hepatocytes, film was firstly allowed to generate peroxide groups by ammonium peroxydisulfate (APS) thermal decomposition and was then carboxylated by free radical grafting polymerization using acrylic acid as the monomer and ferrous(II) ions as redox initiator. Collagen was then covalently immobilized through amide bonds between the residue amine groups and carboxylic ones in the presence of water-soluble carbodiimide. The film was characterized by Fourier transformed infrared spectroscopy in attenuated total reflection mode (ATR-FTIR), X-photoelectron spectroscopy (XPS) and confocal laser scanning microscopy (CLSM), etc. The experimental results show that there exist hydroperoxide groups after APS aqueous thermal decomposition, which are subsequently converted into the polymeric free radicals initiating the vinyl monomers to subject grafting polymerization. The maximum collagen immobilization content is about 10.5 μg/cm 2 by ninhydrin assay. Hepatocytes cultured on collagen immobilized film show stable phenotype and normal liver-specific functions more than 20 days as observed by scanning electronic microscopy (SEM) and biochemical parameters determination

A novel method to immobilize collagen on polypropylene film as substrate for hepatocyte culture

Energy Technology Data Exchange (ETDEWEB)

Zhang Yong [Biomaterials Laboratory, Department of Material Science and Engineering, Tsinghua University, Beijing 100084 (China); Wang Wenjie [Biomaterials Laboratory, Department of Material Science and Engineering, Tsinghua University, Beijing 100084 (China); Feng Qingling [Biomaterials Laboratory, Department of Material Science and Engineering, Tsinghua University, Beijing 100084 (China)]. E-mail: biomater@mail.tsinghua.edu.cn; Cui Fuzhai [Biomaterials Laboratory, Department of Material Science and Engineering, Tsinghua University, Beijing 100084 (China); Xu Yingxin [Institute of General Surgery, Chinese PLA General Hospital, Beijing 100853 (China)

2006-05-15

To improve the biocompatibility of polypropylene membrane with hepatocytes, film was firstly allowed to generate peroxide groups by ammonium peroxydisulfate (APS) thermal decomposition and was then carboxylated by free radical grafting polymerization using acrylic acid as the monomer and ferrous(II) ions as redox initiator. Collagen was then covalently immobilized through amide bonds between the residue amine groups and carboxylic ones in the presence of water-soluble carbodiimide. The film was characterized by Fourier transformed infrared spectroscopy in attenuated total reflection mode (ATR-FTIR), X-photoelectron spectroscopy (XPS) and confocal laser scanning microscopy (CLSM), etc. The experimental results show that there exist hydroperoxide groups after APS aqueous thermal decomposition, which are subsequently converted into the polymeric free radicals initiating the vinyl monomers to subject grafting polymerization. The maximum collagen immobilization content is about 10.5 {mu}g/cm{sup 2} by ninhydrin assay. Hepatocytes cultured on collagen immobilized film show stable phenotype and normal liver-specific functions more than 20 days as observed by scanning electronic microscopy (SEM) and biochemical parameters determination.

Rare mutations and potentially damaging missense variants in genes encoding fibrillar collagens and proteins involved in their production are candidates for risk for preterm premature rupture of membranes.

Science.gov (United States)

Modi, Bhavi P; Teves, Maria E; Pearson, Laurel N; Parikh, Hardik I; Chaemsaithong, Piya; Sheth, Nihar U; York, Timothy P; Romero, Roberto; Strauss, Jerome F

2017-01-01

Preterm premature rupture of membranes (PPROM) is the leading identifiable cause of preterm birth with ~ 40% of preterm births being associated with PPROM and occurs in 1% - 2% of all pregnancies. We hypothesized that multiple rare variants in fetal genes involved in extracellular matrix synthesis would associate with PPROM, based on the assumption that impaired elaboration of matrix proteins would reduce fetal membrane tensile strength, predisposing to unscheduled rupture. We performed whole exome sequencing (WES) on neonatal DNA derived from pregnancies complicated by PPROM (49 cases) and healthy term deliveries (20 controls) to identify candidate mutations/variants. Genotyping for selected variants from the WES study was carried out on an additional 188 PPROM cases and 175 controls. All mothers were self-reported African Americans, and a panel of ancestry informative markers was used to control for genetic ancestry in all genetic association tests. In support of the primary hypothesis, a statistically significant genetic burden (all samples combined, SKAT-O p-value = 0.0225) of damaging/potentially damaging rare variants was identified in the genes of interest-fibrillar collagen genes, which contribute to fetal membrane strength and integrity. These findings suggest that the fetal contribution to PPROM is polygenic, and driven by an increased burden of rare variants that may also contribute to the disparities in rates of preterm birth among African Americans.

A 48 kDa collagen-binding phosphoprotein isolated from bovine aortic endothelial cells interacts with the collagenous domain, but not the globular domain, of collagen type IV.

Science.gov (United States)

Yannariello-Brown, J; Madri, J A

1990-01-15

We have identified collagen-binding proteins in detergent extracts of metabolically labelled bovine aortic endothelial cells (BAEC) by collagen type IV-Sepharose affinity chromatography. The major collagen type IV-binding protein identified by SDS/PAGE had a molecular mass of 48 kDa, which we term the 'collagen-binding 48 kDa protein' (CB48). The pI of CB48 was 8.0-8.3 in a two-dimensional gel system, running non-equilibrium pH gel electrophoresis in the first dimension and SDS/PAGE in the second dimension. Under these conditions CB48 separated into two major (a and b) and one minor isoform (c); a was the most basic of the three isoforms. Two-dimensional chymotryptic peptide maps derived from each individual isoform were virtually identical. The charge differences between the isoforms were due in part to differential H3(32)PO4 incorporation by the protein. CB48 bound to intact collagen type IV and the collagenous region of collagen type IV, but not to the globular NC1 domain. Cell-surface labelling and indirect immunofluorescence experiments localized the bulk of CB48 intracellularly in the endoplasmic reticulum Golgi region, with a minor population of molecules on the cell surface. A specific rabbit polyclonal anti-CB48 serum did not inhibit the attachment or spreading of BAEC to collagen type IV in an 'in vitro' adhesion assay, suggesting that the cell-surface population of CB48 is not involved in BAEC adhesion. We conclude that CB48 is a collagen-binding phosphoprotein that interacts with the collagenous domain of collagen type IV and may be involved in intracellular transport of collagen molecules.